Annual Review of Immunology Volume 24 2006

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Contents

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Contents

Annual Review of Immunology Volume 24, 2006

Annu. Rev. Immunol. 2006.24:1-31. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:1-31. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Boardpage

The author in his ofﬁce holding an enlargement of the structure of the MHC class II/HA 306-318 complex, from Stern et al. (71).

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The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger Department of Molecular and Cellular Biology, Harvard University, Cambridge, Massachusetts 02138; email: [email protected]

Annu. Rev. Immunol. 2006. 24:1–31 First published online as a Review in Advance on November 28, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090703 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0001$20.00

Key Words bacterial cell walls, penicillin, histocompatibility proteins, MHC, HLA, NK cells

Abstract After starting out to become a physician, by a series of accidents I found myself at NIH in 1951 during its most productive growth phase. At age 26, I had a fully funded, independent laboratory and did not know what to work on. With advice from colleagues, I initiated a study of how penicillin kills bacteria. Twenty years later, my lab had outlined the structure and biosynthesis of the peptidoglycan of bacterial cell walls and had discovered that penicillin inhibited the terminal step in its biosynthesis catalyzed by transpeptidases. I then switched ﬁelds, moving to Harvard in 1968 and beginning the study of human HLA proteins. Twenty-ﬁve years later, the last half of which was spent in a stimulating collaboration with the late Don Wiley, our labs had isolated, crystallized, and elucidated the three-dimensional structures of these molecules and shown that their principal function was to present peptides to the immune system in initiating an immune response. More recently, the laboratory has focused on natural killer cells and their roles in peripheral blood and in the pregnant uterine decidua. It has been a wonderful scientiﬁc journey.

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EARLIEST MEMORIES 1925–1942

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I was born in 1925, the ﬁrst of three brothers whose grandparents had all emigrated in the 1890s from Eastern Europe: Belarus, Ukraine, and Lithuania. My grandparents met in the great melting pot of immigrants, New York City. I am not absolutely certain of the origin of the paternal grandfather who gave me my surname. The best memories in our family say it was Czernowitz, which is now in the southwest corner of Ukraine but was then part of the Austro-Hungarian Empire; at the time of his naturalization, he renounced allegiance to Franz Josef I, Emperor of Austria and Pontiﬁcal King of Hungary. He was ﬁrst employed as a carpenter in the growing city, but by the time I was born he was the relatively prosperous owner of a construction company. He built the building at 3320 Ditmars Blvd. in Astoria, Queens, a poor working-class area, as a graduation gift to his only son, my father, who was a dentist. My father never sent a patient a bill or asked for payment, maybe in gratitude for his family’s good fortune. I can remember patients returning years later to pay their debts. My earliest memory is of my father’s ofﬁce and waiting room on the second ﬂoor, which are still there. Behind the waiting room was our family apartment. On the ground ﬂoor were a drug store and tailor shop, both sources of rental income for our family. On the third ﬂoor was a small apartment for a couple who cared for the property and cleaned my father’s ofﬁce. By the time I started school, we had moved to an apartment in a row of two-story buildings across the street from the school I ﬁrst attended, quite close to the Steinway piano factory. We soon moved to another rental in a two-family house in Flushing. Then my grandfather built two adjacent homes for himself and my parents and their growing family. On many evenings, my father, my grandfather, and his two sons-in-law gathered to play pinochle and smoke cigars. There were no television sets then; I suppose that today families gather similarly to watch TV. I attended

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P.S. 32 and then Bayside High School. In sixth grade I was moved ahead a year, and in high school I graduated second in my class by a slim margin. I was surprised but extremely pleased when I was admitted to Harvard, despite its known quota on admission of Jewish students. I have no idea what made me stand out. My two brothers went to Yale and to the University of Chicago. Both became university professors, one of pediatrics at Washington University in St. Louis and the other of neuroscience at Albany Medical College.

HARVARD AND CHELSEA NAVAL HOSPITAL, 1942–1944 I began college in the summer of 1942, only a few months after Pearl Harbor and the engagement of the United States in World War II. I was not yet 17 and was too young to be drafted or to enlist in the armed service training program at Harvard or other universities. Fifteen months later, however, I enlisted in the Navy V-12 program at Harvard College. Kirkland House, where I lived, had been commandeered by the V-12 program. I have vivid memories of that period—up at dawn to run a three-mile loop around the bridges that cross the Charles River. Lights were out at dark because of the fear of attack, presumably from enemy submarines. That did not give much time for studying, so after dark Alex Rich (who lived in the same entryway and remains a close friend) and I would sneak under the stairs at the basement level with a blanket over our heads and read by ﬂashlight. Harvard had three semesters per year and no summer vacation. I was only at Harvard for about 20 months, but in that time I completed 29 of the 32 courses required for graduation. I majored in psychology while taking the minimum number of courses that would satisfy the requirements for entrance to medical school. I was not much interested in biology or the other natural sciences, and I thought that psychology would be an excellent preparation for

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medical school (or, as a I later realized, for running a laboratory). I worked as a volunteer at Massachusetts General Hospital and was on duty the night of the Coconut Grove ﬁre that claimed nearly 500 lives. I can still see the charred bodies lining a main corridor of the hospital. I was at Harvard for only ﬁve and a half terms. I do not know how I managed to take so many courses in that period, but the last ﬁve were easy. Students were continuously being sent away for military service, so the university had a policy that if you completed the midterm of a course, you got full credit. As soon as the Navy realized in March 1944 that I had completed my premedical requirements, they assigned me as a corpsman to Chelsea Naval Hospital across the Mystic River. It still stands as a condominium complex today. At the naval hospital, I had the top deck of a triple-decker bunk; one morning I broke my big toe leaping out of bed when I realized that the commandant was coming up the stairs for an inspection. I was assigned to the plaster room, where my job was to roll plaster after various kinds of broken bones had been set by the naval doctor. The plaster room was run by a marine who had returned from the South Paciﬁc and was often sufﬁciently inebriated so that the running of the plaster room was left to me and another V-12 corpsman (Sig Gunderson). I was the oldest grandchild on both sides of the family and the family’s ambition was that I become a doctor. It is a familiar story of upwardly mobile immigrant families that I see repeated today in the children of recent Asian immigrants in their quest for both economic stability and a useful humanitarian life. I had no interest in science and no exposure to it. U.S. Navy policy in those days was that a V-12 corpsman could apply to medical school; those who were admitted were sent to school as naval trainees. I was admitted to Yale Medical School in September 1944, after only ﬁve months at Chelsea Naval Hospital.

YALE MEDICAL SCHOOL, 1944–1948 Yale Medical School was then and is now an unusual place. My class had only 44 students, so I knew all of them; a third became close friends. There were no examinations and no forced attendance at any class. The only requirements were to pass Part I of the Examination of the National Board of Medical Examiners (Basic Science) after two years and Part II (Clinical Science) after four years. In addition, an original thesis was required at graduation. We took the usual courses in the ﬁrst two years. During the ﬁrst year, while taking physiology, I met John Brobeck, then an assistant professor in physiology. He was engaged in the study of hypothalamic hyperphagia induced by lesions in the ventromedial nucleus of the hypothalamus placed in rats with electrodes attached to the stereotactic Horsley-Clarke instrument. He needed someone to help him by measuring the food intake of these rats and their controls on various diets and with various activities, to weigh them, and later to help in placing lesions. Medical school seemed a little cut and dry, and, although I am not sure how, I got the job. It was simple, but I liked it. It allowed me to escape the routine of medical school, and John was a stimulating mentor. The job lasted through medical school, and I wrote my thesis on the topic. I was the ﬁrst recipient of the Borden Undergraduate Research Award at Yale Medical School. I wrote four papers on this topic, none of them startling, but I was ﬁrst author of three and sole author of one, the ﬁrst entry in my publication list. I was also the ﬁrst author of an abstract, presented at the Federation meetings, but I do not remember whether I presented it or John did. I was hooked. Research was fun, and there was a small conference room where I played bridge with my friends and any nursing students we could persuade to join us. Female medical students were very rare. A seventh paper also early appears in my publication list, entitled “A National Science

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Foundation.” It is a summary of the congressional debate regarding whether to establish NSF. It was written in connection with my membership in the Association of Interns and Medical Students (AIMS) in which the Yale chapter was very active, discussed more below. The paper was republished recently as a “classic” by the Yale Journal of Biology and Medicine in which it originally appeared (1). I am not sure whether it is considered a classic because of what it said or because its author had by then become a prominent scientist. It was wartime when I began medical school. I was still in the Navy, but our uniforms had changed from those of seamen to those of ofﬁcers in training. Medical school was on an accelerated basis, i.e., three semesters per year, so that eight semesters, or four years, could be completed in slightly less than three years. Suddenly, in 1945, the war ended, ﬁrst in Europe and then in Asia. I completed my second year 16 months after I had started, in March 1946, and I was discharged from the Navy. The third year would not begin again on the regular schedule until September of that year. I had six months with nothing to do. I decided on an adventure. Another assistant professor in physiology, Bob Livingston, was from Seattle, and his family knew the owner of a logging company based in Seattle but operating in British Columbia, the O’Brien Logging Company. He got me a job with the company. I thought it would be great fun, but John Brobeck did not agree. He made me a deal: If I worked for his friend, Leo Samuels, the chairman of the Biochemistry Department at the relatively new Medical School of the University of Utah in Salt Lake City, for three of the six months, John would help with my transportation expenses. Salt Lake City also seemed like a great adventure to someone who had never been off the East Coast, so I took him up on the offer. My job was to measure the body fat of rats. This involved macerating the whole rat in a meat grinder, extracting the lipids, and then titrat-

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ing the fatty acids with sodium hydroxide, my ﬁrst experience in biochemistry. For the remaining three months, I went to Seattle and then on to the coast of British Columbia, somewhere between Stillwater and the Queen Charlotte Islands, where the O’Brien Company was operating. Its owner was extremely concerned that a medical student could hurt his hands in the logging operation, so to my extreme disappointment they put me to work washing dishes for the crew. It took me several weeks to talk my way out of that. I was then promoted to pounding spikes on a railroad being built from the site of the logging camp to the coast, so that the logs could be taken down to the sea and ﬂoated. Finally, after another few weeks, I managed to get where I wanted to be, working in the crew that was harvesting gigantic cedar trees, 6 to 10 ft in diameter. My job was to “choke” the log that had been topped by the “toppers” and other crews that had then felled the gigantic trees and cut them into sections of about 20 ft. Choking the log meant hauling a small steel cable wire from the top of a crane that had been erected at a collecting site, to a log, then around it and back to the crane. A very heavy chain could then be pulled out by the wire to “choke” the log, i.e., to fasten the heavy chain around the log. The log would then be pulled back to a pile at the site of the crane and eventually loaded on a railroad car bound for the sea. It was a solitary, isolated life among men of a totally different background than I had experienced. I had a lot of time to think. Medical school began again in September with the clinical courses. I remember only a few things from those two years. Francis G. Blake was professor of medicine and a prominent American physician. While I was taking my medicine clerkship in 1946, he received a new drug to administer to a patient at the Yale–New Haven Hospital. It was called penicillin, and it was said to cure infectious disease. This was given to a patient at the hospital with miraculous results; I believe it was the ﬁrst patient treated with penicillin in the

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United States. It made a deep and lasting impression on me. I also delivered several babies, or more precisely, I caught them as they were given birth by multiparous women. During my surgery clerkship, I rebuilt a sailboat from a hull that had been washed up on Lighthouse Beach during the famous hurricane of 1938; the rigging and sails were intact in a nearby barn. I continued to work with John Brobeck. I passed Part II of the National Board of Medical Examiners, ﬁnishing second or third in my class, to the surprise of the faculty, my fellow classmates, and myself.

INTERNSHIP AND POSTDOCTORAL FELLOWSHIP AT WASHINGTON UNIVERSITY SCHOOL OF MEDICINE, 1948–1951 I had obtained my internship in internal medicine on the Ward Medical Service at Barnes Hospital in St. Louis afﬁliated with Washington University School of Medicine. The professor of medicine, W. Barry Wood, had been appointed in his early thirties and was one of the young stars of American medicine. Earlier, he had been an allAmerican football player at Harvard College (and liked to demonstrate his football prowess by inviting all the interns and residents on Sundays for a pickup game of touch football at his beautiful suburban residence). He was also extremely interested in research. I did not like being an intern. It seemed very routine. I decided not to pursue a residency but instead to become a postdoctoral fellow. On the advice of the dean of Yale Medical School, C.N.H. Long, a biochemist whom I knew well enough to ask for guidance, I applied to Oliver H. Lowry, who had been appointed chairman of the Department of Pharmacology at Washington University and was in his mid- to late-thirties. Very few fellowships were available at that time, but ultimately I received one under circumstances that were so accidental and unusual that I cannot relate them

here. Science at Washington University was famous worldwide. Joseph Erlanger (physiology) and Herbert Gasser (pharmacology) were early neurophysiologists who had uncovered the relationship between neuroﬁber size and velocity of nerve conduction. Importantly, Carl and Gerty Cori were widely known for their studies of the mechanism of glycogen synthesis and of glycogen storage diseases. Carl had recently vacated the chairmanship of pharmacology to become the head of biochemistry one ﬂoor below (in terms of contemporary issues, Gerty was a research associate; she became a professor after they shared a Nobel Prize). Research in physiology, pharmacology, biochemistry, and microbiology, where Jacques Bronfenbrenner had been involved in the discovery of bacteriophage and Al Hershey had initiated his famous studies of these bacterial viruses, was very active. Martin Kamen, who had used C11 (a very short-lived carbon isotope, the only radiocarbon known at the time) as a tracer in biology and had then created C14 as a very long-lived tracer, was carrying out research in the Radiology Department. And Ollie Lowry had undertaken a difﬁcult problem, the dissection of single cells from the nervous system and the measurement of their enzyme activities. What an atmosphere in which to be introduced to research! I stayed for two years, although I did not do extremely well. Somewhat discouraged, I opted to go back to ﬁnish my residency in medicine at the University of Chicago, where the young woman I later married was a student. But then the drama began. The Korean War had started in 1951, and those of us who had been trained by the Navy but had not served were called back into service in the spring. I learned that Washington University School of Medicine had been asked to help vitalize medical education in Bangkok, Thailand, by providing staff for the medical school at Siriraj Hospital and that I could join this program as an ofﬁcer in the U.S. Public Health Service as an alternative to the U.S. Navy. I enlisted. Ann and I were www.annualreviews.org • My Scientific Journey

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married in May and went to Bangkok a month later, we thought for two years. By the time we left for Thailand, I had published two abstracts and completed the work for one paper.

BANGKOK AND A SECURITY HEARING, 1951: HOW I BECAME A BIOCHEMIST

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Bangkok was extraordinarily exotic then, a relatively small city of 500,000 people. Many of the streets were canals (klongs). Each morning, I took a small boat to Siriraj Hospital across the Menam Chao Praya, a large, dirty river that drained much of the country. I carefully sat in the middle of the boat, not wishing to be splashed with the unsanitary water. My task was to help Professor Ouay Ketusinh, the chairman of pharmacology, modernize teaching. I met the members of his department and we talked and consulted every day. I also had many tasks to establish a life in Bangkok: renting a house from a prince (of which there were many because the kings had formerly been polygamous), purchasing a car, and ﬁnding furniture. It did not last long. After only two or three months, I was abruptly called to the ofﬁce of the chief of the Public Health Service mission to whom I was responsible and then to the American ambassador. A cable had been received that I was to leave Bangkok within 24 hours and return to Washington. Why? I asked. The cable message contained a coded reference; the ambassador opened a reference book that showed that I was being recalled under a code that deﬁned me as a security risk. I had no idea why. In protesting the haste, because I had a car to sell, furniture to deal with, and a rental contract from which to disengage myself, I was given an additional 48 hours. I wanted to stop in Europe on the return trip, but that was not permitted. However, the travel ofﬁce carefully arranged for my plane from Bangkok to arrive in Amsterdam after the last (possibly the only) departure for the United States. So in fact, I had a 20-hour stopover. 6

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Soon after we arrived in Washington, I called a friend, Arthur Murphy, who had married a young woman I had known at Wellesley when I was an undergraduate. She had recently given birth to their second child and had been taken to the hospital that day with a high fever. She died four days later of bulbar polio; post-partum women were unusually susceptible. It was only a few years before the introduction of polio vaccination. Why do I mention this personal episode? It gave me enormous perspective during the not-soserious events that followed. The next day, I reported to the chief of the Division of Commissioned Ofﬁcers in the Public Health Service, who asked me to resign my commission or the president would revoke it. Why? I asked again. I was told that I was a security risk, but they would not provide any details. “Please resign your commission.” Fortunately for me, my brother, who was a medical student in St. Louis then, managed to notify Oliver Lowry and Robert Moore, the dean of the Medical School, about what was happening to me. After all, I had originally gone to Thailand under the auspices of Washington University. They protested mightily, and I was told that I would receive a bill of charges and ultimately a security hearing, if I wanted one. My friend Arthur Murphy was a Justice Department lawyer just out of law school. He could not help me in a case that involved the government, but he told me of several law ﬁrms in Washington that were taking cases such as mine pro bono. First, I went to see Mr. Abraham Fortas, a member of the ﬁrm of Fortas and Porter at that time (and later a Supreme Court Justice). He listened sympathetically but told me that their ﬁrm already had too many pro bono cases. He and Arthur Murphy both suggested another Washington ﬁrm, Covington and Burling. There, another young lawyer, Paul Warnke, Arthur’s classmate and a former law clerk with Dean Acheson, agreed to be my lawyer (Dean Acheson had, by that time, taken leave from the ﬁrm to be secretary of state). Paul later became assistant secretary of defense for international

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security affairs and still later was appointed as ambassador to negotiate the Strategic Arms Limitations Treaty (SALT) with the Soviet Union. I met many young lawyers in Washington during this period, and, aside from “the problem,” it was a very interesting experience. Paul immediately asked for the bill of charges, which the Loyalty Review Board reluctantly agreed to provide after several months. The bill listed three charges: (a) I had had friends in medical school who were allegedly members of the Communist Party; (b) I had a library of communist books; and (c) I had subscribed to the Daily Worker, the publication of the American Communist Party. None of these was true to my knowledge. What had happened? At the time I left, the FBI had not completed its investigation of me, routine for those assigned to an overseas mission. When they did, they decided I was a security risk. As mentioned above, I had been an active member of the Association of Interns and Medical Students (AIMS). AIMS had campaigned for the introduction of health insurance in the United States, which, at that time, did not exist in any form, private or public. As a result, the powerful American Medical Association branded the organization communist. All the individuals listed in my bill of charges had been AIMS members. The politics of some were to the right and others to the left of mine, but I had never asked any of my friends about their formal political afﬁliations, nor did I think it was appropriate for the Loyalty Review Board to ask. I had obtained my so-called “library” of communist books in a political science course at Yale while in medical school, one of three that I took in late afternoons to complete the requirements for my AB degree from Harvard. There were texts by Engel, Marx, Lenin, and others. My alleged Daily Worker subscription was actually the Progressive, the publication of the party by the same name led by Henry Wallace, of which my landlady, through whose mailbox my own mail came, told the FBI when they interviewed her about me.

The security hearing was short, and I was cleared. However, the Public Health Service told me that I could not return to Bangkok because that would require an additional clearance by the State Department, which had a much tougher Loyalty Review Board, and an additional six months to schedule a hearing. Instead, they suggested an assignment to the National Institutes of Health for the remainder of my appointment as a commissioned ofﬁcer. NIH? What was that? I had never heard of it. At that time, NIH was a small operation in Bethesda, Maryland, composed of only ﬁve or six buildings. But it was a stroke of good fortune. It was really the beginning of my scientiﬁc career. I owe it to Senator Joseph McCarthy!

NATIONAL INSTITUTES OF HEALTH, 1951–1954 To join the NIH, I had to ﬁnd a lab that would accept me. I soon discovered that there were many interesting young scientists at NIH, and I spoke with many of them. I was interested in the National Heart Institute, but the director told his laboratory chiefs that he would not agree to my placement; he did not want his institute associated with someone who would have a security hearing. In the end, only two individuals offered me placements within their laboratories: Sanford Rosenthal, chief of the Laboratory of Pharmacology in the National Institute of Arthritis and Metabolic Diseases, and Seymour Kety, the newly appointed director of the newly established National Institute of Mental Health. The latter, however, was so new that it did not yet have laboratory space, so I gratefully accepted Sanford Rosenthal’s offer. Among the many honors I have received, I treasure the invitation to give the lecture at NIH in honor of his eightieth birthday. The Laboratory of Pharmacology was a fortunate choice. On my ﬁrst day at work, I walked into Sanford’s ofﬁce in Building 4 and asked what he would like me to work on. He said that my one-unit laboratory was www.annualreviews.org • My Scientific Journey

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adjacent to his ofﬁce and that I could work on anything I wanted. I was stunned. I was 26 years old and had little research experience, but I had a fully funded independent laboratory. What should I work on? Before leaving Oliver Lowry’s neurochemistry laboratory in St. Louis, I had been thinking about focusing my effort on how drugs work in the nervous system, e.g., why does morphine obtund pain without loss of consciousness, whereas barbiturates put you to sleep? These drugs must have differential effects within the complex cellular components of the nervous system. Could the effects be studied with the single-cell dissection techniques pioneered by Lowry? I decided it was much too difﬁcult a problem for 1951 and that, until we could understand how a drug like penicillin worked on a homogenous cell population like bacteria, unraveling the complexities of the nervous system would be too difﬁcult. Thus, penicillin’s mechanism of action was on my mind when Alan Mehler, a member of the Rosenthal laboratory and earlier Severo Ochoa’s only graduate student, pointed out an interesting abstract that was published in the most recent Federation Proceedings. It was based on the PhD thesis of James T. (Ted) Park at the University of Wisconsin, who had isolated from penicillin-inhibited staphylococci a very unusual nucleotide. It contained uridine diphosphate, linked to a previously undescribed sugar whose structure had not yet been elucidated, to which was linked a peptide. Could this nucleotide be involved in the then mysterious relationship between nucleic acid biosynthesis and protein biosynthesis? Alan also told me that Ted Park was in the armed services and had been assigned to the Army Bacteriological Warfare Laboratory at Fort Detrick, Maryland, very close to Bethesda. Park was not able to continue to work on the nucleotide he had discovered. I phoned him and we met for a picnic lunch at a brook outside the gates of Fort Detrick because I was not allowed in. He was very generous in providing me with advice and helping me get started. I had found a problem that really interested me

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and a way to get into it. Was this nucleotide that accumulated in Staphylococcus aureus related to the mechanism of action of penicillin? Did it have anything to do with protein and nucleic acid synthesis? Alan Mehler was a well of knowledge about biochemistry as was Herb Tabor, another member of the Rosenthal lab and now the editor of the Journal of Biological Chemistry. Through them I was introduced to the biochemical ferment then being generated at NIH by young scientists in Building 3 (several of whom were close friends with Herb Tabor and Alan Mehler), such as Arthur Kornberg (now known for his studies of DNA synthesis), Bernard Horecker (mechanism of carbon dioxide ﬁxation for sugar synthesis in plants), Jesse Rabinowitz (1-carbon transfer involving folic acid derivatives), Bruce Ames (environmental carcinogens), Leon Heppel and Herman Kalckar (nucleotide metabolism), Osamu Hayaishi (oxygenases in mammals), Earl and Terry Stadtman (microbial biochemistry), Christian Anﬁnsen (protein structure), Bernard Brodie and Julius Axlerod (drug metabolism), Herb Tabor (spermine and related polyamines), Dan Steinberg (phytanic acid storage disease), and many, many others. Three of these individuals later received Nobel prizes, and all have ultimately been elected to the National Academy of Science. My work evolved in two directions: the work I did independently on the mechanism of penicillin action, and the work I did in collaborations (principally with Herman Kalckar and Leon Heppel, which allowed me to gain expertise in enzymology). First, I learned how to use the Park method to isolate the unusual uridine nucleotides from penicillin-inhibited S. aureus (2). It involved fractional precipitation of mercury salts by ethanol and ultimate measurement of the amount of nucleotide by the UV absorption of uridine. It was clear that I would not make very much progress using this time-consuming and inexact procedure. Then I came across an obscure paper that described a method for measuring glucosamine. Could it be adapted

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to measure the Park nucleotide after mild acid hydrolysis to free the acetylamino sugar? The molar extinction coefﬁcient of the chromagen formed in the published procedure was exceedingly low, but Oliver Lowry was a methodologist, and I had learned that playing with conditions of colorimetric reactions could be rewarding. Late one afternoon, I decided to try different buffers at different pHs. The last one I used was sodium tetraborate, an internal salt that was used in the Lowry laboratory as a pH standard, pH 9.15, and that I had on my reagent shelf for that purpose. I added it, went home for supper, allowing the various trials to incubate at 37◦ . When I came back after supper, my eyes bulged. A tube to which I had added borate was an intense dark purple color. This small thing was the break I needed, and it provided me with a quantitative method of measuring the accumulation of the Park nucleotides. I only published the details of the method a few years later when I encountered Luis Leloir in Copenhagen and learned that a student in his laboratory had developed a similar method. We published it together, four years later in 1955 (3). Two years after that, I published the use of the method to measure the kinetics of accumulation of the nucleotide and many of its features, using 50– 100 ml cultures (4). Almost nothing had to be rushed into publication in the late 1950s. During the same period, my work with Leon Heppel taught me how to purify enzymes and to keep careful track of units of activity, yields, and speciﬁc activity during the puriﬁcation procedure (5), knowledge that proved extremely important later. With Herman Kalckar, who had recently come to NIH from his native Denmark, we isolated another uridine diphospho-sugar compound (UDP-glucose) that had been described by Leloir. Again, we began with precipitation of mercury salts from yeast, but now used the recently introduced anion exchange chromatography to purify the nucleotide. We built our own fraction collector (they could not be purchased then). We knew that UDPglucose was important, but we did not have

a good idea what to do with it. Then one afternoon, in the men’s room, I encountered Julius Axlerod and, while standing side by side, asked him what he was doing. He was studying the synthesis of morphine glucuronide in liver slices. Soon after, we designed an experiment in which UDP-glucose was added to minced liver slices together with the oxidizing agent, NAD (nicotinamide adenine dinucleotide) [then called DPN (diphosphopyridine nucleotide)]. UDP-glucuronic acid was formed and used for the synthesis of the glucoronide. In one afternoon, we had carried out the ﬁrst metabolic transformation of a sugar while attached to a nucleotide (6). This technique has had many extensions for other nucleotides of this type and for other sugars. For example, Herman Kalckar went on to use it to study the utilization of galactose in mammals and to discover that mutation in a uridyl transferase in which galactose 1-phosphate is exchanged for glucose 1-phosphate in UDPglucose is the defect in the metabolic disease congenital galactosemia (7).

SABBATICAL LEAVE IN COPENHAGEN, DENMARK, AND CAMBRIDGE, UK, 1955 After two years at NIH, I knew I wanted to leave. Despite the scientiﬁc excitement, the security hearing had left a bad taste in my mouth for government service. Oliver Lowry invited me to return to his department in St. Louis as an assistant professor. I said that I would like to take the job but that I wanted to see how science is conducted in Europe ﬁrst. He gave me the ﬁrst year of my new appointment as a sabbatical leave, and I obtained support from the Commonwealth Fund for the year in Europe. I wanted to go to Cambridge University, but Ernest Gale, a microbiologist in the Biochemistry Department who was studying penicillin, had no room in his lab that year, and Roy Markham could only take me for the second half. Lowry had worked in Copenhagen with the prominent Danish protein chemist, Kaj Linderstrom-Lang at the www.annualreviews.org • My Scientific Journey

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Carlsberg Laboratory, and suggested I spend the ﬁrst six months there. It turned out to be an amazingly fortuitous experience. The Carlsberg Laboratory on the grounds of and funded by the Carlsberg Brewery was at that time one of the principal research institutions in biochemistry in Europe. Linderstrom-Lang was a joyful, gregarious laboratory head whose laboratory had previously seen development of the ﬁrst method for nitrogen determination by Kjeldahl and then the introduction of the concept of pH by Sorenson. A very important feature of the laboratory has been often noted by others, but I will say it again. It had a huge two-way refrigerator. One side opened to the laboratory and the other side to the brewery from which it was kept stocked. I needed the cap of a bottle of Carlsberg Beer to illustrate the match shooting game that took place every day at lunch time. The other postdoctoral fellows and sabbatical visitors there at the same time included Chris Anﬁnsen (NIH and Johns Hopkins), Bill Harrington (Johns Hopkins), Fred Richards (Yale), Ieuan Harris (Cambridge, UK), John and Charlotte Schellman (Oregon), and Don Wetlaufer (Minnesota). Chris and Bill lived only a few blocks from us in Hellerup, a Copenhagen suburb. Bill was the only one with a car, so the three of us commuted across town to the Carlsberg Laboratory, 30 minutes each way, more time to talk about science. My bench space, about four feet, had a cupboard full of copper pots that I wanted to clear out. Nothing doing; they were Kjeldahl’s pots. Everyone was working on some aspect of protein or peptide chemistry, so I thought I might as well do the same. I worked out the sequence of the pentapeptide in UDP-acetylmuramylpentapeptide, the Park nucleotide: l-Ala-γ-dGlu-l-Lys-d-Ala-d-Ala, using simple partial acid hydrolysis and paper chromatography, coupled with the stereospeciﬁcity of several enzymes to identify the stereoisomer (8). There was a steady ﬂow of important scientiﬁc visitors; three of them were critical to me. Milton Salton from Cambridge said he

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had described a new morphological structure called the bacterial cell wall and that it contained a very unusual sugar. Later I showed that this sugar and the one in the uridine nucleotide were the same. Luis Leloir from Buenos Aires came, and during discussion we discovered that we each had invented a new method for acetylglucosamine determination, which I have already described above. Esmond Snell from Berkeley arrived and talked about a peculiar dipeptide that was present in large amount in lactobacilli and had been identiﬁed as d-alanyl-d-alanine. That was it, I was on the track. But the six-month stay in Copenhagen came to an end. By then, Fred Richards had moved to Cambridge and, because Ann was by this time eight months pregnant, he had found a ﬂat for us on Brunswick Walk facing Midsummer Common. Any readers who have been to Cambridge will know that beautiful spot. The ﬂat, however, was a little squalid. As a result, Ann was allowed to have a hospital delivery at Mill Road Maternity Hospital, which is fortunate because it was a difﬁcult delivery and our son Andy had to be extracted quickly. A month later, Alex and Jane Rich moved in for a week’s stay with our two-year old son, Paul, allowing Ann, me, and Andy to have a driving holiday in Scotland all the way to Skye. I was at the Molteno Institute for Parasitology, where Roy Markham had his laboratory. I learned a great deal about separation of nucleotides by paper electrophoresis and worked with Leslie Mapson across the road on UDP-glucose dehydrogenase from plants. Roy’s laboratory was on the top ﬂoor. A wonderful older scientist, David Keilin, was on the ﬂoor below. He had discovered cytochromes while looking at translucent worms with a hand spectroscope and later found them in many other organisms, including translucent Antarctic ﬁsh that have no hemoglobin and swim slowly under the ice. His museum was on the top ﬂoor with Roy’s lab. It doubled as a small lecture theater. Then one day, Max Perutz began his lecture course on protein

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crystallography in that lecture space, the ﬁrst such course anywhere in the world, for perhaps a dozen students. I had no choice but to listen, although I understood very little, because Max had asked Roy to request that that “young American” not run the centrifuge while he was lecturing.

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WASHINGTON UNIVERSITY SCHOOL OF MEDICINE, ST. LOUIS, 1956–1963 Time passed quickly, and I was soon back in St. Louis as an assistant professor of pharmacology. I shared a lab initially with Bob Furchgott who had just discovered endothelial relaxing factor, later identiﬁed as nitric oxide, using a Rube-Goldberg-type apparatus common to pharmacologists at that time. Bob left six months later to become head of pharmacology at Downstate Medical School in New York, and much later he was widely recognized for the work he had done in St. Louis and New York. I had my own small lab again. On the way back from Europe, I had met Ted Park again. He was working on the same phenomenon that I was, namely, that the uridine nucleotide that accumulated in penicillin-treated staphylococci was a precursor of the bacterial cell wall. We published two papers together rather than compete with separate publications (9, 10). Isotope studies strongly supported the conjecture (11). I spent the next eight years showing that the nucleotide was in fact a biosynthetic precursor of the cell wall, isolating cell walls, determining their structure with Jean-Marie Ghuysen using the speciﬁcities of bacteriolytic enzymes to dissect the structure (12), and working out the mode of biosynthesis of UDPacetylmuramyl-pentapeptide (13). Finally, I got my foot in the door and we developed a cell-free system to study the utilization of two nucleotides, UDP-acetylglucosamine and UDP-acetylmuramyl-perntapeptide, for cell wall peptidoglycan biosynthesis (14), and we also extended the study of transformations of sugars while attached to nucleotides (15). I had

my ﬁrst excursion into immunology [although in fact the components of the l-lysine and meso-diaminopimelic acid–containing peptidoglycans, whose structures were elucidated, have in the past year turned out to be key molecules in understanding a component of innate immunity, the intracellular nucleotidebinding oligomerization domain (NOD) receptor for these components of the peptidoglycans] (16). Teichoic acid, a speciesspeciﬁc “ornament” attached to the cell wall of S. aureus, has an acetylglucosamine residue as a substitution on polyribitol phosphate. Rabbit antisera prepared against S. aureus cells agglutinated cell walls of this organism and were speciﬁc for α-acetylglucosaminyl ribitol, which accounted for only 15% of the total acetlyglucosamine, whereas the remaining 85% of β-acetylglucosaminyl ribitol was immunologically inert (17). Another excursion into immunobiology was a minisabbatical at the Wanderforschungs Institute (now the Max Planck Institute for Immunology) in Freiburg, Germany. The work there was focused on the chemical structure of lipopolysaccharides (LPS) of Gram-negative bacteria (18). A particular focus was the relation between the serological speciﬁcities of the many strains of Salmonella and Escherichia coli and the structures and linkages of the sugar components of LPS, including several unusual sugars, 3,6-dideoxyhexoses such as colitose and tyvelose, in a collaboration with Fritz Kaufmann, a prominent bacterial serologist at the Statens Serologische Institut in Copenhagen. More about how this work got me into HLA below.

UNIVERSITY OF WISCONSIN, 1964–1968 I had found myself increasingly frustrated by the opportunities available at Washington University and about the lack of scientiﬁc stimulation. The school was in temporary doldrums. So, when an offer came to be chairman of the Pharmacology Department at the University of Wisconsin, I took it. The university www.annualreviews.org • My Scientific Journey

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was extremely generous to a department chairman. In addition to secretarial help, they provided positions for a senior research associate, a technical assistant, and a dishwasher and substantial funds for supplies. Those four years at the University of Wisconsin were extremely productive. The foundation for my work had already been laid at NIH and at Washington University. The structure of cell walls (of which I am especially proud) had been outlined, particularly the presence in them of peptide cross bridges between acetylmuramyl-peptide subunits. The structural analysis led Donald Tipper to the structural analog hypothesis of penicillin action (see below). A particulate enzyme system that catalyzed cell wall biosynthesis from two uridine nucleotide precursors had been established (14), or, at least, we thought it had been established. Initially, in the radioactive incorporation assay, we found 50 cpm of labeled precursor incorporated above a background of 50 cpm. As we continued to work with the system, it soon went up to 500–1000 cpm above the background. I do not know why. We did not change anything. Years later, the same story was repeated in the yields of HLA proteins. You have to have intuition and faith sometimes. A long series of papers have described the steps in the biosynthesis of the peptidoglycan of bacterial cell walls, including the species speciﬁcity in the nature of the peptide cross bridges and the utilization of diaminopimelic acid–containing nucleotides (rather than those containing lysine) in E. coli (19). With each new step in the biosynthesis that we discovered, we tested its sensitivity to penicillin. None were sensitive until the very last step. Finally, we could show that the formation of cross bridges in the particulate system was inhibited by penicillins, and we postulated that the LD-cyclic dipeptide, penicillin, was an analog of the C-terminal end of acetylmuramyl-pentapeptide, i.e., dalanyl-d-alanine, that released d-alanine in the trans-peptidation reaction to form the cross bridge (20). Only recently has it been shown that the hypothesis of the structural

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analogy was correct, i.e., that substrate and penicillin lay in the same space in the enzyme (21). Michio Matsuhashi and Kazuo Izaki identiﬁed the double-headed enzymes that catalyzed both trans-glycosylation and trans-peptidation (19). “Enzymes” is plural because multiple penicillin-binding proteins were described that have different roles in cell wall synthesis and that catalyzed this reaction. Along the way, lipid intermediates in peptidoglycan synthesis were also described by John Anderson and later identiﬁed by Yasuo Higashi as C55 -isoprenyl phosphate, the lipid that is involved in the transport of these molecules across the cell membrane (22, 23). There was another research vein also, the description of unusual bacterial sugars and of the mechanism of their biosynthesis. These included 3,6-dideoxyhexoses and two new ones, 4,6-dideoxy-4-acetamidohexoses, that are involved in speciﬁcity determination among different bacterial strains (24). These studies had important consequences later. What has this to do with immunology? Surprisingly, a lot. I had often wondered why no immunological system involving the peptidoglycan or its various components had been described—no antibodies and no T cell recognition phenomena. They are unique bacterial components, and they seemed like logical targets for the immune system of mammals. It took 30–40 years longer for their importance in immunology to be outlined. JeanFrancois Petit, who had worked on peptidoglycan structure and synthesis in my lab (25), returned to Paris and was involved in the discovery that muramyl dipeptide [really Nacetylmuramyl dipeptide (26)] and its derivatives are powerful immunoadjuvants (27), but he had no explanation of the mechanism. Only in the past 10 years, since the innate immune system has become a center-stage player in immunology, has it become clear that these substances are ligands for receptors in the innate immune system helping to initiate immune system responses (28). And, in the past year, a second innate immune system to deal with intracellular bacteria has been uncovered

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(16). Acetylmuramyl-l-Ala-γ-d-Glu-mesoDap, or more simply γ-d-Glu-meso-Dap, called iE-DAP, was shown to be a ligand that initiates immune responses as a ligand for the NOD1 receptor (approximately 50 years after its discovery as a component of the cell wall peptidoglycan of some Gram-negative bacteria!). NOD2 recognizes N-acetylmuramyl-lAla-γ-d-Glu, “muramyl dipeptide.”

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HARVARD UNIVERSITY 1968–PRESENT In the midst of this exciting work, a phone call came from Konrad Bloch and John Edsall, who invited me to join a committee on biochemistry and molecular biology at Harvard as professor of biology, with the expectation that the committee would become a department within a few years. It was an irresistible opportunity to join a small group of outstanding scientists to form a new department in a major university. The other members were Paul Doty, Matt Meselson, and Jim Watson. I was invited to ﬁll a niche in research and instruction in microbial biochemistry, a central theme in biology at that time. Neither they, nor I, knew that I would soon turn my attention to molecular immunology. I had done research in pharmacology for the past 17 years and had been associated with a medical institution for more than 20 years. I welcomed the opportunity to join the faculty of arts and sciences with the stimulation and challenges that a new environment was certain to bring. I had thought for about a year that the penicillin problem was almost solved and that I should look for new pastures. Of course, 40 years later, it still has not been completely solved, and much interesting current work that builds on our old work has been stimulated by the emergence of drug-resistant bacteria. I had only recently passed my fortieth birthday, and I thought that if I were to have an exciting scientiﬁc future, I should ﬁnd another problem. Transplantation biology was in the air, brought there by the advent of kidney grafts and then, in the early 1960s, by heart grafts.

Little was known about the mechanism of allograft rejection, and there was no knowledge of the transplantation antigens that had been named by Peter Gorer (29) during his studies of graft acceptance or rejection in outbred mice. Gorer’s work stimulated the development of inbred strains of mice by George Snell (30). When I moved to Harvard, I decided that it was now or never to change ﬁelds. At ﬁrst, only two brave postdoctoral fellows (Wolf Droege and Dieter Malchow) were interested in working in this new area. The rest of the lab continued to work on bacterial cell walls, the mechanism of their biosynthesis, and how penicillin worked. Over a period of 15 years, however, the number of people working in the former slowly increased and those working in the latter decreased. By 1985, no one who came to the lab was interested in microbial biochemistry. I knew there was interesting work still to do in that area, but I had no choice but to stop working on the penicillin and bacterial cell wall problem. I had been continuously supported by an NIH grant in this area for 29 years, from 1956 when I moved from NIH to Washington University and obtained my ﬁrst research grant, until 1985 when I did not renew it. One notable event inﬂuenced my decision to move toward the study of histocompatibility proteins. In the mid-1960s, Salvador Luria from MIT obtained a grant from NSF to organize a yearly conference that he called “Microdermatology”—the study of the surfaces of microorganisms. The meetings were always held in some interesting place, and one year it was held at the Abbay´e-Royaumont near Paris. Salva always invited an additional speaker working in some distantly related area. That year, the speaker was Allan Davies from the Microbial Research Establishment at Porton, UK (Britain’s equivalent of Fort Detrick). Allan had discovered a number of the 3,6-dideoxyhexoses that give speciﬁcity to bacterial surfaces. He described this work and ended with the speculation that the speciﬁcity of transplantation antigen might also be www.annualreviews.org • My Scientific Journey

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Human leukocyte antigen (HLA): solving the structure of HLA proteins was especially important in the 1970s because it had become clear from the work of Jean Dausset, Jon van Rood, and many others that these substances were critical in transplantation rejection.

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determined by unusual sugars. I traveled back with him to London, and he offered to be my guide on a sightseeing trip. First we went to Westminster Abbey, just across the street from the Houses of Parliament, and sat down on a bench in front of the statue of Abraham Lincoln at one side of Parliament Square. We never did any more sightseeing. He spent the next few hours introducing me to transplantation biology. I was enthralled, and it moved this ﬁeld to the top of my ideas for future projects. Whether or not sugars were involved, it was certainly a fascinating area for research. After an abortive start trying to study the homing of lymphocytes to spleen and lymph nodes and its inhibition by pertussis toxin (as had been described by Jim Gowans), two important contributions from former postdoctoral fellows who had worked on the bacterial cell wall/penicillin problem ﬁnally got me started working on HLA antigens. When Stan Nathenson left my lab, he went to work with Allan Davies on murine transplantation antigens and discovered that they could be solubilized from the surfaces of cells by papain (31). At that time, one could not purify a membrane-bound substance, so this contribution was critical. But how could one purify a substance without any idea about its chemical nature or any way to measure it? Arnold Sanderson, who had worked on teichoic acid (32), came to Boston Children’s Hospital on a sabbatical leave from the Blond Institute at Queen Elizabeth Hospital in East Grinstead, UK. There, a Center for Transplantation Biology had been established under the direction of the Danish immunologist, Morten Simonsen. Arnold had been attempting to study human histocompatibility proteins and had even solubilized them with papain and separated the HLA-A and HLA-B speciﬁcities by ion-exchange chromatography (33). I asked if he would show me the assays. First, he showed me the trypan blue exclusion assay that was then used in tissue typing. The dye is excluded from lymphocytes, but, in the presence of pregnancy alloantiserum of Strominger

the right speciﬁcity and complement, holes are punched in the membrane and the dye enters. Thus, the percentage of blue cells and the inhibition of their appearance by soluble HLA antigen could be scored microscopically. Then he showed me the chromium51 release assay that Hans Wigzell had recently developed and that he had used in a more quantitative way. I had learned much about methods from Ollie Lowry, and with a little ﬁddling I could easily make the assay for HLA antigens strictly quantitative. I had had a great experience learning how to purify enzymes with Leon Heppel (a master of the trade), and so I set about purifying HLA proteins as though they were an enzyme, keeping very careful track of speciﬁc activity, fold puriﬁcation, and yield at each step. I quickly realized that to obtain any substantial amount of the protein, very large amounts of starting material would be needed. Fortunately, at this juncture, several generous colleagues appeared who realized how interesting and potentially important the problem was and offered to grow B cell lymphoblastoid cell lines (LCLs) in 100-l batches, ∼100 gm of packed cell (particularly Dean Mann at NIH and Jim Woody and Mike Strong at the U.S. Naval Medical Research Laboratory across the street, together with Navy Corpsman Tommy Williams, who was fantastically devoted to the project). LCLs were only a few years old at the time. Epstein Barr virus (EBV) had only very recently been discovered and been shown to be able to transform and establish LCL in vitro. Thus, one could establish LCL from the B cells of any individual. But these cell lines all had the difﬁculty that, because the products of all three HLA loci that were then known, HLA-A, HLA-B, and HLA-C, were expressed codominantly, a mixture of six speciﬁcities resulted. Then I thought that using homozygous cell lines obtained from the offspring of consanguineous marriages would simplify the problem. There were very good church records of ﬁrst-cousin marriages in the Netherlands, but still another source might be inbred religious

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populations in the United States. Dean Mann had come from such a community, the Indiana Amish, so he established a line from an individual in that community, JY. The JY cell line was HLA-A2/HLA-B7 homozygous, or so we thought. Years later, we learned that JY expresses two closely related subtypes of HLA-B7, and when class II MHC loci were discovered later, JY was heterozygous at those loci (HLA-DR4, HLA-DR6)! However, other homozygous cell lines were extremely useful in doubling the yield and simplifying the problem of separating speciﬁcities, which was essential to deﬁne the polymorphism. A further problem was funding the research. The NIH Study Section had given my application a low priority score for several reasons: (a) The amount of material was too small to be able to obtain enough to work with; (b) we were using the wrong solubilization method—we should have been using LiCl solubilization that had been reported by another group; and (c) I had no prior experience in immunology (despite my success in microbial biochemistry). Fortunately, I had sufﬁcient funds in the bacterial cell wall/penicillin problem that I could divert a small amount to support one or two people working on HLA antigens. I rewrote the grant application and paid special attention to emphasizing the LiCl extraction method. However, we continued to use the papain solubilization procedure, which we knew was far superior in yield and reproducibility. The application was eventually funded, and, although it supported important continuing work, by that time Mervyn Turner and Peter Creswell had nearly completed the ﬁrst puriﬁcation to homogeneity of human histocompatibility proteins (HLA-A2 and HLA-B7) (see Table 1) (34). Pete Creswell had already shown that papain-solubilized radioactive material seemed to contain two peptide fragments (35). In a collaboration with Howard Grey, they showed that the small subunit of the HLA antigen was β2microglobulin, a small urinary protein with an immunoglobulin-like sequence that had

been isolated form nephrotic human urine and was thought to be a free immunoglobulin domain (36, 37). Tim Springer then demonstrated that detergent-solubilized HLA antigen contains an additional transmembrane region and an intracytoplasmic region and was also composed of two subunits (38), thus ruling out the hypothesis that the two subunits were the products of papain proteolysis. Harry Orr and Jose Lopez de Castro obtained amino acid sequence information using an inherited sequenator left behind by Klaus Weber when he moved with Jim Watson to Cold Spring Harbor (39, 40). Sequencing was a struggle learned by one postdoc after another, until ﬁnally in 1982 I established the protein-sequencing facility, now a world-class and widely used facility called the Harvard Microchemistry Facility run by Bill Lane. The sequences of the heavy (α) chain allowed us to determine that it was composed of three domains and that the third of these (α3) was an immunoglobulin-like domain (39, 40). That marked the beginning of the immunoglobulin superfamily of proteins. Later, DNA sequence information showed that the three domains were encoded in separate exons, conﬁrming the deduction from the protein sequences and also later conﬁrmed by the X-ray determined three-dimensional structure, although α1 and α2 folded together to form a superdomain. Many of the sites of polymorphism could also be deduced from the ﬁrst several sequences.

CLASS II MHC PROTEINS Soon after puriﬁcation of HLA antigens (in reality what we now refer to as class I MHC proteins, or MHCI), a new postdoctoral fellow, Bob Humphreys, observed that during the last step of a puriﬁcation of the papain-solubilized protein by ion-exchange chromatography, a second protein peak of approximately equal size was eluted just before the HLA antigen (41). He ran an SDS gel and found that this material also contained two polypeptide chains of 23 kDa and 30 kDa. www.annualreviews.org • My Scientific Journey

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Table 1

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Purification of HLA antigensa

Step HL-A2b 1. Crude membranes 2. Membranes after zonal centrifugation (Peak II) 3. a. Membranes after papain b. Papain digest 4. CM52 chromatography 5. Sephadex G-150 chromatography 6. DE52 chromatography

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HL-A7b 1. Crude membranes 2. Membranes after zonal centrifugation (Peak II) 3. a. Membranes after papain b. Papain digest 4. CM52 chromatography 5. Sephadex G-150 chromatography 6. DE52 chromatography

Titer

Volume (ml)

14,600 30,900

53 23

290 25,000 50,000 15,500

Total units

Total protein (mg)

Specific activity

Purification

Yield %

776,000 710,000

986 225

787 3,150

(1) 4

(100) 91

14 9 1.4 10

4,060 225,000 70,000 155,000

145 45 ND 6

28 5,000

6.3

25,800

33

0.5 29 9 20

35,400

3.5

124,000

0.49

253,000

320

16

1,500 3,100

53 23

79,500 72,300

986 225

80.6 321

1 4

100 91

115 3,300 13,000 2,000

14 9 1.4 10

1,610 29,700 18,200 20,000

145 45 ND 6

11.1 660

8.2

3,330

41

2 37 25 25

2,500

4.4

11,000

0.56

20,000

248

14

a

In this preparation, 53 ml of packed frozen cells (RPMI 4265) were employed. First published in Reference 34. MHC class I HL-A2 is now called HLA-A2. HL-A7 is now called HLA-B7. They are products of different loci, not fully known at the time of this puriﬁcation. ND, not determined.

b

We had to study it. Rabbits were immunized, and the immune serum was found to react with a protein that was found on human peripheral B cells but not on other cells among peripheral blood lymphocytes. Just at that point, I was invited to a meeting in Aarhus, Denmark, the 1975 Histocompatibility Testing Workshop, at which the speciﬁcities of pregnancy alloantisera were deﬁned. In addition to the workshop, a symposium on recent developments was held to which I had been invited. I was very surprised to learn that Jon van Rood and his colleagues in Leiden had found some pregnancy alloantisera that also detected a B lymphocyte–speciﬁc protein. The HLA-D speciﬁcity had been previously deﬁned by the people that participated in this workshop as proteins involved in mixed-lymphocyte culture (MLC) proliferation that could not be classiﬁed as HLA-A, B, or C. Van Rood and colleagues referred to 16

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the new antisera as HLA-DR (d-Related) because these antisera also seemed to block the MLC. These speciﬁcities were found to be genetically linked to the HLA antigens (class I) in a region called the major histocompatibility complex (MHC). They were the human analog of the immune response genes that had been characterized in rodents. Structural studies taken up by Jim Kaufman and Debbie Shackelford ended with an illustration of the striking structural similarities of class I and class II MHC proteins (Figure 1) (42), differing at that stage in our knowledge mainly in the linkage of the four extracellular domains and in the demonstration using alloantisera provided by the Leiden group that a second class II MHC heterodimer, distinct from HLA-DR, could be immunoprecipitated (43). This material was called DC1. DC (now called HLA-DQ) was Dora Centis, the research assistant in Roberto Tosi’s

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laboratory in Rome who had ﬁrst noticed this type of unusual pregnancy alloantiserum. The third class II protein isotype, originally called SB but renamed HLA-DP, was discovered by Stephen Shaw at NIH. It had originally been thought that humans expressed only one class II MHC protein, whereas mice expressed two! Thus, by 1979 we had gotten as far as we could with protein chemistry of class I and class II MHC proteins, but we still had not answered the fundamental questions that had driven the initiation of this work. Where exactly was the polymorphism located within the molecule? What was the evolutionary force that had driven it? It could not have been the exchange of surgical grafts that originally deﬁned transplantation proteins. And what was the function of the polymorphism? The next steps in answering some of these questions were cDNA and genomic cloning, which had recently been introduced into biochemistry and molecular biology, and three-dimensional structure determination by X-ray crystallography. Both were in their infancy, but advances in techniques soon brought them to the center stage of modern biology.

cDNA AND GENOMIC CLONING OF HLA ANTIGENS Hidde Ploegh ﬁrst came to the lab to work on the penicillin problem as an undergraduate from Groningen, the Netherlands. He later returned as a PhD student from Leiden and began to study the biosynthesis of the HLA proteins. Members of the lab knew very little about the emerging ﬁeld of molecular biology, but Wally Gilbert was just down the hall working out the Maxam-Gilbert DNA sequencing method. When Hidde said he would like to try to clone a class I MHC cDNA, I urged him to give it a try but not to let go of his biosynthetic work until he was sure the cloning would work. The cloning did work, and he reported the ﬁrst cDNA clone for an MHC protein (44). It was not a very big clone, but Harry Orr used it to show that there were at least six class I–like genes in the human genome

(45). He later identiﬁed the additional three as the molecules now called HLA-E, HLAF, and HLA-G (46). At this point, everybody jumped in, and additional clones were rapidly reported. In our lab, the HLA-DR α cDNA was obtained by a very unusual method. A monoclonal antibody that recognized the heavy chain of HLA-DR was used to immunopurify polysomes containing only this mRNA in a single step (47)! cDNA and genomic cloning and sequencing led to the sequence of the HLA-DR α chain (48), and continued analysis revealed the complexity of the MHC class II α chain loci (49). Many labs cloned HLA-DR β chain genes; our contribution was a cDNA clone for HLA-DP (then called SB) (50). Then genomic cloning and cosmid cloning came to the fore. Michael Steinmetz in Lee Hood’s lab had been unusually successful in constructing a complete map of the MHC class II region in the mouse using cosmids (51), and we wanted to do the same for the human MHC class II region. It was not that easy. Eventually, however, Kiyo Okada made a good map from cosmid clones of subregions of the class II region of the human www.annualreviews.org • My Scientific Journey

Figure 1 An illustration of the structures of class I and class II MHC proteins as visualized in 1984 (42, 105).

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MHC (52, 53), and Thomas Spies made an overlapping map of the entire MHC class I, class II, and class III regions using the discovery that TNF genes were encoded near the center to link the two halves (54, 55). Many new genes were discovered in the class III subregion (56), although the functions of most are still to be identiﬁed. Notably, however, soon after he established an independent lab. Thomas identiﬁed two new genes near the HLA-B locus called MICA and MICB, which have recently assumed importance as ligands on target cells for natural killer (NK) cells (57).

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X-RAY CRYSTALLOGRAPHY AND THE THREE-DIMENSIONAL STRUCTURE OF CLASS I AND CLASS II HLA PROTEINS As I have said, we had gone about as far as we could in analyzing the structure of HLA proteins by 1979. Protein sequence analysis had shown that the polymorphic residues were spread throughout the linear sequence of the ﬁrst two domains of the structure (40, 58). Attempting to determine the three-dimensional structure was the next logical step. During the period that I was serving my three-year term as chairman of the department (1970–1973), a young crystallographer, Don Wiley, a student in the Biophysics Program at Harvard who did his PhD thesis on aspartic transcarbamylase with Bill Lipscomb as his advisor, was appointed as an assistant professor. He was sufﬁciently promising so that we appointed him directly after he received his PhD degree without having done a postdoctoral fellowship. Don was a generation younger than me, and so he became friends with several of my students and postdoctoral fellows. One of them tried to crystallize a class I HLA protein, with advice from Don of course, but without much success. The class of graduate students that had entered in 1978 included Pamela Bjorkman. Like all of our graduate students, she did at least three rotations in her ﬁrst year 18

Strominger

to try to decide which lab to join. She was interested in protein chemistry or crystallography. One of her rotations was in my lab. One of my graduate students, Jim Kaufman, who was a year or two ahead of Pam and was working on MHC class II proteins, persuaded her that the crystallography of HLA proteins was a dynamite project. Don, who had only recently been promoted to professor, and I had talked about this project casually over the years. He was interested in surface glycoproteins and had been working on inﬂuenza hemagglutinin so it was a natural extension of his interest. Despite the small amounts of HLA protein that could be obtained from cultured LCL and the huge effort required, it seemed like an attractive project to Pam, so at the end of the year she elected to begin her thesis work in my lab with the view of doing the crystal structure in Don’s lab if she succeeded in crystallizing one of these proteins. One other interesting event is worth noting in terms of the forces that guide science in our country now. My NIH research grant to work on MHC proteins was up for its ﬁve-year renewal. The main focus of the renewal application was the three-dimensional structure of HLA proteins. It was turned down by Study Section (now called Scientiﬁc Review Group) with a score of around 275, the lowest score I had received in 25 years of applications for research work on the penicillin problem, the HLA problem, and other projects I describe below. This grant at that time was the main support of my lab, and if it was not funded I would have to close up shop. At this point I learned that Study Sections do not make funding decisions. They only make recommendations to Institute Council, who together with the institute director make the funding decisions. I appealed to Council and asked them to overrule the Study Section. They did not, but instead they funded the grant for one additional year and asked me to revise the application and resubmit. What was the basis for the Study Section score? My memory is a little dim 25 years later, but I do remember two things. One was a

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reverberation of the review of my ﬁrst application to work on HLA proteins (the amount that could be obtained was too small to make crystallization and subsequent analysis by Xray crystallography feasible). Neither I nor the reviewers could know that the techniques for X-ray crystallography would improve dramatically in the next few years. The other criticism that I remember was that the application was not sufﬁciently well written or focused. I went back and looked at the application, of course, and it did not look that bad to me. I did not see much to change, but I did go through it and improved its language and focus. I also asked on resubmission that it be reviewed by a different Study Section. The new Study Section gave it a dramatically different score, in my memory around 110, well within the funding range at that time. The message is that even an established investigator has difﬁculty being funded when he wants to branch out and do something new, different, and difﬁcult. If established investigators are not encouraged to undertake important, difﬁcult problems, then who is going to tackle them? Something needs to be done to improve the system in that respect for both junior and senior investigators. Pamela Bjorkman worked in my lab for about a year or a year and a half. By that time, she had crystals of HLA-A2 (MAC, the ﬁrst polymorphic leukocyte antigen that had been detected by Jean Dausset). The crystals were small, thin, and fragile. She then moved to the Wiley lab to learn X-ray crystallography, and she completed her thesis as a joint student with Don and me. The thin crystals did not do well in a conventional X-ray source, i.e., they disintegrated so that a complete data set could not be obtained. However, sufﬁcient information was obtained for a ﬁrst publication (59). Importantly, the data showed that the molecule had approximate twofold rotational symmetry. In the illustration that Jim Kaufman created comparing MHC class I and MHC class II proteins, only one of several possible alignments for the three domains of the heavy chain of the MHC class I protein was shown, the one favored by Jim that made

MHC class I look much like MHC class II. For example, in one alignment not shown in Jim’s illustration, the three domains of the heavy chain would be linear. Pam’s initial Xray data showing twofold rotational symmetry supported Jim’s guess and certainly informed the illustration. The next important event was the advent of synchrotron radiation sources for protein crystallography. One of the ﬁrst available was DESY at Hamburg, Germany, so Pamela and Bill Bennett, one of Don’s postdocs, made frequent transatlantic trips to collect data there. Later, data were collected at the Cornell High Energy Synchrotron Source (CHESS) when it became available for protein crystallography, which of course was much more convenient. High-intensity X-rays produced by a synchrotron source were much less destructive to the small, thin crystals of HLA-A2, so complete native and heavy atom derivative data sets could be obtained, and, with important help from other members of Don’s laboratory, particularly Mark Saper, the structure was solved (60). One seemingly trivial but important problem was also solved. The technique for identifying amino acids at each step of degradation of a protein or peptide was not as precise in the late 1970s as it is today, and a few errors in the sequence were corrected by the sequence obtained from a genomic clone (61). The X-ray data ﬁt the sequence that was determined. It was a bombshell, now so well known to most of you that it does not need any description other than the structure that I show in Figure 2. Two striking features of the structure are the “extra density” that lies in a cleft between the two α-helices, a mixture of foreign and self peptides that are presented to the immune system and the location of polymorphic residues. The presence of the extra density and the need to establish which part of the X-ray diffraction pattern was due to the extra density and which part to the main chain of the HLA-A2 molecule were the last and most difﬁcult parts of the structure to establish. Nearly all the polymorphic residues that www.annualreviews.org • My Scientific Journey

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characterized MHC class I sequences were pointing into the peptide-binding cleft (62)! Thus, it was clear that the polymorphism was designed to establish in the population a variety of MHC proteins that could bind a very

large number of peptides. The evolutionary force that had driven it was the need to protect the population as a whole from catastrophic variation in an infectious agent that might otherwise decimate the population, or from differences in the geographic distribution of infectious agents. Next, Tom Garrett, along with Pam and Mark, solved the structure of HLA-Aw68, which differed from HLA-A2 by only 13 residues and demonstrated the existence of speciﬁcity pockets that accommodate the side chains of peptide antigens (anchor residues) (63). Joan Gorga solubilized and crystallized HLA-B27, and its structure was solved by Dean Madden (64, 65). It was notable because ˚ permitthe high-resolution data set (2.1 A) ted details of the mode of peptide binding to ←−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−− Figure 2 Structures of class I and class II MHC proteins as determined by 1994. MHC structures are in the left column and MHC class II are in the right column. (a) Ribbon diagrams are shown. The clefts between the two helices at the top are the sites of peptide binding (60, 62, 69, 71). (b–e) Peptides in the clefts of class I and class II MHC proteins viewed from the top and looking down into the clefts. (b) van der Waals surface representation of the mixture of peptides in the clefts of HLA-A2 and HLA-DR1 (60, 69, 106). (c) Single peptide complexes of the inﬂuenza nucleoprotein peptide 91–99 in the cleft of HLA-A68 and of the inﬂuenza hemagglutinin peptide 306–318 in the cleft of HLA-DR1 (66, 71). (d) Side views show the location of anchor residues of the peptides that extend into pockets in the structure of both classes of molecules. Those at P1 and P9 in both class I and class II proteins are particularly evident, but other anchor residues in each class are also important. (e) Mechanism of peptide binding to class I and class II MHC proteins. Hydrogen bonds with the main chains of the bound peptides are shown: class I (HLA-A2) and class II (HLA-DR1). Note that in class I hydrogen bonds are located at the two ends, whereas in class II they are distributed all along the length of the peptide (107–109). These ﬁgures are the result of 15 years of collaboration between members of Don Wiley’s laboratory and my laboratory between 1979 and 1994. Don died accidentally in 2001, and this ﬁgure honors his memory.

20

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Table 2 Yields of purified MHC antigensa Antigen

Yield mg/10 g cells

DR

12

DQ

2

DP

0.2

Class I

0.5

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a

First published in Reference 67.

be described. The single-peptide complex of HLA-Aw68 binding an inﬂuenza virus peptide was also solved (66). While all of this work was going on, people in my lab were carefully examining the possibility of crystallizing a class II MHC protein. The ﬁrst problem was to separate the three isotypes, HLA-DR, -DQ, and -DP. Immunoafﬁnity chromatography and monoclonal antibodies speciﬁc for each of the isotypes made that possible (67). Fortunately, the yield of the major MHC class II isotype HLA-DR was much greater than that of MHC class I isotypes (Table 2), but HLA-DR was resistant to papain proteolysis. After that problem was solved, seven allotypes of HLADR were crystallized by Joan Gorga (68). All the crystals were the same, relatively small and fragile, and HLA-DR1 was chosen for further study, probably because it had the number 1. Progress was again slow, but eventually a structure was obtained from data assembled from three different types of crystal, the native molecule (Jerry Brown), a superantigen complex of it (Ted Jardetzky), and a single peptide complex of hemagglutinin 306–318 bound to HLA-DR1 that had been obtained in the meantime using recombinant DNA technology to produce it (Larry Stern) (all three in Don’s lab) (69–71) (Figure 2). There are many differences between MHC class II and MHC class I proteins, but in outline the molecules look remarkably similar. The major difference is that the peptide-binding cleft is open at both ends in MHC class II, so that the peptide extends through the cleft, consistent with the fact that long peptides had been obtained after denaturation of HLA-DR pro-

teins and sequenced (71a,b). Thirteen amino acids lay within the cleft, the central nine residues being bound by anchor residues in pockets (as is true for the MHC class I proteins) and by substantial hydrogen bonds between the peptide backbone and the MHC class II protein. Much more work followed on the structures of both class I and class II MHC proteins, both in our laboratories and those of others, but none of it changed the outlines in the structures that had been determined. I am often asked how Don and I interacted during these exciting years. Don and I had very different styles and interests, and maybe that is why the collaboration was so successful. He was very focused and precise, and I was much more interested in where the problem was going to go. In the beginning, Don wanted me to work directly with him and learn to be a crystallographer, but he may have soon realized that I was a slow learner. I knew some of the language from Max Perutz’ course, and I learned much, much more from Don, his students, and his post-docs. However, I focused on the proteins. After Pam Bjorkman, Mark Saper, Tom Garrett, and others solved the structures of HLA-A2 and HLA-Aw68, products of alleles at the HLA-A locus, Joan Gorga solubilized and crystallized HLA-B27, and Dean Madden (in Don’s lab) solved its structure. Joan, together with Vaclav Horejsi and David Johnson, separated the three MHC class II isotypes and then solubilized and crystallized the HLA-DR proteins. Jerry Brown, Ted Jardetsky, and Larry Stern (all in Don’s lab) did the difﬁcult structure determination that was ﬁnally solved by combining data from the three different crystals. Don became one of the outstanding structural biologists in the world, and I was very fortunate to have him as a natural collaborator within the department. Don’s ofﬁce and lab were on the ﬁrst ﬂoor of the building, and mine were on the fourth. When I came into the building in the morning and passed the door that led to his lab and ofﬁce, I would often stop for a few minutes, most often with nothing particular in mind, but www.annualreviews.org • My Scientific Journey

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sometimes with some speciﬁc question. I always came away stimulated and with new insights. Don died accidentally in 2001 (71c). He left an enormous intellectual hole for me and for our department. I loved the way he did science. My own interests turned after that to attempting to use the structural information to further our understanding of autoimmune diseases. Nearly all these diseases are linked to speciﬁc allotypes of class II proteins (72). An important drug for the treatment of multiple sclerosis (MS) (Copolymer 1, Copaxone® , Glatiramer acetate) was developed by Michael Sela, Ruth Arnon, and their colleagues in Israel (73) and is in wide use for treatment today. Although it reduces the frequency of relapses by only 30%, this reduction is highly signiﬁcant for patients with this disease. This disease is linked to HLA-DR2, and by using the structural information obtained for the manner in which MBP(85−99) (a myelin basic protein region that is thought to be encephalitogenic in MS and is thus a major candidate autoantigen for the disease) is bound to the MHC class II protein, new copolymers were designed that were more effective in binding to HLA-DR2 and in inhibiting MBP(85−99) -speciﬁc, HLA-DR2-restricted T cell lines. Importantly, they are also more effective in ameliorating experimental autoimmune encephalomyelitis, the rodent model of MS, using several different mouse models and modes of administration of the copolymers (74–77). However, mice are not humans, and only clinical trials will answer the question whether these new molecules are improvements over Copolymer 1. Work on these materials is ongoing.

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NATURAL KILLER CELLS Soon after the identiﬁcation of many new genes in the class III region of the human MHC, Moretta and colleagues, who had been studying human NK cells, reported that a resistance gene that regulated NK cell activity was linked to the MHC (78). They sug22

Strominger

gested that we collaborate in identifying this gene, believing that it might be one of the newly discovered genes. However, immunogenetic studies established that the resistance gene was located in the large 2-mb region between HLA-B and HLA-A (79). Although several genes had been identiﬁed in this region, only one of them, HLA-C, was known to be functional and expressed on the cell surface. Moretta and colleagues had also cloned NK cells from human peripheral blood and found that the two dominant sets of clones were mutually exclusive in their ability to lyse NK targets, i.e., cells that were lysed by group 1 clones were not lysed by group 2 clones and vice versa (80). A few target cells were resistant to both group I and group II clones. Marco Colonna, then a postdoctoral fellow, noticed that HLA-C was also dimorphic (81). One group of HLA-C allotypes had serine at position 77 and asparagine at position 80, whereas the second group of allotypes had asparagine 77 and lysine 80. He quickly established the relationship between the two groups of HLAC allotypes and the two groups of human NK clones, i.e., NK 1 clones were inhibited by members of HLA-C allotype group II, and NK 2 clones were inhibited by members of HLA-C allotype group molecules (82). Later, residue 80 was shown to be critical for the inhibitory function of HLA-C (83). These studies greatly clariﬁed how NK clones recognize (or fail to recognize) target cells. Later, Marco cloned the NK inhibitory receptors, now called KIR2DL1, 2, or 3 (as did another group) (84, 85). He also showed that the serological speciﬁcities Bw4 and Bw6 that had been deﬁned by van Rood and his colleagues corresponded to a similar dimorphism of HLA-B allotypes at residue 80 that were related to inhibition of lysis by other types of NK clones (86). The receptor for this inhibition, KIR3DL1, was subsequently cloned (87). My laboratory has continued to be involved in studies of human NK cells, principally in trying to understand the formation and structure of inhibitory and activating NK synapses and their relationships to each other

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(88, 89; K. Krzewski, X. Chen, J.S. Orange, and J. Strominger, manuscript submitted). In addition, we are engaged in studies of human decidual NK cells, which represent 70%–90% of the lymphocytes found in the uterine decidua during pregnancy (91–93). These cells are likely involved in maternal fetal tolerance. The enigma of acceptance of the hemiallogeneic fetal graft by the pregnant female was pointed out by Peter Medawar more than 50 years ago, and its explanation is still incomplete. Both of these topics also represent current research projects in my laboratory.

THE DANA-FARBER CANCER INSTITUTE, 1974-PRESENT Soon after I moved to Harvard in 1968, Emil (Tom) Frei, my classmate at Yale Medical School and a close friend, became the chief of medicine of what was then called the Children’s Cancer Research Foundation, founded by Sidney Farber. Dr. Farber was the director, but he died a few years later, and Tom then became the director. A large new research building that included a small research hospital (the Dana Building) was under construction. It was initially called the Sidney Farber Cancer Institute, but the name was later changed to the Dana-Farber Cancer Institute as a result of a large contribution by the Dana Foundation. The new building had several hundred thousand square feet of research space, and stafﬁng it was a huge opportunity and responsibility. Around that time, I had begun to work on EBV, which had been discovered only a few years earlier as an agent closely associated with Burkitt’s lymphoma, a disease endemic in the malaria belt of Africa. EBV was soon found in sporadic cases in the United States, in endemic malarial regions of the world, and in other regions. EBV was particularly useful in research because it transformed human lymphocytes in vitro and thus provided a means of establishing cell lines that express a variety of HLA haplotypes, including homozygous cell lines. EBV is a very large herpes virus, and the mechanism by which it transformed lym-

phocyte was unknown. One of my graduate students, Roger Yocum, thought it would be interesting to study, and I thought it would be an interesting diversiﬁcation of our work. However, soon after we began Jim Watson, then a member of the department, became concerned that growing the virus in our relatively poor facilities could be dangerous, even though the virus is found in the throats of nearly all individuals. Neither the potential for increased tumorigenicity as a result of virus variation nor the potential for large doses of virus in aerosols to produce disease were well understood. He was uncomfortable with my growing it in our building. I then asked Tom whether he had a small lab I could use for growing EBV at the Farber, as the institute is called. He responded by asking me to become the institute’s director of Basic Sciences and to help him get it off to a good start. After some discussion, I agreed to do it for three years. It was not an easy job. Sidney Farber had not been especially careful in appointing staff, very few of whom had Harvard appointments. One ground rule we established was that all future appointments would need to be Harvard faculty appointments. There was also tension between excellent clinical scientists and the basic scientists that Tom and I both wanted to bring into the institute. I had, and still have, the strong view that clinical science prospers in the presence of outstanding basic science, and vice versa. I am very proud of the excellent trajectory for science that was established by the appointments made in those three years. That trajectory has been maintained until the present time, so that the Farber has become an outstanding scientiﬁc institution as well as an outstanding center for clinical research and treatment. At the end of three years, I returned full-time to my position in Cambridge, Massachusetts, but Tom asked me to keep a laboratory at the Farber, and excellent work was carried out there by postdoctoral fellows, some of whom became faculty members. Notable among those contributions were the www.annualreviews.org • My Scientific Journey

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cloning of the EBV genome (Jim Skare) (94); the identiﬁcation of a very large polycistronic message derived from EBV and its differential splicing, together with the subsequent identiﬁcation of the complex promoter usage by EBV (Sam Speck) (95); the identiﬁcation of C3d as the EBV receptor (Joyce Fingeroth) (96); the identiﬁcation of VLA (very late antigens, now called the β1-integrin family of molecules) by Martin Hemler (97); the identiﬁcation of a second T cell receptor, the γδ T cell receptor, by Mike Brenner and Mike Krangel (98); the identiﬁcation of the two subunits of the TAP transporter as well as the ﬁrst genetic map of the entire MHC, including the identiﬁcation of many new genes by Thomas Spies (99); and most recently the discovery of the role of caspases and nitric oxide in the maturation of dendritic cells and the light this discovery shed on the intracellular transport of proteins, including MHC class II, by Laura Santambrogio and Siew-Heng Wong (100, 101). An extremely fruitful and pleasant collaboration with Steve Burakoff resulted in the ﬁrst suggestion that MHC class II–restricted T cell clones are CD4+ , whereas MHC class I–restricted clones are CD8+ (102). This collaboration was my introduction to cellular immunology, and I learned an enormous amount.

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NIH R35 CA 47554 (OUTSTANDING INVESTIGATOR GRANT) Having taken several digs at NIH several times above, I have to conclude by expressing my deep appreciation of the support I have received over my career, most speciﬁcally the support of the Outstanding Investigator Grant. In 1976, when I ﬁnally had organized a laboratory at the Farber, I had seven or eight R01 grants to support the two laboratories. An enormous amount of time was being spent on writing up competing renewals and noncompeting renewals. I learned about the OIG awards and applied. When I re-

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ceived the award, program ofﬁcers at the National Cancer Institute (NCI) were extremely generous. It was an umbrella grant that replaced all my existing grants, including those from other institutes and my EBV grant from NCI. The yearly noncompeting renewal process was greatly simpliﬁed, and after seven years the competing renewal was a review of progress made under that grant by a panel constituted for that purpose. In many ways, the OIGs were equivalent to the investigatorships offered by the Howard Hughes Medical Institute, but they were not encumbered by the necessity of attending annual meetings or including NCI in the title lines of papers as one of the institutions from which the work had come. My grant could not have been administered in a better way. After 13 years, however, NCI terminated the program. I am not sure why. I believe that programs of this type would greatly simplify the administrative burdens now placed on investigators. The OIG and NCI played a very large part in my success. Immunoland has truly been a wonderland for me since I entered the ﬁeld as a biochemist in about 1970. During this period, two extremely important immunological tools were introduced, the generation of monoclonal antibodies by George Kohler and Cesar Milstein (103) and the development of the ﬂuorescence activated cell sorter (FACS) by Leonard Herzenberg (104). Without these inventions, progress would have been much, much slower. Developmental processes, cellcell interactions, signaling processes, and biochemical mechanisms in the immune system were all made more accessible to investigation during the period that I worked in this ﬁeld. I was very fortunate to be there at the beginning. I conclude by thanking and expressing my deep gratitude to the many postdoctoral fellows, graduate and undergraduate students, research assistants, administrative assistants, laboratory aides, and collaborators who worked with me during this adventure.

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LITERATURE CITED 1. Strominger J. 1948. A national science foundation. Yale. J. Biol. Med. 20:577 2. Park JT. 1952. Uridine-5 -pyrophosphate derivatives. II. Isolation from Staphylococcus aureus. J. Biol. Chem. 194:877–84 3. Reissig JL, Strominger JL, Leloir LF. 1955. A modiﬁed colorimetric method for the estimation of N-acetylamino sugars. J. Biol. Chem. 217:959–66 4. Strominger JL. 1957. Microbial uridine-5 -pyrophosphate N-acetylamino sugar compounds. I. Biology of the penicillin-induced accumulation. J. Biol. Chem. 224:509– 23 5. Strominger JL, Heppel LA, Maxwell ES. 1959. Nucleoside monophosphate kinases. I. Transphosphorylation between adenosine triphosphate and nucleoside monophosphates. Biochim. Biophys. Acta 32:412–21 6. Strominger JL, Kalckar HM, Axelrod J, Maxwell ES. 1954. Enzymatic oxidation of uridine diphosphate glucose to uridine diphosphate glucuronic acid. J. Am. Chem. Soc. 76: 6411 7. Anderson EP, Isselbacher KJ, Kalckar HM. 1957. Defect in uptake of galactose-1phosphate into liver nucleotides in congenital galactosemia. Science 125:113–14 8. Strominger JL. 1959. The amino acid sequence in the uridine nucleotide-peptide from Staphylococcus aureus. Comp. Rend. Trav. Lab. Carlsberg 31:181 9. Park JT, Strominger JL. 1957. Mode of action of penicillin. Science 125:99–101 10. Strominger JL, Park JT, Thompson RE. 1959. Composition of the cell wall of Staphylococcus aureus: its relation to the mechanism of action of penicillin. J. Biol. Chem. 234:3263–68 11. Nathenson SG, Strominger JL. 1961. Effects of penicillin on the biosynthesis of the cell walls of Escherichia coli and Staphylococcus aureus. J. Pharmacol. Exp. Ther. 131:1–6 12. Strominger JL, Ghuysen JM. 1967. Mechanisms of enzymatic bacteriaolysis. Cell walls of bacteria are solubilized by action of either speciﬁc carbohydrases or speciﬁc peptidases. Science 156:213–21 13. Strominger JL. 1970. Penicillin-sensitive enzymatic reactions in bacterial cell wall synthesis. In Harvey Lectures, pp. 179–213. New York: Academic 14. Meadow PM, Anderson JS, Strominger JL. 1964. Enzymatic polymerization of UDPacetylmuramyl.l-ala.d-glu.l-lys.d-ala.d-ala and UDP-acetylglucosamine by a particulate enzyme from Staphylococcus aureus and its inhibition by antibiotics. Biochem. Biophys. Res. Commun. 14:382–87 15. Strominger JL, Okazaki T, Okazaki R. 1963. Oxidation and reduction of nucleotidelinked sugars. In The Enzymes, ed. P Boyer, H Lardy, K Myrback, p. 161. New York: Academic 16. Kobayashi KS, Chamaillard M, Ogura Y, Henegariu O, Inohara N, et al. 2005. Nod2dependent regulation of innate and adaptive immunity in the intestinal tract. Science 307:731–34 17. Juergens WG, Sanderson AR, Strominger JL. 1963. Chemical basis for an immunological speciﬁcity of a strain of Staphylococcus aureus. J. Exp. Med. 117:925–35 18. Krueger L, Luederitz O, Strominger JL, Westphal O. 1962. [On the immunochemistry of O antigens of Enterobacteriaceae. VII. The relation of hexoses and 6-desoxyhexoses in Salmonella lipopolysaccharides to the D and L group.]. Biochem Z 335:548–58 19. Izaki K, Matsuhashi M, Strominger JL. 1966. Glycopeptide transpeptidase and D-alanine carboxypeptidase: penicillin-sensitive enzymatic reactions. Proc. Natl. Acad. Sci. USA 55:656–63

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20. Tipper DJ, Strominger JL. 1965. Mechanism of action of penicillins: a proposal based on their structural similarity to acyl-D-alanyl-D-alanine. Proc. Natl. Acad. Sci. USA 54:1133– 41 21. Lee W, McDonough MA, Kotra L, Li ZH, Silvaggi NR, et al. 2001. A 1.2-A snapshot of the ﬁnal step of bacterial cell wall biosynthesis. Proc. Natl. Acad. Sci. USA 98:1427– 31 22. Anderson JS, Matsuhashi M, Haskin MA, Strominger JL. 1965. Lipid-Phosphoacetylmuramyl-pentapeptide and lipid-phosphodisaccharide-pentapeptide: presumed membrane transport intermediates in cell wall synthesis. Proc. Natl. Acad. Sci. USA 53:881– 89 23. Higashi Y, Strominger JL, Sweeley CC. 1967. Structure of a lipid intermediate in cell wall peptidoglycan synthesis: a derivative of a C55 isoprenoid alcohol. Proc. Natl. Acad. Sci. USA 57:1878–84 24. Matsuhashi M, Strominger JL. 1964. Thymidine diphosphate 4-acetamido-4, 6dideoxyhexoses. I. Enzymatic synthesis by strains of Escherichia coli. J. Biol. Chem. 239:2454–63 25. Roberts WS, Petit JF, Strominger JL. 1968. Biosynthesis of the peptidoglycan of bacterial cell walls. 8. Speciﬁcity in the utilization of L-alanyl transfer ribonucleic acid for interpeptide bridge synthesis in Arthrobacter crystallopoietes. J. Biol. Chem. 243:768–72 26. Strominger JL, Threnn RH. 1959. Accumulation of a uridine nucleotide in Staphylococcus aureus as the consequence of lysine deprivation. Biochim. Biophys. Acta 36:83–92 27. Adam A, Petit JF, Lefrancier P, Lederer E. 1981. Muramyl peptides. Chemical structure, biological activity and mechanism of action. Mol. Cell. Biochem. 41:27–47 28. Janeway CA Jr, Medzhitov R. 2002. Innate immune recognition. Annu. Rev. Immunol. 20:197–216 29. Gorer PA. 1937. The genetic and antigenic basis for tumor transplantation. J. Pathol. Bacteriol. 44:691–97 30. Snell GD, Lyman S, Gorer PA. 1948. Studies on the genetic and antigenetic basis of tumor transplantation. Linkage between a histocompatibility gene and “fused” in mice. Proc. R. Soc. London Ser. B Biol. Sci. 135:499–505 31. Nathenson SG, Davies DA. 1966. Solubilization and partial puriﬁcation of mouse histocompatibility antigens from a membranous lipoprotein fraction. Proc. Natl. Acad. Sci. USA 56:476–83 32. Sanderson AR, Strominger JL, Nathenson SG. 1962. Chemical structure of teichoic acid from Staphylococcus aureus, strain Copenhagen. J. Biol. Chem. 237:3603–13 33. Sanderson AR, Batchelor JR. 1968. Transplantation antigens from human spleens. Nature 219:184–86 34. Turner MJ, Cresswell P, Parham P, Strominger JL, Mann DL, Sanderson AR. 1975. Puriﬁcation of papain-solubilized histocompatibility antigens from a cultured human lymphoblastoid line, RPMI 4265. J. Biol. Chem. 250:4512–19 35. Cresswell P, Turner MJ, Strominger JL. 1973. Papain-solubilized HL-A antigens from cultured human lymphocytes contain two peptide fragments. Proc. Natl. Acad. Sci. USA 70:1603–7 36. Grey HM, Kubo RT, Colon SM, Poulik MD, Cresswell P, et al. 1973. The small subunit of HL-A antigens is beta 2-microglobulin. J. Exp. Med. 138:1608–12 37. Cresswell P, Springer T, Strominger JL, Turner MJ, Grey HM, Kubo RT. 1974. Immunological identity of the small subunit of HL-A antigens and beta2-microglobulin and its turnover on the cell membrane. Proc. Natl. Acad. Sci. USA 71:2123–27

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38. Springer TA, Strominger JL. 1976. Detergent-soluble HLA antigens contain a hydrophilic region at the COOH-terminus and a penultimate hydrophobic region. Proc. Natl. Acad. Sci. USA 73:2481–85 39. Orr HT, Lancet D, Robb RJ, Lopez de Castro JA, Strominger JL. 1979. The heavy chain of human histocompatibility antigen HLA-B7 contains an immunoglobulin-like region. Nature 282:266–70 40. Orr HT, Lopez de Castro JA, Lancet D, Strominger JL. 1979. Complete amino acid sequence of a papain-solubilized human histocompatibility antigen, HLA-B7. 2. Sequence determination and search for homologies. Biochemistry 18:5711–20 41. Humphreys RE, McCune JM, Chess L, Herrman HC, Malenka DJ, et al. 1976. Isolation and immunologic characterization of a human B-lymphocyte-speciﬁc, cell surface antigen. J. Exp. Med. 144:99–112 42. Kaufman JF, Auffray C, Korman AJ, Shackelford DA, Strominger J. 1984. The class II molecules of the human and murine major histocompatibility complex. Cell 36:1–13 43. Shackelford DA, Mann DL, van Rood JJ, Ferrara GB, Strominger JL. 1981. Human B-cell alloantigens DC1, MT1, and LB12 are identical to each other but distinct from the HLA-DR antigen. Proc. Natl. Acad. Sci. USA 78:4566–70 44. Ploegh HL, Orr HT, Strominger JL. 1980. Molecular cloning of a human histocompatibility antigen cDNA fragment. Proc. Natl. Acad. Sci. USA 77:6081–85 45. Orr HT, Bach FH, Ploegh HL, Strominger JL, Kavathas P, DeMars R. 1982. Use of HLA loss mutants to analyse the structure of the human major histocompatibility complex. Nature 296:454–56 46. Koller BH, Geraghty DE, DeMars R, Duvick L, Rich SS, Orr HT. 1989. Chromosomal organization of the human major histocompatibility complex class I gene family. J. Exp. Med. 169:469–80 47. Korman AJ, Knudsen PJ, Kaufman JF, Strominger JL. 1982. cDNA clones for the heavy chain of HLA-DR antigens obtained after immunopuriﬁcation of polysomes by monoclonal antibody. Proc. Natl. Acad. Sci. USA 79:1844–48 48. Korman AJ, Auffray C, Schamboeck A, Strominger JL. 1982. The amino acid sequence and gene organization of the heavy chain of the HLA-DR antigen: homology to immunoglobulins. Proc. Natl. Acad. Sci. USA 79:6013–17 49. Auffray C, Kuo J, DeMars R, Strominger JL. 1983. A minimum of four human class II alpha-chain genes are encoded in the HLA region of chromosome 6. Nature 304:174–77 50. Roux-Dosseto M, Auffray C, Lillie JW, Boss JM, Cohen D, et al. 1983. Genetic mapping of a human class II antigen beta-chain cDNA clone to the SB region of the HLA complex. Proc. Natl. Acad. Sci. USA 80:6036–40 51. Steinmetz M, Minard K, Horvath S, McNicholas J, Srelinger J, et al. 1982. A molecular map of the immune response region from the major histocompatibility complex of the mouse. Nature 300:35–42 52. Okada K, Boss JM, Prentice H, Spies T, Mengler R, et al. 1985. Gene organization of DC and DX subregions of the human major histocompatibility complex. Proc. Natl. Acad. Sci. USA 82:3410–14 53. Spies T, Sorrentino R, Boss JM, Okada K, Strominger JL. 1985. Structural organization of the DR subregion of the human major histocompatibility complex. Proc. Natl. Acad. Sci. USA 82:5165–69 54. Spies T, Morton CC, Nedospasov SA, Fiers W, Pious D, Strominger JL. 1986. Genes for the tumor necrosis factors α and β are linked to the human major histocompatibility complex. Proc. Natl. Acad. Sci. USA 83:8699–702 www.annualreviews.org • My Scientific Journey

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55. Carroll MC, Katzman P, Alicot EM, Koller BH, Geraghty DE, et al. 1987. Linkage map of the human major histocompatibility complex including the tumor necrosis factor genes. Proc. Natl. Acad. Sci. USA 84:8535–39 56. Spies T, Blanck G, Bresnahan M, Sands J, Strominger JL. 1989. A new cluster of genes within the human major histocompatibility complex. Science 243:214 –17 57. Bahram S, Spies T. 1996. The MIC gene family. Res. Immunol. 147:328–33 58. Orr HT, Lopez de Castro JA, Parham P, Ploegh HL, Strominger JL. 1979. Comparison of amino acid sequences of two human histocompatibility antigens, HLA-A2 and HLA-B7: location of putative alloantigenic sites. Proc. Natl. Acad. Sci. USA 76:4395– 99 59. Bjorkman PJ, Strominger JL, Wiley DC. 1985. Crystallization and X-ray diffraction studies on the histocompatibility antigens HLA-A2 and HLA-A28 from human cell membranes. J. Mol. Biol. 186:205–10 60. Bjorkman PJ, Saper MA, Samraoui B, Bennett WS, Strominger JL, Wiley DC. 1987. Structure of the human class I histocompatibility antigen, HLA-A2. Nature 329:506– 12 61. Koller BH, Orr HT. 1985. Cloning and complete sequence of an HLA-A2 gene: analysis of two HLA-A alleles at the nucleotide level. J. Immunol. 134:2727–33 62. Bjorkman PJ, Saper MA, Samraoui B, Bennett WS, Strominger JL, Wiley DC. 1987. The foreign antigen binding site and T cell recognition regions of class I histocompatibility antigens. Nature 329:512–18 63. Garrett TP, Saper MA, Bjorkman PJ, Strominger JL, Wiley DC. 1989. Speciﬁcity pockets for the side chains of peptide antigens in HLA-Aw68. Nature 342:692–96 64. Gorga JC, Madden DR, Prendergast JK, Wiley DC, Strominger JL. 1992. Crystallization and preliminary X-ray diffraction studies of the human major histocompatibility antigen HLA-B27. Proteins 12:87–90 65. Madden DR, Gorga JC, Strominger JL, Wiley DC. 1992. The three-dimensional structure of HLA-B27 at 2.1 A˚ resolution suggests a general mechanism for tight peptide binding to MHC. Cell 70:1035–48 66. Silver ML, Guo HC, Strominger JL, Wiley DC. 1992. Atomic structure of a human MHC molecule presenting an inﬂuenza virus peptide. Nature 360:367–69 67. Gorga JC, Horejsi V, Johnson DR, Raghupathy R, Strominger JL. 1987. Puriﬁcation and characterization of class II histocompatibility antigens from a homozygous human B cell line. J. Biol. Chem. 262:16087–94 68. Gorga JC, Brown JH, Jardetzky T, Wiley DC, Strominger JL. 1991. Crystallization of HLA-DR antigens. Res. Immunol. 142:401–7 69. Brown JH, Jardetzky TS, Gorga JC, Stern LJ, Urban RG, et al. 1993. Three-dimensional structure of the human class II histocompatibility antigen HLA-DR1. Nature 364:33– 39 70. Jardetzky TS, Brown JH, Gorga JC, Stern LJ, Urban RG, et al. 1994. Three-dimensional structure of a human class II histocompatibility molecule complexed with superantigen. Nature 368:711–18 71. Stern LJ, Brown JH, Jardetzky TS, Gorga JC, Urban RG, et al. 1994. Crystal structure of the human class II MHC protein HLA-DR1 complexed with an inﬂuenza virus peptide. Nature 368:215–21 71a. Chicz RM, Urban RG, Lane WS, Gorga JC, Stern LJ, et al. 1992. Predominant naturally processed peptides bound to HLA-DR1 are derived from MHC-related molecules and are heterogeneous in size. Nature 358:764–68

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71b. Chicz RM, Urban RG, Gorga JC, Vignali DAA, Lane WS, Strominger JL. 1993. Speciﬁcity and promiscuity among naturally processed peptides bound to HLA-DR alleles. J. Exp. Med. 178:27–47 71c. Strominger JL. 2002. Don Craig Wiley (1944–2001): a reminiscence. Nat. Immunol. 2:103–4 72. Svejgaard A, Platz P, Ryder LP. 1983. HLA and disease 1982—a survey. Immunol. Rev. 70:193–218 73. Teitelbaum D, Webb C, Meshorer A, Arnon R, Sela M. 1973. Suppression by several synthetic polypeptides of experimental allergic encephalomyelitis induced in guinea pigs and rabbits with bovine and human basic encephalitogen. Eur. J. Immunol. 3:273–79 74. Fridkis-Hareli M, Santambrogio L, Stern JN, Fugger L, Brosnan C, Strominger JL. 2002. Novel synthetic amino acid copolymers that inhibit autoantigen-speciﬁc T cell responses and suppress experimental autoimmune encephalomyelitis. J. Clin. Invest. 109:1635–43 75. Stern JN, Illes Z, Reddy J, Keskin DB, Sheu E, et al. 2004. Amelioration of proteolipid protein 139–151-induced encephalomyelitis in SJL mice by modiﬁed amino acid copolymers and their mechanisms. Proc. Natl. Acad. Sci. USA 101:11743–48 76. Illes Z, Stern JN, Reddy J, Waldner H, Mycko MP, et al. 2004. Modiﬁed amino acid copolymers suppress myelin basic protein 85–99-induced encephalomyelitis in humanized mice through different effects on T cells. Proc. Natl. Acad. Sci. USA 101:11749–54 77. Stern JN, Illes Z, Reddy J, Keskin DB, Fridkis-Hareli M, et al. 2005. Peptide 15-mers of deﬁned sequence that substitute for random amino acid copolymers in amelioration of experimental autoimmune encephalomyelitis. Proc. Natl. Acad. Sci. USA 102:1620–25 78. Ciccone E, Pende D, Viale O, Tambussi G, Ferrini S, et al. 1990. Speciﬁc recognition of human CD3-CD16+ natural killer cells requires the expression of an autosomic recessive gene on target cells. J. Exp. Med. 172:47–52 79. Ciccone E, Colonna M, Viale O, Pende D, Di Donato C, et al. 1990. Susceptibility or resistance to lysis by alloreactive natural killer cells is governed by a gene in the human major histocompatibility complex between BF and HLA-B. Proc. Natl. Acad. Sci. USA 87:9794–97 (and correction in Proc. Natl. Acad. USA 88:5477) 80. Ciccone E, Pende D, Viale O, Di Donato C, Tripodi G, et al. 1992. Evidence of a natural killer (NK) cell repertoire for (allo) antigen recognition: deﬁnition of ﬁve distinct NKdetermined allospeciﬁcities in humans. J. Exp. Med. 175:709–18 81. Colonna M, Spies T, Strominger JL, Ciccone E, Moretta A, et al. 1992. Alloantigen recognition by two human natural killer cell clones is associated with HLA-C or a closely linked gene. Proc. Natl. Acad. Sci. USA 89:7983–85 82. Colonna M, Borsellino G, Falco M, Ferrara GB, Strominger JL. 1993. HLA-C is the inhibitory ligand that determines dominant resistance to lysis by NK1- and NK2-speciﬁc natural killer cells. Proc. Natl. Acad. Sci. USA 90:12000–4 83. Mandelboim O, Reyburn HT, Vales-Gomez M, Pazmany L, Colonna M, et al. 1996. Protection from lysis by natural killer cells of group 1 and 2 speciﬁcity is mediated by residue 80 in human histocompatibility leukocyte antigen C alleles and also occurs with empty major histocompatibility complex molecules. J. Exp. Med. 184:913–22 84. Colonna M, Samaridis J. 1995. Cloning of immunoglobulin-superfamily members associated with HLA-C and HLA-B recognition by human natural killer cells. Science 268:405– 8 85. Wagtmann N, Biassoni R, Cantoni C, Verdiani S, Malnati MS, et al. 1995. Molecular clones of the p58 NK cell receptor reveal immunoglobulin-related molecules with diversity in both the extra- and intracellular domains. Immunity 2:439–49 www.annualreviews.org • My Scientific Journey

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86. Cella M, Longo A, Ferrara GB, Strominger JL, Colonna M. 1994. NK3-speciﬁc natural killer cells are selectively inhibited by Bw4-positive HLA alleles with isoleucine 80. J. Exp. Med. 180:1235–42 87. D’Andrea A, Chang C, Franz-Bacon K, McClanahan T, Phillips JH, Lanier LL. 1995. Molecular cloning of NKB1. A natural killer cell receptor for HLA-B allotypes. J. Immunol. 155:2306–10 88. Davis DM, Chiu I, Fassett M, Cohen GB, Mandelboim O, Strominger JL. 1999. The human natural killer cell immune synapse. Proc. Natl. Acad. Sci. USA 96:15062–67 89. Orange JS, Harris KE, Andzelm MM, Valter MM, Geha RS, Strominger JL. 2003. The mature activating natural killer cell immunologic synapse is formed in distinct stages. Proc. Natl. Acad. Sci. USA 100:14151–56 90. Deleted in proof 91. Orange JS, Fassett MS, Koopman LA, Boyson JE, Strominger JL. 2002. Viral evasion of natural killer cells. Nat. Immunol. 3:1006–12 92. Koopman LA, Kopcow HD, Rybalov B, Boyson JE, Orange JS, et al. 2003. Human decidual natural killer cells are a unique NK cell subset with immunomodulatory potential. J. Exp. Med. 198:1201–12 93. Kopcow HD, Allan DS, Ge B, Rybalov B, Andzelm MM, et al. 2005. Human decidual NK cells form immature activating synapses and are not cytotoxic. Proc. Natl. Acad. Sci. USA. 43:15563–68 94. Skare J, Strominger JL. 1980. Cloning and mapping of BamHi endonuclease fragments of DNA from the transforming B95-8 strain of Epstein-Barr virus. Proc. Natl. Acad. Sci. USA 77:3860–64 95. Schaefer BC, Woisetschlaeger M, Strominger JL, Speck SH. 1991. Exclusive expression of Epstein-Barr virus nuclear antigen 1 in Burkitt lymphoma arises from a third promoter, distinct from the promoters used in latently infected lymphocytes. Proc. Natl. Acad. Sci. USA 88:6550–54 96. Fingeroth JD, Weis JJ, Tedder TF, Strominger JL, Biro PA, Fearon DT. 1984. EpsteinBarr virus receptor of human B lymphocytes is the C3d receptor CR2. Proc. Natl. Acad. Sci. USA 81:4510–14 97. Hemler ME, Ware CF, Strominger JL. 1983. Characterization of a novel differentiation antigen complex recognize by a monoclonal antibody (A-1A5): unique activation-speciﬁc molecular forms on stimulated T cells. J. Immunol. 131:334–40 98. Brenner MB, McLean J, Dialynas DP, Strominger JL, Smith JA, et al. 1986. Identiﬁcation of a putative second T-cell receptor. Nature 322:145–49 99. Spies T, Bresnahan M, Strominger JL. 1989. Human major histocompatibility complex contains a minimum of 19 genes between the complement cluster and HLA-B. Proc. Natl. Acad. Sci. USA 86:8955–58 100. Wong SH, Santambrogio L, Strominger JL. 2004. Caspases and nitric oxide broadly regulate dendritic cell maturation and surface expression of class II MHC proteins. Proc. Natl. Acad. Sci. USA 101:17783–88 101. Santambrogio L, Potolicchio I, Fessler SP, Wong SH, Raposo G, Strominger JL. 2005. Involvement of caspase-cleaved and intact adaptor protein 1 complex in endosomal remodeling in maturing dendritic cells. Nat. Immunol. 6:1020–28 102. Krensky AM, Clayberger C, Reiss CS, Strominger JL, Burakoff SJ. 1982. Speciﬁcity of OKT4+ cytotoxic T lymphocyte clones. J. Immunol. 129:2001–3 103. Kohler G, Milstein C. 1975. Continuous cultures of fused cells secreting antibody of predeﬁned speciﬁcity. Nature 256:495–97

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104. Herzenberg LA, Herzenberg LA. 2004. Genetics, FACS, immunology, and redox: a tale of two lives intertwined. Annu. Rev. Immunol. 22:1–31 105. Ploegh HL, Orr HT, Strominger JL. 1981. Major histocompatibility antigens: the human (HLA-A, -B, -C) and murine (H-2K, H-2D) class I molecules. Cell 24:287–99 106. Chicz RM, Urban RG, Gorga JC, Vignali DA, Lane WS, Strominger JL. 1993. Speciﬁcity and promiscuity among naturally processed peptides bound to HLA-DR alleles. J. Exp. Med. 178:27–47 107. Stern LJ, Wiley DC. 1994. Antigenic peptide binding by class I and class II histocompatibility proteins. Structure 2:245–51 108. Madden DR. 1995. The three-dimensional structure of peptide-MHC complexes. Annu. Rev. Immunol. 13:587–622 109. Latron F, Pazmany L, Morrison J, Moots R, Saper MA, et al. 1992. A critical role for conserved residues in the cleft of HLA-A2 in presentation of a nonapeptide to T cells. Science 257:964–67

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Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh,1 Nacksung Kim,2 Yuho Kadono,1 Jaerang Rho,3 Soo Young Lee,4 Joseph Lorenzo,5 and Yongwon Choi1 1

Department of Pathology and Laboratory Medicine, AFCRI, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104; email: [email protected], [email protected], [email protected]

2

MRC for Gene Regulation, Chonnam National University Medical School, Gwangju 501–746, Korea; email: [email protected]

3

Department of Microbiology and RCTCG, Chungnam National University, Daejon 305–764, Korea; email: [email protected]

4

Division of Molecular Life Sciences, CCSR, Ewha Womans University, Seoul 120–750, Korea; email: [email protected]

5

Department of Medicine, University of Connecticut Health Center, Farmington, Connecticut 06030-1317; email: [email protected]

Annu. Rev. Immunol. 2006. 24:33–63 First published online as a Review in Advance on October 12, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090646 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0033$20.00

Key Words T lymphocyte, dendritic cell, osteoblast, osteoclast, TRANCE-RANK, costimulation

Abstract Studies of bone and the immune system have converged in recent years under the banner of osteoimmunology. The immune system is spawned in the bone marrow reservoir, and investigators now recognize that important niches also exist there for memory lymphocytes. At the same time, various factors produced during immune responses are capable of profoundly affecting regulation of bone. Mechanisms have evolved to prevent excessive interference by the immune system with bone homeostasis, yet pathologic bone loss is a common sequela associated with autoimmunity and cancer. There are also developmental links, or parallels, between bone and the immune system. Cells that regulate bone turnover share a common precursor with inﬂammatory immune cells and may restrict themselves anatomically, in part by utilizing a signaling network analogous to lymphocyte costimulation. Efforts are currently under way to further characterize how these two organ systems overlap and to develop therapeutic strategies that beneﬁt from this understanding. 33

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INTRODUCTION HSC: hematopoietic stem cell

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Osteoblast (OB): bone-producing cell that also provides signals for osteoclast formation Osteoclast (OC): specialized multinucleated giant cell that resorbs bone TNF: tumor necrosis factor TNF-related activation-induced cytokine (TRANCE): a TNF family cytokine that is essential for osteoclast development Osteoporosis: a condition characterized by decreased bone mass and density that causes bones to become fragile

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The emergence of the niche ﬁeld of osteoimmunology represents a conceptual rethinking of multiple phenomena, relating biological events in bone and the immune system (1). The root of exploration of this interplay begins with the basic understanding that bone provides a microenvironment that is critical for the development of the hematopoietic stem cells (HSCs), from which all cells of the mammalian immune system derive, and that various immunoregulatory cytokines inﬂuence the fate of bone cells. The reasons bone is an ideal anatomic microenvironment for HSC maintenance and differentiation have become clearer with recent data indicating that bone matrix– generating osteoblasts (OBs) provide key factors to the development of the HSC niche. There is growing evidence that bone continues to play a role in adaptive immunity at steps beyond lymphocyte development. We now know that long-lived memory T and B cells return to specialized niches in the bone marrow. Why these cells are retained in the bone and what molecular and environmental factors they rely on while there represent a series of challenging questions that will clearly beneﬁt from experiments designed in the context of osteoimmunology. How the immune system exerts its inﬂuence on bone is equally interesting. Ontogenically, skeletal development proceeds independently of early development of the immune system. Hence, it is unlikely that there are developmental inﬂuences of the immune system on skeletal and marrow cavity formation. However, bone homeostasis and remodeling occur throughout life in all bony animals. Anatomically, bone marrow spaces are loosely compartmentalized, allowing immune cells and bone cells to interact and inﬂuence each other (Figure 1). Hence, bone homeostasis is often inﬂuenced by immune responses, particularly when the immune system has been activated or becomes diseased. During pathological conditions like arthritis, inﬁl-

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trating lymphocytes and other mononuclear cells provide several key factors that inﬂuence bone metabolism by altering the balance between bone-forming OBs and bone-resorbing osteoclasts (OCs). In the past, whether these interactions also inﬂuence normal bone homeostasis had been unclear. However, the discovery that activated T cells express the TNF superfamily member TRANCE, coupled with the subsequent ﬁnding that TRANCE is a key differentiation factor for OCs, represents critical evidence that a link exists between normal immune cell function and bone metabolism. During their lifetimes, mammals are challenged with various infectious agents, which results in a gradual change in the composition of the T cell compartment toward an accumulation of TRANCEexpressing memory cells that preferentially reside in bone. Hence, with age the immune system might exert a greater inﬂuence on bone homeostasis. Finally, how relevant are osteoimmunologic approaches in biomedical research to the treatment of human disease and the maintenance of good health? Although most research in osteoimmunology currently focuses on inﬂammatory bone diseases, such as those associated with osteoarthritis or rheumatoid arthritis (RA), it is important to consider the vast public health implications arising from common metabolic bone diseases, such as osteoporosis, that may be caused by or associated with inﬂammatory molecules. To prevent and treat these conditions, it will be necessary to develop anti-inﬂammatory agents that have a high degree of speciﬁcity so that both the immune and bone systems may function with minimal complications. With these issues in mind, we ﬁrst brieﬂy review bone development and remodeling and then focus on several key areas of crosstalk between the bone and immune system that we believe will most beneﬁt from interdisciplinary approaches aimed at elucidating underlying cellular and molecular mechanisms of action. The work thus far of many groups on the physiologic and clinical relevance of

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Figure 1 Schematic diagram of the bone microenvironment as a loosely compartmentalized lymphoid organ: T∗ (memory T cells and circulating T cells), B∗ (B cells, differentiation of which occurs via interaction with stromal cells; memory B cells also interact with stromal cells; in addition, there are circulating mature B cells), stromal cells (bone marrow stromal cells are of mesenchymal origin, but not fully characterized), M∗ (monocyte and its derivatives), and osteoid [newly formed, but not yet calciﬁed matrix, composed mostly of type I collagen (∼90%) and noncollagenous proteins (∼10%)].

TRANCE-RANK signaling has served as a cornerstone of osteoimmunology research. As such, we provide a current review of the role of TRANCE in the osteoimmune system. Additionally, we review the overlapping signaling networks within cells of the immune system and bone and attempt to build a conceptual framework of parallel signaling systems in immune and bone cells.

A BRIEF INTRODUCTION TO BONE Bone Development The skeleton is among the largest organs in the body. It is composed of a mineralized framework that is maintained by a complex cellular network (2). In addition to providing structural integrity, the skeleton is a

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Receptor activator of NF-κB (RANK): the signaling receptor for TRANCE

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Osteoid: uncalciﬁed bone matrix produced by osteoblasts and consisting mainly of collagen, but also of some noncollagenous proteins BMP: bone morphogenetic protein

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storehouse for calcium, a critical ion for a variety of metabolic processes, and it is the site of hematopoiesis. Bone development occurs along two pathways. Endochondral bone, which includes the long bones and vertebrae, begins in the embryo as a cartilaginous template (3) (Figure 2). A second type of bone, called intramembranous bone (e.g., the ﬂat bones of the skull, scapula, and ileum), forms directly from the condensation of mesenchymal cells, which directly differentiate into OBs (3). A second level of organization in endochondral bone is its division into cortical and trabecular or cancellous bone. Cortical bone forms the outer surface of endochondral bones and provides the structural integrity for many of the long bones. It forms up to 80% of the skeleton and is typically dense bone with a well-organized pattern of collagen ﬁbrils that are aligned along stress lines to provide bone with maximum strength (2, 3). Trabecular bone is thinner and less well organized, and it is primarily found traversing the bone marrow space. However, in some bones with a high degree of trabecular bone, like vertebrae, trabecular bone provides much of the structural integrity. A major function of trabecular bone is to provide a large surface area for metabolic processes. Bone turnover, which consists of bone resorption (removal) and its replacement with new bone, occurs much more frequently in trabecular bone than in cortical bone. OBs initially produce an osteoid matrix that is calciﬁed extracellularly. The major structural protein of bone is type I collagen, which provides bone with a resistance to fracture in a way that is similar to the effect of reinforcing bars in modern reinforced concrete buildings. In addition, osteoid contains a large number of other noncollagen proteins that have a variety of critical functions in bone. The mineral crystal of bone is hydroxyapetite, which is a calcium-phosphate salt, containing hydroxyl ions.

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Bone Cells OBs are derived from a mesenchymal progenitor cell that is multipotential and can also differentiate into marrow stromal cells and adipocytes (4) (Figure 3). The signals that regulate the decision of mesenchymal progenitor cells to form OBs are not fully understood. However, a number of critical paracrine signals and transcription factors have been identiﬁed. These include the transcription factors Runx2 and osterix, which when absent prevent OB formation, and members of the bone morphogenetic protein (BMP) family (5–7), which initiate the signals for OB differentiation. Most recently, it was demonstrated that Wnt signaling pathways are involved in the decision of the mesenchymal progenitor cell to become either an adipocyte or an OB (8–12). As matrix calciﬁes under the inﬂuence of the OB-produced enzyme bone-speciﬁc alkaline phosphatase, a portion of the OBs are entrapped in the calciﬁed matrix and persist in bone as unique cells called osteocytes. These cells are believed to sense mechanical force on bone and to send signals via cellular processes, termed canaliculi (13), that connect osteocytes to each other and to OBs on the surface of bone. OCs are specialized multinucleated giant cells that resorb bone (14). They are hematopoietic in origin and derive from a myeloid precursor that also gives rise to macrophages and dendritic cells (DCs) (Figure 3). The signals that stimulate OCs to form and resorb bone involve a series of transcription factors and paracrine cytokines, which are discussed in detail later in this review. OCs attach themselves to bone through a specialized structure called the sealing zone. This structure allows them to create a resorption space that is isolated from the extracellular space. OCs can acidify the resorption space to solubilize the mineral component of bone (14). To remove the organic components of bone, OCs produce lysosomal enzymes, including cathepsin K, that are released into

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Figure 2 Schematic diagram of endochondral bone development. During embryogenesis, the initial step in long bone development is the patterning of the future bone by cartilage (1). Shortly thereafter, a collar of intramembranous bone forms around the center of the cartilaginous shaft (2), and simultaneously the remaining cartilage near the center of the developing bone calciﬁes (3). The calciﬁed cartilage is then removed (4), and vascular cells invade the developing marrow space to establish a blood supply. The next step in this process is the production of mineralized bone by OBs (5). As the development of the embryo progresses, mineralized tissue, produced by OBs, replaces cartilage in the majority of the embryonic bone (6–7). However, an area of cartilage is maintained at either end of growing bones after birth. This forms a structure called the epiphyseal plate or growth plate, which in humans facilitates bone growth through childhood and into adolescence. Linear growth in humans ends after puberty when the growth plate is lost as a result of the actions of sex steroid hormones (8). www.annualreviews.org • Insights into Osteoimmunology

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Figure 3 Schematic summary of bone cell differentiation. Mesenchymal stem cells, which also give rise to myoblasts, adipocytes, chondrocytes, and some as yet uncharacterized stromal cells (also called marrow stromal cells, marrow ﬁbroblasts, or the reticular network), differentiate into preosteoblasts and then become OBs on the bone surface. OBs either incorporate into bone as osteocytes or remain on the surface as lining cells. A common myeloid lineage precursor, which can also give rise to macrophages and dendritic cells, commits to becoming a preosteoclast and then fuses to become a mature, multinucleated osteoclast.

the resorption space (14). To facilitate the resorption process, OCs polarize their structure and form a unique element called the rufﬂed border, which allows the surface area available for active transport of H+ ions through a unique vacuolar proton pump. The products of resorption traverse the OC by a process that is termed transcytosis and leave the OC through the basolateral membrane opposite the resorption space. OCs are highly motile, move across the bone surface, and resorb relatively large areas of bone. OCs die by apoptotic processes that appear to be regulated by

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paracrine-acting cytokines and possibly factors in bone matrix (14).

The Coupling of Bone Resorption and Formation The skeleton is a dynamic organ that is constantly remodeling. To accomplish this function there must be a tight coupling between bone resorption and formation. Although investigators have proposed a local coupling factor linking bone resorption to subsequent formation, its nature remains elusive

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(15). During growth and continuing to midadulthood in humans, bone mass increases (16). Genetics plays a major role in the peak bone mass that is achieved. Sex steroid hormones are also critical regulators of the skeleton (2). Loss of either estrogens in women or androgens in men is associated with an enhancement of resorption rates in bone without an equivalent increase in bone formation. In women, loss of estrogens with menopause causes roughly a doubling of the rate of bone loss and increases the risk of developing osteoporosis. Because women generally live longer and have a lower peak bone mass than men, they are more prone to develop osteoporosis (2).

INTERPLAY BETWEEN OBs AND THE IMMUNE SYSTEM The Role of OBs in the HSC Niche OBs on the endosteal surface of bone, which is adjacent to the marrow cavity, function as critical support cells for HSCs in bone marrow (17–19). Investigators have shown that HSCs are adjacent to OBs and that their number is ∼2.3-fold higher in mice upon deletion of the BMP receptor 1A (BMPR1A). Significantly, BMPR1A-deﬁcient mice also have a similar increase in OB number (17). The OBs involved are on the marrow surface of the endosteum and are likely early in their commitment to the OB lineage. Adhesion of HSC and OBs appears to be mediated by interaction of N-cadherin and β-catenin (17). Similarly, it was demonstrated that expansion of the OB population in bone by stimulation of the PTH/PTHrP (parathyroid hormone/parathyroid hormone–related protein) receptor on osteoblastic cells increased the number of HSCs in bone marrow (18). This effect appeared to be mediated by Jagged-1-Notch-1 signaling because Jagged-1 levels were increased in mice with OB-targeted activation of the PTH/PTHrP receptor. In addition, the increase in the number of HSCs in cultures of cells from trans-

Hormone and Bone Health Estrogen therapy for the treatment of osteoporosis in women had been the standard of care until the release in 2002 of the ﬁndings of the Women’s Health Initiative. This study, which was among the largest ever funded by the United States National Institutes of Health, demonstrated for the ﬁrst time that oral estrogen replacement therapy prevented the development of osteoporotic fractures in postmenopausal women. However, the study also showed that this therapy, both with and without progesterone, was associated with an increased risk of thromboembolic disease and strokes. In addition, it was reported that women who took the combination of oral estrogen plus progesterone had an increased risk of breast cancer and myocardial infarctions. Although estrogen therapy is effective for the prevention of postmenopausal osteoporosis, the United States Food and Drug Administration has recommended that it should only be considered for the treatment of women at signiﬁcant risk of osteoporosis who cannot take nonestrogen medications, and it should be used at the lowest doses and for the shortest duration possible to reach treatment goals.

genic mice with OB-targeted activation of the PTH/PTHrP receptor was abrogated by inhibitors of Notch signaling. In a converse experiment, researchers found that targeted destruction of osteoblastic cells in mice led to a decrease in HSCs in bone marrow (19). Interactions of HSCs and OBs are also mediated by interactions of Tie2 on HSCs and Angiopoietin-1 on OBs. This signaling system inhibits cell division in HSCs while maintaining their capacity for self-renewal (20).

Bone as a Reservoir for Memory B and T Cells The bone marrow has long been recognized as the site of early lymphocyte development, but more recent ﬁndings indicate that endstage and memory lymphocytes preferentially inhabit the bone marrow. Upon completing differentiation in germinal centers, antibodyproducing B cells, or plasma cells, downregulate CXCR5 and upregulate CXCR4, www.annualreviews.org • Insights into Osteoimmunology

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facilitating migration from the spleen to the bone marrow, where stromal cells highly express the CXCR4 ligand, CXCL12 (21–23). Bone marrow stromal cells, which appear to be different from mature OBs, retain plasma cells via expression of adhesion molecules VCAM-1 (vascular cell adhesion molecule 1) and E-selectin (22, 23). BrdU incorporation studies have demonstrated that plasma cells found in the bone marrow have life spans typically in excess of 90 days (21, 24), suggesting the existence in bone marrow of specialized survival niches. Although it has been reported that individual bone marrow plasma cells exhibit no intrinsic survival advantage over splenic plasma cells (24), bone marrow may be capable, unlike the red pulp of the spleen, of providing the necessary space for plasma cells to thrive as a population (25). Given the short half-life of bone marrow plasma cells when cultured in vitro, the bone marrow microenvironment may contain a combination of factors, including IL-5, IL-6, SDF-1α, TNF-α, and CD44 ligands, that are required to sustain resident bone marrow plasma cells (26). Of considerable signiﬁcance is the role of bone as a source of the B cell survival factor BAFF/BLYS (B cell– activating factor/B lymphocyte stimulator). A BAFF/BLYS receptor, BCMA (B cell maturation factor), has recently been shown to be critical for long-term plasma cell survival (27, 28). Recent ﬁndings in multiple myeloma patients show that in addition to neutrophils and monocytes, OCs serve as a greater source of BAFF/BLYS than do bone marrow stromal cells (29). Although there is little evidence that the bone marrow microenvironment is required for optimal function of plasma cells, additional research is required to clearly determine the anatomical advantage for plasma cells of relocating from secondary lymphoid organs to this site. T lymphocyte precursor cells leave the bone marrow and relocate to the thymus for further differentiation. Mature T cells function in the secondary lymphoid organs and at sites of infection. Investigators have re-

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cently shown, however, that in the absence of secondary lymphoid tissues the bone marrow can support productive cytotoxic T cell and memory cell generation (30). Although the importance of bone marrow as a secondary lymphoid organ during normal immune responses requires further study, it has potential relevance to host defense against bacterial infections in bone (e.g., osteomyelitis), where OBs may facilitate immune responses by producing a variety of immunomodulatory molecules (31). Memory T cells form after the expansion and contraction of both CD4+ and CD8+ primary response T cells. In normal mice, memory CD8+ T cells preferentially reside and homeostatically proliferate in the bone marrow (32). One homing study indicates that long-lived central memory CD8+ T cells have a special afﬁnity for the bone marrow, employing VCAM-1 for sticking and L-selectin for rolling within the bone marrow microvessels (33). The advantage of central memory CD8+ T cells relocating to the bone marrow may involve the presence of the crucial survival factor IL-15 (30, 33).

Regulation of OBs by Immune Cells and Cytokines A variety of cytokines are known to regulate osteoblastic cells. TNF-α inhibits the differentiation of OBs (34). IL-1, TNF-α, and IFN-γ inhibit collagen synthesis in OBs (35– 38). IL-4 and IL-13 suppress prostaglandin synthesis in bone and are reported to be chemoattractants for OBs (39, 40). IL-4 acts as a direct stimulator of proliferation and inhibitor of differentiation in an osteoblastic cell line (41). Similarly, IL-4-overexpressing mice exhibit a decrease in bone formation and differentiated OBs on the bone surface (42). The role of cytokines in OB apoptosis has also been studied. TNF-α is potently proapoptotic for OBs (43). Activated T lymphocytes also produce products that drive differentiation of human bone marrow stromal cells toward an osteoblastic phenotype (44). B7-H3 is an Ig

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superfamily member that is expressed on the surface of antigen-presenting cells (APCs). Recently, B7-H3 was found to be expressed on developing OBs, with its expression increasing during cell maturation (45). Furthermore, B7-H3-knockout mice have decreased cortical bone mineral density compared with littermate controls (45).

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INTERPLAY BETWEEN OCs AND THE IMMUNE SYSTEM The ﬁrst observation that immune cells could inﬂuence the activity of OCs came from the ﬁnding that supernatant from normal human phytohemagglutinin-stimulated peripheral blood monocytes contained factors that stimulated bone resorption (46). This activity was named OC-activating factor (OAF). When it was eventually puriﬁed and sequenced, the principal stimulator of bone resorption in these crude OAF preparations was found to be the cytokine IL-1 (47). Subsequently, a long list of cytokines have been identiﬁed to have effects on bone. Stimulators of bone resorption include IL-1, TNF-α, IL-6, IL-11, IL-15, and IL-17. Inhibitors of resorption include IL-4, IL-10, IL-13, IL-18, GM-CSF, and IFN-γ. TGF (transforming growth factor)-β and prostaglandins can have either stimulatory or inhibitory effects on resorption, depending on the conditions under which these factors are examined (48). Production of cytokines by immune cells has been linked to human diseases that involve bone. Perhaps the most extensive studies have been on the role of cytokines in the development of osteolytic lesions observed in RA and other inﬂammatory bone diseases, including periodontal disease. Although the immunologic perspective toward RA is typically limited to how RA is initiated (i.e., by failure of immunological tolerance) and its resultant synovitis (the conventional deﬁnition of disease culmination), a hallmark of RA is the rapid erosion of periarticular bone, which is often followed by general secondary osteoporosis (or osteopenia). OCs are found to be

prevalent at the site of focal erosion and are critical for bone erosion (49, 50). In addition to inﬂammatory bone diseases, altered immune responses or cytokine production may cause other osteolytic diseases. Estrogen withdrawal after menopause is associated with a rapid and sustained increase in the rate at which bone is lost. This phenomenon seems to result from an increase in bone resorption that is not met by an equivalent increase in bone formation. Interestingly, activated T cells may cause rapid bone loss under conditions of estrogen deﬁciency by enhancing TNF-α production (51). In a series of experiments involving ovariectomy (OVX)-induced bone loss in mice, an animal model for postmenopausal bone disease, it was reported that nude mice did not lose bone mass after OVX, suggesting that T cells are critical for this response (51). However, similar experiments using nude rats (52), RAG2- or TCR-α-deﬁcient mice, and SCID rats demonstrated OVX-induced trabecular bone loss that was equivalent with that seen in wild-type mice (Y. Choi, Y. Kadono, and J. Lorenzo, unpublished data). Curiously, loss of cortical bone upon OVX was different between T cell–deﬁcient and wild-type models and depended on the bone that was examined. These results suggest there may be compartmental and bone-speciﬁc effects of T cell depletion on OVX-induced bone loss. Additional experiments are required to determine how T cells are involved in this response of bone. These studies will likely require the use of mutant mouse models deﬁcient in speciﬁc immunoregulatory molecules to mechanistically examine the causes of OVX-induced bone loss. Special consideration must be given to the genetic background of any mutant mice examined as part of future studies. The reduction in bone density observed as a result of OVX is roughly only 30%–50%, whereas the difference in average bone density observed between 129/J and C57BL/6 mice is signiﬁcant, with 129/J exhibiting ∼40% higher bone density than C57BL/6. Together, 129/J and www.annualreviews.org • Insights into Osteoimmunology

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Osteoprotegerin (OPG): the decoy receptor for TRANCE found in soluble form Osteopetrosis: increased bone density resulting from an imbalance between the formation and breakdown of bone

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C57BL/6 mice constitute most of the knockout mice analyzed. The role of cytokines in malignancyrelated bone disease has also been studied extensively (53). Hematological malignancies such as lymphoma, multiple myeloma, and adult T cell leukemia are associated with increased OC formation and activity, possibly through dysregulation of various cytokines, including IL-1, TNF-α, and IL-6. Unlike bone disease associated with solid tumors, which is typically mediated by PTHrP, hematological malignancies are often characterized by an uncoupling of resorption from formation and the frequent development of purely lytic bone lesions. It should be noted, however, that most cytokines believed to play a role in regulating bone cells are produced by nonimmune cells, like ﬁbroblasts, as well as by immune cells, and exert pleiotropic effects on various cell types. Despite extensive research over the past two decades on this topic, the molecular and cellular signiﬁcance of these cytokines in vivo, speciﬁcally in the context of an immune response, has only recently begun to be elucidated with the identiﬁcation of the TRANCE-RANK-OPG axis (54–60).

THE TRANCE-RANK-OPG AXIS AND OSTEOIMMUNOLOGY Characterization of the functions of TRANCE and its receptors [RANK and osteoprotegerin (OPG)] have contributed signiﬁcantly to the emergence of osteoimmunology, speciﬁcally with respect to examination of the interplay between active immunity and maintenance of bone homeostasis (58–60). Because there are a number of recent reviews on the diverse physiologic function of the TRANCERANK-OPG axis (58, 59), we focus here on its role in the context of osteoimmunology (Figure 4).

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Regulation of Bone Homeostasis by the TRANCE-RANK-OPG Axis Bone homeostasis is maintained via functional balance of two cell types: OBs, which build bone, and OCs, which resorb bone (14, 60). In an ongoing cycle, OCs remove bone, and subsequently OBs ﬁll the cavity with new bone. This balance enables the continuous remodeling of the bone matrix necessary to maintain skeletal strength and a reservoir for hematopoiesis. Much recent effort has gone into determining the developmental processes underlying OC differentiation and activation. OPG was initially identiﬁed as a soluble decoy-like factor, capable of inhibiting osteoclastogenesis in vitro (61, 62) and inducing osteopetrosis when transgenically overexpressed in mice (62). Furthermore, OPGdeﬁcient mice were described as osteoporotic and found to have an excess of OCs (63). Interestingly, these mice are also susceptible to arterial calciﬁcation, highlighting a potential genetic link between osteoporosis and vascular calciﬁcation (63). The gene identiﬁed as encoding the ligand for OPG was determined to be identical to the gene originally characterized as encoding the activated T cell factor TRANCE (54–56). TRANCE is capable of inducing OC differentiation, maturation, and activation in vitro and, importantly, can do so in the absence of bone marrow stromal cells (55– 57). Many well-known osteotropic factors, including IL-1, IL-6, and IL-11, are now believed to exert most of their osteoclastogenic activity by inducing TRANCE expression on OBs (60, 64). Not surprisingly, TRANCEdeﬁcient mice are severely osteopetrotic owing to a cell nonautonomous defect in OC development (65, 66). These mice also exhibit failed tooth eruption, a common defect associated with developmental osteopetrosis, and diversion of hematopoiesis to the spleen and liver because a functional bone marrow cavity fails to form in the absence of OCs (65, 66). RANK, the signaling receptor for TRANCE, was initially identiﬁed through

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Figure 4 A model for the cellular interactions linking T cell immunity and bone homeostasis. Pathogenic stimuli or self antigens are phagocytosed and presented to naive T cells by DCs. T cells provide activating signals to DCs through CD40L and in return receive optimal activating and costimulatory signals through MHC:TCR and B7:CD28 interactions, respectively. The activated T cells are induced to express TRANCE, which provides further activating and survival signals to the DCs. The DCs may negatively regulate TRANCE:RANK signaling through upregulation of the TRANCE decoy receptor, OPG. Inﬂammatory cytokines (IL-1, TNF-α) produced during successful T cell immune responses, as well as calciotropic factors (PGE2 or VitD3), induce TRANCE expression by OBs, which cooperate with effector T cells to induce OC differentiation via provision of TRANCE to OC precursors. TRANCE signaling in mature OCs induces bone-resorbing function. OBs block TRANCE binding through secretion of OPG, whereas IFN-γ and IL-4 produced by effector T cells inhibit RANK signaling. Without proper regulation, excessive bone resorption leads to osteoporosis, arthritic joint erosion, and periodontal tooth loss. www.annualreviews.org • Insights into Osteoimmunology

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TRAF: TNF receptor–associated factor

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TRAF6: a TRAF family adapter essential for RANK signaling as well as IL-1R/TLR family signaling

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a large-scale analysis of genes expressed in DCs (67). RANK expression at the RNA level is detected in most cell types or tissues examined (67). RANK-deﬁcient mice were demonstrated to phenocopy the defect in OC development observed in the TRANCEknockout mouse, conﬁrming the exclusive speciﬁcity of TRANCE for OC-expressed RANK (68). In humans, gain-of-function mutations in RANK are associated with familial expansile osteolysis and with expansible skeletal hyperphosphatasia, which is characterized by increased OC and OB activity, resulting in fragile bones, pain, and deformity (69–73). Efforts aimed at elucidating the signaling mechanisms involved in TRANCEmediated osteoclastogenesis have been informative (58, 60, 64). RANK signal transduction is mediated by adapter proteins called TNF receptor–associated factors (TRAFs) (74–76). Of the six known TRAFs, RANK interacts with TRAFs 1, 2, 3, and 5 in a membranedistal region of the cytoplasmic tail and with TRAF6 at a distinct membrane-proximal ProX-Glu-X-X-(aromatic/acid residue) binding motif (74–77). Genetic experiments show that TRAF6-deﬁcient mice have severe osteopetrosis, implying that the key signals sent through RANK in OC precursors are mediated by the adapter molecule TRAF6 (78, 79; Y. Choi, N. Kim, and Y. Kadono, unpublished data). Downstream of TRAF6, TRANCE signaling in OCs activates PI3K, TAK1, c-Src, JNK1, p44/42 ERK, p38 MAPK, Akt/PKB, and mTOR, and subsequently a series of transcription factors including NF-κB, c-Fos, Fra-1, and NFATc1. This aspect of TRANCE signaling has been recently reviewed elsewhere (58–60, 64, 80). In addition to the signaling pathways mentioned above, TRANCE stimulation also triggers reactive oxygen species (ROS) production (81). ROS, such as superoxide anions, hydroxyl radicals, and H2 O2 , have been associated with many cellular responses, including metabolic bone diseases found in aged osteoporotic women (82). Walsh et al.

Recent reports suggest that ROS act as a key second messenger during osteoclastogenesis (81), such that TRANCE stimulation induces the production of ROS in OC precursors via the small GTPase Rac1 and the ROSinducing factor NADPH oxidase (Nox) 1. How ROS cross-regulates the signaling pathways necessary for OC differentiation is unclear, although one interesting hypothesis is that ROS may potentiate mitogen-activated protein kinase (MAPK) activation by inactivating protein tyrosine phosphatase activity in a manner similar to mechanisms recently described in B cells (83).

Modulation of Immunity by the TRANCE-RANK-OPG Axis The signiﬁcance of TRANCE-RANK-OPG signaling in regulating the immune system continues to emerge. Initial studies of TRANCE- and RANK-deﬁcient mice demonstrated the importance of these signals for secondary lymphoid organ development, as these animals display a lack of peripheral lymph nodes and abnormalities in B cell follicle formation and marginal zone integrity in the spleen (58, 59). In this section, however, we focus on the role TRANCE-RANK plays in shaping the immune response in the adult immune system. To date, most reported data indicate that TRANCE modulates immunity through DCs (Figure 4). DCs are the most potent professional APCs and are required to initiate T cell–mediated immunity in vivo (84). DCs differentiate from the hematopoietic monocyte/macrophage progenitor cell lineage and, as close relatives of OCs, can be generated in vitro by treating a common precursor cell with GM-CSF. GM-CSF suppresses c-Fos and Fra-1 (85, 86), which are key transcription factors for OC differentiation. These results highlight a mechanism of developmental divergence between these two cell types. Upon receipt of inﬂammatory or activating stimuli, DCs home to the T cell areas of the lymph nodes to activate antigen-speciﬁc

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T cells. Productive activation relies on numerous DC-speciﬁc factors, including alteration of the chemokine receptor repertoire, upregulation of costimulatory molecules, and cytokine production. These modiﬁcations are induced by exogenous inﬂammatory stimuli as well as by signals transmitted by TNF family members, TNF-α and CD40L. TRANCE signaling has also been implicated in DC function, particularly with regard to regulation of DC survival. Activated DCs are relatively short-lived cells, with a halflife as low as 1–2 days upon arrival in the lymph node (87), yet in situ imaging studies suggest that individual T-DC couplings may last 37 h or longer (88–90). TRANCEprolonged DC survival is attributed to upregulation of the antiapoptotic protein Bcl-xL (91) through a pathway requiring the NF-κB components p50 and c-Rel (92). Treatment of DCs with TRANCE also activates the antiapoptotic serine/threonine kinase, Akt/PKB, through recruitment of PI3-K by TRAF6 and Cbl-b to RANK, in a mechanism dependent on the kinase activity of c-Src (75, 93). TRANCE-prolonged DC survival also has in vivo relevance, as pretreatment of peptidepulsed DCs with TRANCE prior to subcutaneous injection into recipient mice results in signiﬁcantly elevated DC persistence in draining lymph nodes and in enhanced Th1 cytokine production and T cell memory formation (94). DC vectors intended for use in immunotherapy persist longer when pretreated with TRANCE (95), and enforced autocrine TRANCE-RANK signaling but not CD40L-CD40 signaling on DCs enhances antitumor immunity (96). Opg−/− DCs potentiate in vitro mixed lymphocyte reaction (MLR), despite CD86, MHC class II, and antigen-presentation levels identical to syngeneic opg+/− DCs (97). Blockade of TRANCE signaling in vivo results in a slightly reduced CD4+ T cell response to LCMV infection, although the response is severely inhibited in the absence of CD40 signaling (98). These experiments highlight the requirement for TNF family

member signaling in the generation of antiviral immunity, as well as the degree to which TRANCE-RANK and CD40L-CD40 function overlap. However, physiologic signaling through RANK is more limited in scope than that through CD40 in that treatment of immature DCs with TRANCE cannot initiate activation, and TRANCE signaling does not complement the cd40−/− defect in germinal center formation and B cell afﬁnity maturation (91, 98). This disconnect is likely not explained by intrinsic signaling differences, as RANK and CD40 activate the same set of signaling cascades, but instead is explained by differential expression patterns and kinetics. For example, on T cells CD40L is rapidly and transiently expressed and is limited only to the CD4+ subset (99). In contrast, TRANCE is expressed on both CD4+ and CD8+ T cells (100) and is capable of binding both its functional (RANK) and decoy (OPG) receptors. These interactions are also likely to succeed CD40L-CD40 signaling, as CD40L is a key inducer of RANK and OPG expression by DCs (101). The physiologic role of CD40L-CD40 versus TRANCE-RANK signaling in DC function may, therefore, depend on the phase of the immune response. CD40L-CD40 signaling may be more prominent during the initiation and effector phases, when many cellular components of the immune system are strongly activated. By contrast, TRANCE-RANK signaling may be more important during the waning phases to ensure that T memory formation is established and then to wind down remaining T-DC interactions, possibly through OPG interference with TRANCE signaling. The severe phenotype of TRANCE- and RANKknockout mice has thus far not allowed a thorough examination of the role of TRANCE in memory cell formation. Evidence also suggests that TRANCE may be important for survival of interstitial DCs engaged in antigen surveillance during the interim period separating immune responses. Human CD34+ immature DCs express both TRANCE and RANK and can therefore www.annualreviews.org • Insights into Osteoimmunology

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provide an autocrine survival signal. Peripheral maturation of these DCs leads to a downregulation of TRANCE, suggesting a requirement for an independent source of TRANCE to validate DC activation (102). TRANCE may also be involved in actively inducing tolerance. TRANCE signaling has been directly implicated in the induction of oral tolerance in mice. Feeding low-dose ovalbumin to mice concomitant with intravenous TRANCE treatment results in T cells refractory to rechallenge and correlates with in vitro production of the suppressive cytokine IL-10 by mucosal DCs (103). Another study has demonstrated that TRANCEmediated signaling is required to prevent the onset of autoimmune disease in a TNF-αinducible mouse model of diabetes and that blockade of TRANCE-RANK interactions parallel a diminution of CD4+ CD25+ regulatory lymphocytes, which are necessary to prevent CTL-mediated islet cell destruction (104). In a recent study of murine cardiac allograft tolerance, TRANCE-RANK signaling was shown to be important for the generation of regulatory T cells via intratracheal delivery of alloantigen (105). It remains unclear, though, whether TRANCE directly triggers T lymphocyte suppression or, alternatively, acts through DC intermediaries. TRANCE is also induced preferentially, among key costimulatory molecules, on T cells activated by tolerogenic DCs (106). Further study of this issue should yield insights into the generation and maintenance of T lymphocyte tolerance. In addition to regulation of DCs, TRANCE might inﬂuence B cell development. In OPG-deﬁcient mice there is an expansion of pro-B cells in the bone marrow, whereas the opposite has been observed in TRANCE- or RANK-deﬁcient mice (65, 68, 97, 107), suggesting that TRANCE-RANK interaction might be involved in the proliferation of pro-B cells. Of interest, pro-B cells also expand in the bone marrow of ovariectomized mice (108), in which TRANCE expression on OBs is thought to be increased. Future studies are required to elucidate the

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molecular mechanisms of how TRANCE might regulate the fate of pro-B cells and what the immunological consequences of this regulation might be.

Autoimmunity, Bone, and TRANCE: The Birth of Osteoimmunology TRANCE expression on T lymphocytes is induced upon T cell receptor engagement and depends on Ca2+ mobilization (54, 109). Initial experiments demonstrated that activated T lymphocytes, or even supernatants from activated T lymphocyte cultures, could support osteoclastogenesis in vitro (110). It was subsequently observed that mice lacking CTLA4, in which T lymphocytes are systemically activated, exhibit an osteoporotic phenotype associated with increased OC numbers. Transfer of ctla-4−/− T lymphocytes into rag2−/− mice leads to decreased bone density over time, which can be prevented by OPG treatment. This ﬁnding indicated that activated T cells can disrupt bone homeostasis by modulating TRANCE expression (110), although whether T cell–derived TRANCE per se is responsible for aberrant bone metabolism is unclear. In a complementary study, transgenic overexpression of TRANCE restricted to T or T/B lymphocytes was sufﬁcient to partially correct the osteopetrotic phenotype observed in TRANCE-deﬁcient mice (66; Y. Choi & N. Kim, unpublished data). Together, these data deﬁnitively showed the ability of lymphocytes, by expression of TRANCE, to regulate bone homeostasis in vivo and conﬁrmed a bona ﬁde interplay between the adaptive immune system and bone metabolism, giving birth to the ﬁeld of osteoimmunology (Figure 4). In human arthritis, inﬂammation of the synovial joints is accompanied by bone and cartilage destruction. Various animal models have been established for the study of arthritis, and the role of TRANCE in their pathogenesis has been investigated. Treating adjuvantinduced arthritis in Lewis rats with OPG had no discernible effect on inﬂammation but

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prevented bone loss and cartilage destruction (110). These experiments could not resolve whether preservation of cartilage was an indirect beneﬁt of inhibiting bone erosion or was due to independent mechanisms. A subsequent study demonstrated that bone loss and cartilage destruction were independent in an arthritis model induced by transfer of serum from K/BxN transgenic mice, where T cell activity is not required for onset of disease (111). When K/BxN serum was transferred into TRANCE-deﬁcient mice, inﬂammation and cartilage destruction were comparable to control recipients, but bone erosion was greatly reduced (111). These ﬁndings reinforced the notion that TRANCE per se mediates induction of bone destruction by OCs in animal models of autoimmune arthritis. Examination of the cellular constituents of synovial ﬂuid collected from human arthritis patients revealed that all local T lymphocytes expressed TRANCE, establishing the clinical relevance of the connection between arthritis and immunologically derived TRANCE (110). Recently, investigators have demonstrated that TRANCE, in combination with M-CSF (macrophage colony stimulating factor), can induce transdifferentiation of immature DCs to the OC lineage and that this process is signiﬁcantly enhanced by RA synovial ﬂuid, potentially identifying another mechanism for disease-related bone destruction (112). Periodontitis, induced by infection with various subgingival bacteria, is a major cause of tooth loss and is associated with increased risk for heart failure and stroke (113, 114). To examine the etiology of the disease, peripheral blood lymphocytes (PBL) from patients with localized juvenile periodontitis (LJP) were transferred into rag2−/− mice, which were then orally inoculated with the Gramnegative bacterium Actinobacillus actinomycetemcomitans (114). LJP was recapitulated in the recipient animals and was accompanied by accumulation of OCs at the alveolar sockets (114). Treatment with OPG inhibited the OC inﬁltration and bone damage (114).

In vitro stimulation of PBL showed that TRANCE was induced on CD4+ T lymphocytes activated with A. actinomycetemcomitans antigens and that disease was attenuated when the same cells were speciﬁcally depleted from recipient mice (114). This study demonstrated the importance of CD4+ T lymphocytes in the pathogenesis of periodontitis, specifically with regard to disease-related bone destruction. Bone loss has long been recognized as an extraintestinal complication of inﬂammatory disorders of the gut, such as Crohn’s disease and ulcerative colitis (115). One recent study found that patients with these diseases have elevated levels of serum OPG, which derive from the site of inﬂammation and inversely correlate with severity of bone loss (116), whereas another study found that Crohn’s disease patients have elevated levels of both OPG and soluble TRANCE (117). Mechanistic insight into this link is provided by a study demonstrating that OPG treatment of mice suffering from IL-2-deﬁciency-induced ulcerative colitis results not only in reduced osteopenia, but also in mitigation of colitis owing to reduced colonic DC survival (118). In addition to arthritis, periodontal disease, and inﬂammatory bowel disease, pathologic bone loss is observed in patients suffering from other autoimmune diseases (diabetes mellitus and lupus erythematosus), chronic viral infections (HIV), allergic diseases (asthma), and metastatic breast and lung cancers (49, 50, 119). The contribution to pathogeneses by osteoimmunologic factors merits further investigation and may provide viable therapeutic options for alleviating painful sequelae associated with a variety of conditions. Although autoimmunity is, in some cases, associated with bone loss, each productive T cell response does not have such a deleterious outcome. T cells also secrete cytokines, such as IFN-γ, IL-4, and TGF-β, that inhibit the pro-osteoclastogenic effects of TRANCE (48–50) (Figure 4). The role of the Th1 cytokine IFN-γ, in particular, appears to be www.annualreviews.org • Insights into Osteoimmunology

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crucial in preventing T lymphocyte–mediated osteoclastogenesis (120). TGF-β is characterized as both an osteotropic and immunosuppressive cytokine. Although the largest repository of latent TGF-β is in bone, its role in OC formation is complex and insufﬁciently understood (14). TGF-β downmodulates TRANCE expression in OBs, thereby negatively impacting their ability to mediate osteoclastogenesis in culture (121). However, TGF-β has also been shown to potentiate TRANCE expression in activated T lymphocytes (109) and enhance osteoclastogenesis in cultures supplemented with soluble TRANCE (121). Additional studies are necessary to determine whether TGF-β uses multiple regulatory mechanisms, and if so, what disparate purposes they might serve. Given the variety of T lymphocyte–associated cytokines with osteotropic function, it will also be useful to clarify the correlation between Th1/Th2 cytokine polarization and any attendant osteoimmunologic bone destruction.

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TLR: Toll-like receptor

TOLL-LIKE RECEPTORS, INFLAMMATION, AND BONE METABOLISM Toll-like receptors (TLRs) are members of an ancient receptor family that shares homology with IL-1R and are critical activators of the innate immune response (122). They are most highly expressed on APCs such as DCs, macrophages, and B cells, but some members are expressed on a diverse array of tissues. Ligation of these receptors by conserved microbial molecules or endogenous danger factors results in the upregulation of costimulatory molecules and the elaboration of inﬂammatory cytokines in preparation for an adaptive immune response. TLR signaling is mediated by the adaptors MyD88, TRAF6, and TRIF, which activate various downstream signaling pathways, including IKK-NF-κB, MAPK, and IRF (122). Because macrophages and DCs share a common progenitor with OCs, it is not surprising that TLR expression is also detected 48

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on bone cells (123–125). Direct signaling of various TLRs (including TLR4) on OC precursors inhibits TRANCE-mediated osteoclastogenesis (123). The data that microbial products inhibit OC differentiation via TLRs are counterintuitive because bacterial infection can cause inﬂammatory bone diseases such as periodontitis, osteomyelitis, and bacterial arthritis (126). Bone mineral density is reduced in such diseases because of excessive bone resorption by OCs. In addition, lipopolysaccharide may be a potent stimulator of bone loss by causing an increase in the number of OCs in mice. Moreover, TLR activation can enhance OB-mediated OC differentiation by inducing TRANCE and TNF-α on OBs (124, 125, 127). The basis for the apparent discrepancy between TLR stimulation as a potent negative regulator of osteoclastogenesis and the association of bacterial infection with excessive bone resorption by OCs remains unclear. As described earlier, alveolar bone destruction in periodontitis caused by infection of Gramnegative bacteria is mediated by enhanced osteoclastogenesis owing to T cell responses and subsequent upregulation of TRANCE (114). In the same study, bacterial infection of immunodeﬁcient (SCID) mice did not lead to signiﬁcant levels of alveolar bone loss, suggesting that bacterial products do not have a direct role in osteoclastogenesis because SCID mice have no known defect in OC precursors or OBs (114). Therefore, bone loss associated with bacterial infection is likely an indirect outcome of exacerbated T cell responses. Similar to macrophages or DCs, OC precursors also produce proinﬂammatory cytokines, such as TNF-α, in response to various TLR ligands (123). Moreover, although TLR stimulation inhibits OC differentiation, OC precursors treated with TLR ligands still retain high levels of phagocytic activity, which is a major host-defense mechanism for the clearance of bacterial infection. Therefore, the net outcome of TLR stimulation in OC precursors is likely the enhancement of

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immune responses toward achieving bacterial clearance. This enhancement of immune responses can be achieved by promoting cytokine production from precursor cells and by inhibiting their differentiation into nonphagocytic, nonimmune cells, such as mature OCs. Thus, interaction of these microbial products with TLRs on OC precursors appears to favor the role of OC precursors as part of the proinﬂammatory system by inhibiting their differentiation into mature OCs and by promoting the production of inﬂammatory cytokines. However, because these cells can differentiate into mature OCs if TLR ligands are removed (123), after a microbial infection is cleared the presence of residual, activated T cells can lead to the differentiation of phagocytic precursors into mature, boneresorbing OCs. In addition, TNF-α produced by OC precursors upon TLR stimulation can enhance osteoclastic bone resorption. TLRs are thus likely to regulate the balance of immune responses and bone metabolism during acute attacks of vertebrate hosts by various microbes. However, physiologic in vivo stimulation of TLRs, which are expressed on various cells, may result in different effects on bone metabolism depending on the nature of the given immune responses. In addition, ongoing stimulation of TLRs by commensal bacteria might affect bone metabolism. In support of this idea, recent data show that mice deﬁcient in mediators of the TLR/IL-1R signaling pathway (MyD88 or IRAK-M) exhibit an altered bone metabolism, although it is not clear whether the defects are due to the signals from TLRs or IL-1R (128, 129).

AN OSTEOIMMUNOLOGICAL SIGNALING NETWORK AND GENE REGULATORY MECHANISM In addition to interplay between cellular constituents of the immune system and bone, there are also numerous parallels between the signaling networks used by the cells of each

system. There are cases both of common, shared pathways and of analogous signaling mechanisms, which activate specialized gene targets via system-speciﬁc mediators. In this section, we touch brieﬂy on the better characterized shared pathways (58–60, 64, 80) and focus on some of the novel osteoimmunologic signaling mechanisms that have recently been identiﬁed (Figure 5).

Costimulation The formation and activation of OCs are processes tightly regulated by OBs, which provide at least two known essential factors for osteoclastogenesis, TRANCE and M-CSF. In addition, stromal cells produce various osteotropic factors that inﬂuence OB-induced osteoclastogenesis of bone marrow precursors. These factors can be divided into two groups: those that inﬂuence activity of OBs (e.g., TNF-α that induces TRANCE expression in OBs), and those that affect the OC precursors or OCs per se. A series of experiments showed that M-CSF and TRANCE together appear to be sufﬁcient to induce the differentiation of bone marrow precursors, spleen cells, or blood monocytes to become mature OCs in vitro. However, the expression of M-CSF, TRANCE, and their receptors is not limited to bone cells. For example, M-CSF and TRANCE are important cytokines for the activity/viability of macrophages and DCs. Despite this pleiotropy, OCs are not found in soft tissues, raising the question of why the same set of signaling receptors leads to different functional outcomes in different anatomical environments. One possibility is the existence of costimulatory molecule(s) present only in bone. Alternatively, there could be a powerful inhibitor of osteoclastogenesis in soft tissues that is not found in bone. To address this question, we proposed the hypothesis a few years ago that there exists a mechanism in preosteoclasts analogous to the costimulation requirement for T cell activation (130). Hence, our hypothesis proposed that OC differentiation is controlled www.annualreviews.org • Insights into Osteoimmunology

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Figure 5 A model for a signaling network necessary for TRANCE-induced OC differentiation and its analog in T cell activation. Factors shaded yellowish have been shown experimentally to be important for OC differentiation. Those shaded green are molecules potentially involved in OC differentiation based on their analogous roles in the TCR signaling network. Src family kinases (SFK) necessary for phosphorylation of ITAMs are not identiﬁed. Positive feedback circuits are marked as (+) and shown in red. TREM is constitutively expressed on pOCs (and monocytes), while OSCAR (OC-associated receptor) expression is further upregulated by TRANCE stimulation. The putative ligand for TREM is expressed on pOCs, whereas OSCAR-L is expressed on OBs. The connection to integrin signaling and the cytoskeleton is proposed based on the analogy to T cell signaling requirements. The term BAAF [bone and age associated factor(s)] is proposed to describe putative costimulatory factors for OCs speciﬁc to bone. For simplicity, negative regulators of the signaling network are not illustrated but are described in the text.

not only by two essential factors, M-CSF and TRANCE (analogous to MHC/antigen complexes interacting with TCR/CD4 or TCR/CD8), but also by other nonessential but critical costimulatory molecules (analogous to B7 family proteins interacting with CD28) (131). Because the in vivo concen50

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trations of M-CSF and TRANCE produced by OBs in response to bone-resorbing hormones is likely to be much lower than is provided in in vitro experiments, costimulatory molecules are likely to inﬂuence physiologic differentiation of OCs in a manner analogous to T cell activation, whereby signals from the

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costimulatory receptor CD28 complement requisite signals from the TCR complex (130, 131). In addition, as with T cells, the requirement for a particular set of costimulatory factors/receptors for OCs should vary depending on the microenvironment. Cells expressing ligands for costimulatory receptors expressed on OCs also vary, but those interacting with OCs themselves, such as OBs, most often provide costimulation (analogous to DC provision of B7 family proteins or TNF family proteins like 4–1BBL to T cells) (130). The signals resulting from the interaction of costimulatory factors and their receptors on OC precursors determine the efﬁcacy of the signals from the essential osteoclastogenic receptor, RANK (similar to TCR/CD4 or TCR/CD8 for T cells), and the sum of the two will determine the quality of OC differentiation and activation. In support of our costimulation hypothesis, we have identiﬁed a novel cell surface receptor, OSCAR, which is preferentially expressed on OCs, and we have shown that in addition to normal TRANCE-RANK signaling, interaction of OSCAR with its putative ligand (OSCAR-L) is important for OB-induced OC differentiation (131). Moreover, OSCAR-L expression is most prevalent on osteoblastic cells (131). Therefore, the OSCAR receptor/ligand pairing could be characterized as a putative costimulation receptor/factor for efﬁcient OC differentiation, and may provide bone-speciﬁc costimulation required for the differentiation of OCs in conjunction with the essential factors M-CSF and TRANCE. This signaling combination may provide a mechanistic explanation of why OCs are found only on bone surfaces in vivo (Figure 5). Although the nature of bone-speciﬁc costimulatory molecules, such as OSCAR-L, requires further study, a series of recent experiments have supported our costimulation hypothesis (132, 133). For OC development in vivo, some surface receptors on OC precursors, such as PIR-A, OSCAR, TREM2, and SIRPβ1, associate with ITAM-containing molecules, DAP12 and FcRγ, and provide

necessary costimulation and activation of Ca2+ signaling (132, 133). Hence, although a single deﬁciency for either DAP12 or FcRγ results in only minor OC defects, double deﬁciency results in severe osteopetrosis (132, 133). Additional analysis of mutant mice suggests that these receptors activate calcineurin via Syk and PLCγ (132–134). More signaling proteins have been identiﬁed in lymphocytes that bridge Syk (or ZAP-70) and PLCγ, and lead to Ca2+ activation (135, 136). Indeed, Gab2 has recently been shown to be critical for generation of functional OCs (137). It will not be surprising if additional molecules (or family members) crucial to T cell signaling are identiﬁed as playing an equivalent role in OC differentiation (Figure 5). However, it is important to point out that osteopetrosis in DAP12/FcRγ doubledeﬁcient mice is much less severe than that in TRANCE- or RANK-knockout mice, and that, in contrast to TRANCE- or RANKknockout mice, these animals exhibit signiﬁcant numbers of OCs. (132, 133). This is consistent with our hypothesis that costimulatory receptors for OC differentiation are not essential and that multiple redundancies probably exist (131). Sustained Ca2+ mobilization is necessary for OC differentiation because NFATc1 activation is absolutely required for the process (138). The NFAT family of transcription factors was originally identiﬁed as a set of regulators of gene transcription in activated T cells (139). Recently, researchers found that RANK signaling induces expression of the NFAT family member NFATc1 (NFAT2) and that this factor is critical for OC development, as NFATc1-deﬁcient precursor cells exhibit an absolute failure to differentiate into OCs (138). Like other NFAT family members, the induction and activation of NFATc1 rely on the calcium-regulated phosphatase, calcineurin, thereby explaining negative effects of calcineurin inhibitors like FK506 and cyclosporine on osteoclastogenesis. The ability of NFATc1 to regulate its own expression indicates the existence of an www.annualreviews.org • Insights into Osteoimmunology

OSCAR: osteoclast-associated receptor NFAT: nuclear factor of activated T cells

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autonomic feedback loop, although initial triggering of NFATc1 induction is mediated by TRAF6 and c-fos via TRANCE-RANK stimulation (138). Thus, Ca2+ signaling via costimulatory receptors on pre-OCs is critical for ampliﬁcation of NFATc1 activity to a level sufﬁcient for OC differentiation. Interestingly, NFATc1, in conjunction with MITF and PU.1, transactivates OSCAR expression during TRANCE-induced OC differentiation (Y. Choi & N. Kim, unpublished data). This suggests that there is a positive feedback circuit from TRANCE to NFATc1 via costimulatory receptors, such as OSCAR, during OC differentiation, which ensures a high level of NFATc1 activity (Figure 5). Key to the analogy with lymphocyte costimulation, RANK, like TCR, is still the primary, requisite receptor, the absence of which renders the secondary receptors inconsequential to osteoclastogenesis. One issue that remains to be worked out is a greater understanding of why this system has evolved, and whether there exists a parallel state in OC development that mimics anergy, or induced tolerance, as observed in lymphocytes.

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Countervailing Osteoclastogenic Signaling In addition to OPG (and factors like TGFβ, which induce TRANCE downregulation), there are other negative regulators of RANK signaling in OCs. Although T cells express TRANCE, there is a negative correlation between T lymphocyte activation and signaling through RANK, apparently due to T cell– derived IFN-γ (120). Signaling through the IFN-γR on OCs or OC precursors leads to rapid proteasomal degradation of TRAF6 and to abortive differentiation and function (120). In this way a productive immune response is prevented from having an overlapping, deleterious effect on bone in the surrounding environment. Another regulatory mechanism involves negative feedback induced by RANK itself. Activation of the fos gene by TRANCE 52

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leads to upregulation of IFN-β, which mediates a feedback mechanism blocking further c-Fos-dependent activity (140). As such, mice deﬁcient for the IFN-α/β receptor (IFNAR1) suffer from an osteoporotic phenotype characterized by an increase in OCs (140). Promoter characterization showed that TRANCE-mediated upregulation of IFN-β utilizes AP-1-binding sites and that c-Fosdeﬁcient OC precursors are incapable of inducing IFN-β production (140). To facilitate OC development, therefore, OC precursors need to upregulate the cytokine signaling regulator SOCS3 to inhibit IFN-mediated suppression (141–143). Interferons are cytokines critical to inducing productive immune responses against viruses (IFN-α/β) as well as parasites and bacteria (IFN-γ) (144–146). Type I (IFN-α/β) and type II (IFN-γ) interferons signal through different surface receptors and activate distinct DNA recognition sites [interferon-stimulated response elements (ISRE) and gamma interferon activation sites (GAS), respectively], but they use a common mediator of gene transcription, Stat1. In immune cells, IFN-activated Stat1 induces antiviral and inﬂammatory gene transcription, although in OC precursors Stat1-dependent mechanisms mitigate osteoclastogenesis. As IFNs are associated with inﬂammation and active T cell immunity, which is itself associated with enhanced osteoclastogenic factors, it is notable that they are involved in mitigating the effects of T cells on bone erosion. Interestingly, Stat1-deﬁcient mice exhibit increased OB differentiation and bone mass, as Stat1 plays a role in vivo in attenuating the critical OB transcription factor Runx2 (147). It will be interesting to determine whether the bone phenotype in Stat1-deﬁcient mice is consistent under different immunologic conditions, or whether under inﬂammatory conditions a greater role for Stat1-mediated inhibition of osteoclastogenesis emerges. Finally, since ITAM-containing molecules provide critical costimulatory signals for OC differentiation, future studies are required

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to determine whether ITIM-containing molecules counteract costimulatory signals for OC differentiation via DAP12/FcRγ.

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Quantitatively Distinctive Utilization of TRAF6 by TRANCE It remains puzzling why TRANCE-RANKTRAF6 signaling is so critical for osteoclastogenesis in vivo. Other immune receptors, including CD40 and IL-1R/TLR, are expressed on OC precursors and use TRAF6 to activate overlapping signaling cascades but do not induce osteoclastogenesis. These observations caused us to question whether qualitative or quantitative differences exist between the TRAF6-mediated signals induced by TRANCE/RANK versus other ligandreceptor pairs. Our recent data show that stimulation via overexpressed wild-type CD40 can induce osteoclastogenesis (148). RANK contains three TRAF6-binding sites on its cytoplasmic tail, whereas CD40 has one. Stimulation through modiﬁed CD40 molecules containing additional TRAF6-binding sites in the cytoplasmic tail showed a dose-dependent increase in osteoclastogenesis. Moreover, precursors overexpressing TRAF6 alone differentiate into OCs in the absence of additional signals from TRANCE. Thus, our results suggest that differences in the osteoclastogenic capacity of RANK versus other TRAF6-associated receptors may stem, in part, from a quantitative difference in TRAF6-mediated signals. Similar mechanisms of controlling distinct biological outcomes by growth/differentiation factors, despite their use in overlapping signaling cascades, have been reported (149, 150). For example, although both epidermal growth factor (EGF) and nerve growth factor (NGF) use the same kinase cascade, only NGF can induce neuronal differentiation. When EGF receptor was overexpressed, it induced neuronal differentiation similar to NGF (149). In the case of OC differentiation, any TRAF6-binding receptor might have the potential, but only RANK

can meet the threshold to induce osteoclastogenesis. Whether OC differentiation occurs through various TRAF6-utilizing receptors also correlates with the induction and persistence of NFATc1 activation. In normal OC differentiation, TRANCE-RANK interaction, in conjunction with costimulation via ITAM-containing molecules, sets the level of NFATc1, determining whether the differentiation process becomes irreversible. However, this concept also suggests that as long as NFATc1 is induced beyond the threshold level, then OC differentiation can be mediated by factors other than TRANCE-RANK. In support of this idea, we have recently found that RANK-deﬁcient splenic cells can become bone-resorbing OCs in vitro when they are stimulated by a cocktail of cytokines (Y. Choi, N. Kim, Y. Kadono, J. Lorenzo, unpublished data). Whether such an outcome is possible in vivo, speciﬁcally under abnormal or pathological conditions, requires further investigation.

CONCLUSIONS The ﬁelds of immunology and bone biology have matured such that key cellular and molecular mechanisms governing the homeostasis of the individual systems are largely understood. However, despite extensive crossregulation between bone metabolism and the immune system, the mechanisms by which one regulates the other, and the biological implications of such interactions, are poorly understood. We believe that this lack of understanding is due in part to the challenges typically associated with crossing disciplinary boundaries that form naturally during the separate evolutions of ﬁelds like modern immunology and bone biology. It is difﬁcult enough for scientists/physicians to keep abreast of advances in multiple ﬁelds, but even more so to develop the knowledge base, skills, and materials necessary to address important issues. Therefore, it will be critical to create an environment conducive to the study of www.annualreviews.org • Insights into Osteoimmunology

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intersystem crosstalk. Awareness of intersystem crosstalk will no doubt contribute to our understanding of how both bone and the immune system are regulated in a physiologic context, both at the molecular level and at the level of organ systems. Moreover, this endeavor will lead to better treatments for human diseases involving both systems, including various inﬂammatory and metabolic bone diseases, as well as tumor-induced bone lysis. Many of these pathologic processes are major targets for therapeutic intervention and are being pursued in the absence of solid scientiﬁc understanding of the molecular and cellular processes underpinning these interactions. According to the ﬁrst-ever report by

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the U.S. Surgeon General on bone health, by 2020 one in two Americans over age 50 will be at risk for fractures from osteoporosis or low bone mass. These secondary health concerns become more prominent as people not only live longer but also expect to remain active in old age. Future preventative treatments for chronic bone-related diseases that are often associated with inﬂammation and that impact quality of life will require a high degree of speciﬁcity, especially if tailored for a segment of the population already suffering from, or vulnerable to, other age-related ailments. We believe these issues place osteoimmunology in a position of unique clinical signiﬁcance.

SUMMARY POINTS 1. Bone cells are inﬂuenced by cytokines and cell surface proteins that are expressed on lymphocytes. 2. Bone cells provide key factors necessary for HSC, B cell differentiation and memory, and T cell memory. 3. Identiﬁcation of the TRANCE-RANK-OPG axis clearly established the physiological connection between immune responses and bone metabolism, thus conﬁrming the importance of various osteotropic cytokines for bone metabolism. 4. Identiﬁcation of effector cytokines produced by activated T cells as inhibitors of TRANCE-induced osteoclast differentiation explains why normal T cell responses do not overtly affect normal bone metabolism. 5. Identiﬁcation of costimulatory receptors and their signaling pathways conﬁrms the molecular parallel between T cell activation and osteoclast differentiation.

FUTURE ISSUES TO BE RESOLVED 1. What is the nature of bone marrow stromal cells and what factors are necessary for their interaction with hematopoietic lineage cells? 2. Why are osteoclasts found only on the surface of bone? What are the ligands for costimulatory receptors found on preosteoclasts? 3. To what extent do normal immune responses affect bone homeostasis? And what are the consequences of bone metabolism to adaptive immune responses? 4. How does the interplay between bone and the immune system change with age?

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LITERATURE CITED 1. Arron JR, Choi Y. 2000. Bone versus immune system. Nature 408:535–36 2. Raisz LG, Kream BE, Lorenzo JA. 2002. Metabolic bone disease. In Williams Textbook of Endocrinology, ed. RH Williams, PR Larsen, HM Kronenberg, KS Polosky, pp. 1373– 410. Philadelphia: WB Saunders 3. Ross MH, Kaye GI, Pawlina W. 2003. Bone. In Histology: A Text and Atlas, ed. MH Ross, GI Kaye, W Pawlina, pp. 180–213. Philadelphia: Lippincott Williams & Wilkins 4. Aubin JE. 2001. Regulation of osteoblast formation and function. Rev. Endocr. Metab. Disord. 2:81–94 5. Ducy P, Zhang R, Geoffroy V, Ridall AL, Karsenty G. 1997. Osf2/Cbfa1: a transcriptional activator of osteoblast differentiation. Cell 89:747–54 6. Nakashima K, Zhou X, Kunkel G, Zhang Z, Deng JM, et al. 2002. The novel zinc ﬁngercontaining transcription factor osterix is required for osteoblast differentiation and bone formation. Cell 108:17–29 7. Celeste AJ, Iannazzi JA, Taylor RC, Hewick RM, Rosen V, et al. 1990. Identiﬁcation of transforming growth factor β family members present in bone-inductive protein puriﬁed from bovine bone. Proc. Natl. Acad. Sci. USA 87:9843–47 8. Gong Y, Slee RB, Fukai N, Rawadi G, Roman-Roman S, et al. 2001. LDL receptor-related protein 5 (LRP5) affects bone accrual and eye development. Cell 107:513–23 9. Little RD, Carulli JP, Del Mastro RG, Dupuis J, Osborne M, et al. 2002. A mutation in the LDL receptor-related protein 5 gene results in the autosomal dominant high-bone-mass trait. Am. J. Hum. Genet. 70:11–19 10. Kato M, Patel MS, Levasseur R, Lobov I, Chang BH, et al. 2002. Cbfa1-independent decrease in osteoblast proliferation, osteopenia, and persistent embryonic eye vascularization in mice deﬁcient in Lrp5, a Wnt coreceptor. J. Cell Biol. 157:303–14 11. Boyden LM, Mao J, Belsky J, Mitzner L, Farhi A, et al. 2002. High bone density due to a mutation in LDL-receptor-related protein 5. N. Engl. J. Med. 346:1513–21 12. Bennett CN, Longo KA, Wright WS, Suva LJ, Lane TF, et al. 2005. Regulation of osteoblastogenesis and bone mass by Wnt10b. Proc. Natl. Acad. Sci. USA 102:3324–29 13. Knothe Tate ML, Adamson JR, Tami AE, Bauer TW. 2004. The osteocyte. Int. J. Biochem. Cell Biol. 36:1–8 14. Teitelbaum SL. 2000. Bone resorption by osteoclasts. Science 289:1504–8 15. Howard GA, Bottemiller BL, Turner RT, Rader JI, Baylink DJ. 1981. Parathyroid hormone stimulates bone formation and resorption in organ culture: evidence for a coupling mechanism. Proc. Natl. Acad. Sci. USA 78:3204–8 16. Riggs BL, Melton LJ 3rd. 1986. Involutional osteoporosis. N. Engl. J. Med. 314:1676–86 17. Zhang J, Niu C, Ye L, Huang H, He X, et al. 2003. Identification of the haematopoietic stem cell niche and control of the niche size. Nature 425:836–41 18. Calvi LM, Adams GB, Weibrecht KW, Weber JM, Olson DP, et al. 2003. Osteoblastic cells regulate the haematopoietic stem cell niche. Nature 425:841–46 19. Visnjic D, Kalajzic Z, Rowe DW, Katavic V, Lorenzo J, Aguila HL. 2004. Hematopoiesis is severely altered in mice with an induced osteoblast deﬁciency. Blood 103:3258–64

17, 18, and 20. All three were key papers showing the molecular link between OBs and the HSC niche.

20. Arai F, Hirao A, Ohmura M, Sato H, Matsuoka S, et al. 2004. Tie2/angiopoietin-1 signaling regulates hematopoietic stem cell quiescence in the bone marrow niche. Cell 118:149–61 21. Manz RA, Arce S, Cassese G, Hauser AE, Hiepe F, Radbruch A. 2002. Humoral immunity and long-lived plasma cells. Curr. Opin. Immunol. 14:517–21 www.annualreviews.org • Insights into Osteoimmunology

55

ARI

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22. Shapiro-Shelef M, Calame K. 2005. Regulation of plasma-cell development. Nat. Rev. Immunol. 5:230–42 23. Tokoyoda K, Egawa T, Sugiyama T, Choi BI, Nagasawa T. 2004. Cellular niches controlling B lymphocyte behavior within bone marrow during development. Immunity 20:707– 18 24. Slifka MK, Antia R, Whitmire JK, Ahmed R. 1998. Humoral immunity due to long-lived plasma cells. Immunity 8:363–72 25. Sze DM, Toellner KM, Garcia de Vinuesa C, Taylor DR, MacLennan IC. 2000. Intrinsic constraint on plasmablast growth and extrinsic limits of plasma cell survival. J. Exp. Med. 192:813–21 26. Cassese G, Arce S, Hauser AE, Lehnert K, Moewes B, et al. 2003. Plasma cell survival is mediated by synergistic effects of cytokines and adhesion-dependent signals. J. Immunol. 171:1684–90 27. O’Connor BP, Gleeson MW, Noelle RJ, Erickson LD. 2003. The rise and fall of longlived humoral immunity: terminal differentiation of plasma cells in health and disease. Immunol. Rev. 194:61–76 28. O’Connor BP, Raman VS, Erickson LD, Cook WJ, Weaver LK, et al. 2004. BCMA is essential for the survival of long-lived bone marrow plasma cells. J. Exp. Med. 199:91–98 29. Moreaux J, Cremer FW, Reme T, Raab M, Mahtouk K, et al. 2005. The level of TACI gene expression in myeloma cells is associated with a signature of microenvironment dependence versus a plasmablastic signature. Blood 106:1021–30 30. Feuerer M, Beckhove P, Garbi N, Mahnke Y, Limmer A, et al. 2003. Bone marrow as a priming site for T-cell responses to blood-borne antigen. Nat. Med. 9:1151–57 31. Marriott I. 2004. Osteoblast responses to bacterial pathogens: a previously unappreciated role for bone-forming cells in host defense and disease progression. Immunol. Res. 30:291– 308 32. Becker TC, Coley SM, Wherry EJ, Ahmed R. 2005. Bone marrow is a preferred site for homeostatic proliferation of memory CD8 T cells. J. Immunol. 174:1269–73 33. Mazo IB, Honczarenko M, Leung H, Cavanagh LL, Bonasio R, et al. 2005. Bone marrow is a major reservoir and site of recruitment for central memory CD8+ T cells. Immunity 22:259–70 34. Gilbert L, He X, Farmer P, Boden S, Kozlowski M, et al. 2000. Inhibition of osteoblast differentiation by tumor necrosis factor-α. Endocrinology 141:3956–64 35. Canalis E. 1986. Interleukin-1 has independent effects on deoxyribonucleic acid and collagen synthesis in cultures of rat calvariae. Endocrinology 118:74–81 36. Stashenko P, Dewhirst FE, Rooney ML, Desjardins LA, Heeley JD. 1987. Interleukin-1β is a potent inhibitor of bone formation in vitro. J. Bone Miner. Res. 2:559–65 37. Centrella M, McCarthy TL, Canalis E. 1988. Tumor necrosis factor-α inhibits collagen synthesis and alkaline phosphatase activity independently of its effect on deoxyribonucleic acid synthesis in osteoblast-enriched bone cell cultures. Endocrinology 123:1442–48 38. Smith DD, Gowen M, Mundy GR. 1987. Effects of interferon-γ and other cytokines on collagen synthesis in fetal rat bone cultures. Endocrinology 120:2494–99 39. Onoe Y, Miyaura C, Kaminakayashiki T, Nagai Y, Noguchi K, et al. 1996. IL-13 and IL-4 inhibit bone resorption by suppressing cyclooxygenase-2-dependent prostaglandin synthesis in osteoblasts. J. Immunol. 156:758–64 40. Lind M, Deleuran B, Yssel H, Fink-Eriksen E, Thestrup-Pedersen K. 1995. IL-4 and IL-13, but not IL-10, are chemotactic factors for human osteoblasts. Cytokine 7:78–82 41. Ura K, Morimoto I, Watanabe K, Saito K, Yanagihara N, Eto S. 2000. Interleukin (IL)-4

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42. 43.

44.

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45.

46.

47.

48. 49. 50. 51.

52. 53. 54.

55.

56.

57.

58. 59.

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and IL-13 inhibit the differentiation of murine osteoblastic MC3T3-E1 cells. Endocr. J. 47:293–302 Lewis DB, Liggitt HD, Effmann EL, Motley ST, Teitelbaum SL, et al. 1993. Osteoporosis induced in mice by overproduction of interleukin 4. Proc. Natl. Acad. Sci. USA 90:11618–22 Jilka RL, Weinstein RS, Bellido T, Parﬁtt AM, Manolagas SC. 1998. Osteoblast programmed cell death (apoptosis): modulation by growth factors and cytokines. J. Bone Miner. Res. 13:793–802 Rifas L, Arackal S, Weitzmann MN. 2003. Inﬂammatory T cells rapidly induce differentiation of human bone marrow stromal cells into mature osteoblasts. J. Cell Biochem. 88:650–59 Suh WK, Wang SX, Jheon AH, Moreno L, Yoshinaga SK, et al. 2004. The immune regulatory protein B7-H3 promotes osteoblast differentiation and bone mineralization. Proc. Natl. Acad. Sci. USA 101:12969–73 Horton JE, Raisz LG, Simmons HA, Oppenheim JJ, Mergenhagen SE. 1972. Bone resorbing activity in supernatant ﬂuid from cultured human peripheral blood leukocytes. Science 177:793–95 Dewhirst FE, Stashenko PP, Mole JE, Tsurumachi T. 1985. Puriﬁcation and partial sequence of human osteoclast-activating factor: identity with interleukin 1 β. J. Immunol. 135:2562–68 Martin TJ, Romas E, Gillespie MT. 1998. Interleukins in the control of osteoclast differentiation. Crit. Rev. Eukaryot. Gene Expr. 8:107–23 Goldring SR, Gravallese EM. 2000. Pathogenesis of bone erosions in rheumatoid arthritis. Curr. Opin. Rheumatol. 12:195–99 O’Gradaigh D, Compston JE. 2004. T-cell involvement in osteoclast biology: implications for rheumatoid bone erosion. Rheumatology (Oxford) 43:122–30 Cenci S, Weitzmann MN, Roggia C, Namba N, Novack D, et al. 2000. Estrogen deﬁciency induces bone loss by enhancing T-cell production of TNF-α. J. Clin. Invest. 106:1229– 37 Sass DA, Liss T, Bowman AR, Rucinski B, Popoff SN, et al. 1997. The role of the Tlymphocyte in estrogen deﬁciency osteopenia. J. Bone Miner. Res. 12:479–86 Guise TA, Mundy GR. 1998. Cancer and bone. Endocr. Rev. 19:18–54 Wong BR, Rho J, Arron J, Robinson E, Orlinick J, et al. 1997. TRANCE is a novel ligand of the tumor necrosis factor receptor family that activates c-Jun N-terminal kinase in T cells. J. Biol. Chem. 272:25190–94 Yasuda H, Shima N, Nakagawa N, Yamaguchi K, Kinosaki M, et al. 1998. Osteoclast differentiation factor is a ligand for osteoprotegerin/osteoclastogenesis-inhibitory factor and is identical to TRANCE/RANKL. Proc. Natl. Acad. Sci. USA 95:3597– 602 Lacey DL, Timms E, Tan HL, Kelley MJ, Dunstan CR, et al. 1998. Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell 93:165–76 Fuller K, Wong B, Fox S, Choi Y, Chambers TJ. 1998. TRANCE is necessary and sufficient for osteoblast-mediated activation of bone resorption in osteoclasts. J. Exp. Med. 188:997–1001 Walsh MC, Choi Y. 2003. Biology of the TRANCE axis. Cytokine Growth Factor Rev. 14:251–63 Theill LE, Boyle WJ, Penninger JM. 2002. RANK-L and RANK: T cells, bone loss, and mammalian evolution. Annu. Rev. Immunol. 20:795–823 www.annualreviews.org • Insights into Osteoimmunology

54. This key report identifies TRANCE as a novel TNF family member on activated T cells. 55 and 56. Both are key reports showing TRANCE as the long-sought OC differentiation factor.

57. This paper shows that TRANCE is a survival factor for OCs.

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66 and 110. Key papers establishing that activated T cells and lymphocytederived TRANCE contribute to OC differentiation in vivo.

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60. Suda T, Takahashi N, Udagawa N, Jimi E, Gillespie MT, Martin TJ. 1999. Modulation of osteoclast differentiation and function by the new members of the tumor necrosis factor receptor and ligand families. Endocr. Rev. 20:345–57 61. Tsuda E, Goto M, Mochizuki S, Yano K, Kobayashi F, et al. 1997. Isolation of a novel cytokine from human ﬁbroblasts that speciﬁcally inhibits osteoclastogenesis. Biochem. Biophys. Res. Commun. 234:137–42 62. Simonet WS, Lacey DL, Dunstan CR, Kelley M, Chang MS, et al. 1997. Osteoprotegerin: a novel secreted protein involved in the regulation of bone density. Cell 89:309–19 63. Bucay N, Sarosi I, Dunstan CR, Morony S, Tarpley J, et al. 1998. osteoprotegerin-deﬁcient mice develop early onset osteoporosis and arterial calciﬁcation. Genes Dev. 12:1260– 68 64. Boyle WJ, Simonet WS, Lacey DL. 2003. Osteoclast differentiation and activation. Nature 423:337–42 65. Kong YY, Yoshida H, Sarosi I, Tan HL, Timms E, et al. 1999. OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis. Nature 397:315–23 66. Kim N, Odgren PR, Kim DK, Marks SC Jr, Choi Y. 2000. Diverse roles of the tumor necrosis factor family member TRANCE in skeletal physiology revealed by TRANCE deficiency and partial rescue by a lymphocyte-expressed TRANCE transgene. Proc. Natl. Acad. Sci. USA 97:10905–10 67. Anderson DM, Maraskovsky E, Billingsley WL, Dougall WC, Tometsko ME, et al. 1997. A homologue of the TNF receptor and its ligand enhance T-cell growth and dendritic-cell function. Nature 390:175–79 68. Dougall WC, Glaccum M, Charrier K, Rohrbach K, Brasel K, et al. 1999. RANK is essential for osteoclast and lymph node development. Genes Dev. 13:2412–24 69. Hughes AE, Ralston SH, Marken J, Bell C, MacPherson H, et al. 2000. Mutations in TNFRSF11A, affecting the signal peptide of RANK, cause familial expansile osteolysis. Nat. Genet. 24:45–48 70. Whyte MP, Mumm S. 2004. Heritable disorders of the RANKL/OPG/RANK signaling pathway. J. Musculoskelet. Neuronal Interact.. 4:254–67 71. Whyte MP, Obrecht SE, Finnegan PM, Jones JL, Podgornik MN, et al. 2002. Osteoprotegerin deﬁciency and juvenile Paget’s disease. N. Engl. J. Med. 347:175–84 72. Whyte MP, Reinus WR, Podgornik MN, Mills BG. 2002. Familial expansile osteolysis (excessive RANK effect) in a 5-generation American kindred. Medicine (Baltimore) 81:101– 21 73. Whyte MP, Hughes AE. 2002. Expansile skeletal hyperphosphatasia is caused by a 15base pair tandem duplication in TNFRSF11A encoding RANK and is allelic to familial expansile osteolysis. J. Bone Miner. Res. 17:26–29 74. Darnay BG, Haridas V, Ni J, Moore PA, Aggarwal BB. 1998. Characterization of the intracellular domain of receptor activator of NF-κB (RANK). Interaction with tumor necrosis factor receptor-associated factors and activation of NF-κB and c-Jun N-terminal kinase. J. Biol. Chem. 273:20551–55 75. Wong BR, Besser D, Kim N, Arron JR, Vologodskaia M, et al. 1999. TRANCE, a TNF family member, activates Akt/PKB through a signaling complex involving TRAF6 and c-Src. Mol. Cell 4:1041–49 76. Wong BR, Josien R, Lee SY, Vologodskaia M, Steinman RM, Choi Y. 1998. The TRAF family of signal transducers mediates NF-κB activation by the TRANCE receptor. J. Biol. Chem. 273:28355–59 Walsh et al.

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77. Ye H, Arron JR, Lamothe B, Cirilli M, Kobayashi T, et al. 2002. Distinct molecular mechanism for initiating TRAF6 signalling. Nature 418:443–47 78. Naito A, Azuma S, Tanaka S, Miyazaki T, Takaki S, et al. 1999. Severe osteopetrosis, defective interleukin-1 signalling and lymph node organogenesis in TRAF6-deﬁcient mice. Genes Cells 4:353–62 79. Lomaga MA, Yeh WC, Sarosi I, Duncan GS, Furlonger C, et al. 1999. TRAF6 deﬁciency results in osteopetrosis and defective interleukin-1, CD40, and LPS signaling. Genes Dev. 13:1015–24 80. Takayanagi H. 2005. Mechanistic insight into osteoclast differentiation in osteoimmunology. J. Mol. Med. 83:170–79 81. Lee NK, Choi YG, Baik JY, Han SY, Jeong DW, et al. 2005. A crucial role for reactive oxygen species in RANKL-induced osteoclast differentiation. Blood 106:852–59 82. Maggio D, Barabani M, Pierandrei M, Polidori MC, Catani M, et al. 2003. Marked decrease in plasma antioxidants in aged osteoporotic women: results of a cross-sectional study. J. Clin. Endocrinol. Metab. 88:1523–27 83. Singh DK, Kumar D, Siddiqui Z, Basu SK, Kumar V, Rao KV. 2005. The strength of receptor signaling is centrally controlled through a cooperative loop between Ca2+ and an oxidant signal. Cell 121:281–93 84. Steinman RM, Bonifaz L, Fujii S, Liu K, Bonnyay D, et al. 2005. The innate functions of dendritic cells in peripheral lymphoid tissues. Adv. Exp. Med. Biol. 560:83–97 85. Matsuo K, Owens JM, Tonko M, Elliott C, Chambers TJ, Wagner EF. 2000. Fosl1 is a transcriptional target of c-Fos during osteoclast differentiation. Nat. Genet. 24:184–87 86. Miyamoto T, Ohneda O, Arai F, Iwamoto K, Okada S, et al. 2001. Bifurcation of osteoclasts and dendritic cells from common progenitors. Blood 98:2544–54 87. Kamath AT, Henri S, Battye F, Tough DF, Shortman K. 2002. Developmental kinetics and lifespan of dendritic cells in mouse lymphoid organs. Blood 100:1734–41 88. Stoll S, Delon J, Brotz TM, Germain RN. 2002. Dynamic imaging of T cell-dendritic cell interactions in lymph nodes. Science 296:1873–76 89. Shakhar G, Lindquist RL, Skokos D, Dudziak D, Huang JH, et al. 2005. Stable T celldendritic cell interactions precede the development of both tolerance and immunity in vivo. Nat. Immunol. 6:707–14 90. Lindquist RL, Shakhar G, Dudziak D, Wardemann H, Eisenreich T, et al. 2004. Visualizing dendritic cell networks in vivo. Nat. Immunol.. 5:1243–50 91. Wong BR, Josien R, Lee SY, Sauter B, Li HL, et al. 1997. TRANCE (tumor necrosis factor [TNF]-related activation-induced cytokine), a new TNF family member predominantly expressed in T cells, is a dendritic cell-specific survival factor. J. Exp. Med. 186:2075–80 92. Ouaaz F, Arron J, Zheng Y, Choi Y, Beg AA. 2002. Dendritic cell development and survival require distinct NF-κB subunits. Immunity 16:257–70 93. Arron JR, Vologodskaia M, Wong BR, Naramura M, Kim N, et al. 2001. A positive regulatory role for Cbl family proteins in tumor necrosis factor-related activation-induced cytokine (TRANCE) and CD40L-mediated Akt activation. J. Biol. Chem. 276:30011–17 94. Josien R, Li HL, Ingulli E, Sarma S, Wong BR, et al. 2000. TRANCE, a tumor necrosis factor family member, enhances the longevity and adjuvant properties of dendritic cells in vivo. J. Exp. Med. 191:495–502 95. Zhong L, Granelli-Piperno A, Choi Y, Steinman RM. 1999. Recombinant adenovirus is an efﬁcient and non-perturbing genetic vector for human dendritic cells. Eur. J. Immunol. 29:964–72 www.annualreviews.org • Insights into Osteoimmunology

67 and 91. Key reports identifying RANK as the TRANCE receptor and showing TRANCE is a survival factor for DCs.

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96. Wiethe C, Dittmar K, Doan T, Lindenmaier W, Tindle R. 2003. Enhanced effector and memory CTL responses generated by incorporation of receptor activator of NF-κB (RANK)/RANK ligand costimulatory molecules into dendritic cell immunogens expressing a human tumor-speciﬁc antigen. J. Immunol. 171:4121–30 97. Yun TJ, Tallquist MD, Aicher A, Rafferty KL, Marshall AJ, et al. 2001. Osteoprotegerin, a crucial regulator of bone metabolism, also regulates B cell development and function. J. Immunol. 166:1482–91 98. Bachmann MF, Wong BR, Josien R, Steinman RM, Oxenius A, Choi Y. 1999. TRANCE, a tumor necrosis factor family member critical for CD40 ligand-independent T helper cell activation. J. Exp. Med. 189:1025–31 99. Roy M, Waldschmidt T, Aruffo A, Ledbetter JA, Noelle RJ. 1993. The regulation of the expression of gp39, the CD40 ligand, on normal and cloned CD4+ T cells. J. Immunol. 151:2497–510 100. Josien R, Wong BR, Li HL, Steinman RM, Choi Y. 1999. TRANCE, a TNF family member, is differentially expressed on T cell subsets and induces cytokine production in dendritic cells. J. Immunol. 162:2562–68 101. Yun TJ, Chaudhary PM, Shu GL, Frazer JK, Ewings MK, et al. 1998. OPG/FDCR-1, a TNF receptor family member, is expressed in lymphoid cells and is up-regulated by ligating CD40. J. Immunol. 161:6113–21 102. Cremer I, Dieu-Nosjean MC, Marechal S, Dezutter-Dambuyant C, Goddard S, et al. 2002. Long-lived immature dendritic cells mediated by TRANCE-RANK interaction. Blood 100:3646–55 103. Williamson E, Bilsborough JM, Viney JL. 2002. Regulation of mucosal dendritic cell function by receptor activator of NF-κB (RANK)/RANK ligand interactions: impact on tolerance induction. J. Immunol. 169:3606–12 104. Green EA, Choi Y, Flavell RA. 2002. Pancreatic lymph node-derived CD4+ CD25+ Treg cells: highly potent regulators of diabetes that require TRANCE-RANK signals. Immunity 16:183–91 105. Aramaki O, Inoue F, Takayama T, Shimazu M, Kitajima M, et al. 2005. Various costimulatory pathways are essential for induction of regulatory cells by intratracheal delivery of alloantigen. Transplant Proc. 37:1934–36 106. Hochweller K, Anderton SM. 2005. Kinetics of costimulatory molecule expression by T cells and dendritic cells during the induction of tolerance versus immunity in vivo. Eur. J. Immunol. 35:1086–96 107. Kim D, Mebius RE, MacMicking JD, Jung S, Cupedo T, et al. 2000. Regulation of peripheral lymph node genesis by the tumor necrosis factor family member TRANCE. J. Exp. Med. 192:1467–78 108. Masuzawa T, Miyaura C, Onoe Y, Kusano K, Ohta H, et al. 1994. Estrogen deﬁciency stimulates B lymphopoiesis in mouse bone marrow. J. Clin. Invest. 94:1090–97 109. Wang R, Zhang L, Zhang X, Moreno J, Celluzzi C, et al. 2002. Regulation of activationinduced receptor activator of NF-κB ligand (RANKL) expression in T cells. Eur. J. Immunol. 32:1090–98 110. Kong YY, Feige U, Sarosi I, Bolon B, Tafuri A, et al. 1999. Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand. Nature 402:304–9 111. Pettit AR, Ji H, von Stechow D, Muller R, Goldring SR, et al. 2001. TRANCE/RANKL knockout mice are protected from bone erosion in a serum transfer model of arthritis. Am. J. Pathol. 159:1689–99

Annu. Rev. Immunol. 2006.24:33-63. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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ANRV270-IY24-02

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112. Rivollier A, Mazzorana M, Tebib J, Piperno M, Aitsiselmi T, et al. 2004. Immature dendritic cell transdifferentiation into osteoclasts: a novel pathway sustained by the rheumatoid arthritis microenvironment. Blood 104:4029–37 113. Clark WB, Loe H. 1993. Mechanisms of initiation and progression of periodontal disease. Periodontology 2000 2:72–82 114. Teng YT, Nguyen H, Gao X, Kong YY, Gorczynski RM, et al. 2000. Functional human Tcell immunity and osteoprotegerin ligand control alveolar bone destruction in periodontal infection. J. Clin. Invest. 106:R59–67 115. Lopez I, Buchman AL. 2000. Metabolic bone disease in IBD. Curr. Gastroenterol. Rep. 2:317–22 116. Moschen AR, Kaser A, Enrich B, Ludwiczek O, Gabriel M, et al. 2005. The RANKL/OPG system is activated in inﬂammatory bowel disease and relates to the state of bone loss. Gut 54:479–87 117. Franchimont N, Reenaers C, Lambert C, Belaiche J, Bours V, et al. 2004. Increased expression of receptor activator of NF-κB ligand (RANKL), its receptor RANK and its decoy receptor osteoprotegerin in the colon of Crohn’s disease patients. Clin. Exp. Immunol. 138:491–98 118. Ashcroft AJ, Cruickshank SM, Croucher PI, Perry MJ, Rollinson S, et al. 2003. Colonic dendritic cells, intestinal inﬂammation, and T cell-mediated bone destruction are modulated by recombinant osteoprotegerin. Immunity 19:849–61 119. Mundy GR. 2002. Metastasis to bone: causes, consequences and therapeutic opportunities. Nat. Rev. Cancer 2:584–93 120. Takayanagi H, Ogasawara K, Hida S, Chiba T, Murata S, et al. 2000. T-cellmediated regulation of osteoclastogenesis by signalling cross-talk between RANKL and IFN-γ. Nature 408:600–5 121. Quinn JM, Itoh K, Udagawa N, Hausler K, Yasuda H, et al. 2001. Transforming growth factor β affects osteoclast differentiation via direct and indirect actions. J. Bone Miner. Res. 16:1787–94 122. Takeda K, Akira S. 2004. TLR signaling pathways. Semin. Immunol. 16:3–9 123. Takami M, Kim N, Rho J, Choi Y. 2002. Stimulation by Toll-like receptors inhibits osteoclast differentiation. J. Immunol. 169:1516–23 124. Kikuchi T, Matsuguchi T, Tsuboi N, Mitani A, Tanaka S, et al. 2001. Gene expression of osteoclast differentiation factor is induced by lipopolysaccharide in mouse osteoblasts via Toll-like receptors. J. Immunol. 166:3574–79 125. Hayashi S, Yamada T, Tsuneto M, Yamane T, Takahashi M, et al. 2003. Distinct osteoclast precursors in the bone marrow and extramedullary organs characterized by responsiveness to Toll-like receptor ligands and TNF-α. J. Immunol. 171:5130–39 126. Nair SP, Meghji S, Wilson M, Reddi K, White P, Henderson B. 1996. Bacterially induced bone destruction: mechanisms and misconceptions. Infect. Immun. 64:2371– 80 127. Zou W, Schwartz H, Endres S, Hartmann G, Bar-Shavit Z. 2002. CpG oligonucleotides: novel regulators of osteoclast differentiation. FASEB J. 16:274–82 128. Sato N, Takahashi N, Suda K, Nakamura M, Yamaki M, et al. 2004. MyD88 but not TRIF is essential for osteoclastogenesis induced by lipopolysaccharide, diacyl lipopeptide, and IL-1α. J. Exp. Med. 200:601–11 129. Li H, Cuartas E, Cui W, Choi Y, Crawford TD, et al. 2005. IL-1 receptor-associated kinase M is a central regulator of osteoclast differentiation and activation. J. Exp. Med. 201:1169–77 www.annualreviews.org • Insights into Osteoimmunology

120. This paper shows a mechanism for how most normal T cell immune responses fail to elicit aberrant OC differentiation.

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131. A key paper for the costimulation hypothesis of OC differentiation.

132 and 133. Key papers describing ITAM-containing adapters and how their associated receptors serve as costimulatory receptors for OC differentiation.

138. This paper demonstrated the importance of NFATc1 for OC differentiation.

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130. Allison JP, Krummel MF. 1995. The yin and yang of T cell costimulation. Science 270:932– 33 131. Kim N, Takami M, Rho J, Josien R, Choi Y. 2002. A novel member of the leukocyte receptor complex regulates osteoclast differentiation. J. Exp. Med. 195:201–9 132. Koga T, Inui M, Inoue K, Kim S, Suematsu A, et al. 2004. Costimulatory signals mediated by the ITAM motif cooperate with RANKL for bone homeostasis. Nature 428:758–63 133. Mocsai A, Humphrey MB, Van Ziffle JA, Hu Y, Burghardt A, et al. 2004. The immunomodulatory adapter proteins DAP12 and Fc receptor γ-chain (FcRγ) regulate development of functional osteoclasts through the Syk tyrosine kinase. Proc. Natl. Acad. Sci. USA 101:6158–63 134. Faccio R, Teitelbaum SL, Fujikawa K, Chappel J, Zallone A, et al. 2005. Vav3 regulates osteoclast function and bone mass. Nat. Med. 11:284–90 135. Samelson LE. 2002. Signal transduction mediated by the T cell antigen receptor: the role of adapter proteins. Annu. Rev. Immunol. 20:371–94 136. Rudd CE, Raab M. 2003. Independent CD28 signaling via VAV and SLP-76: a model for in trans costimulation. Immunol. Rev. 192:32–41 137. Wada T, Nakashima T, Oliveira-Dos-Santos AJ, Gasser J, Hara H, et al. 2005. The molecular scaffold Gab2 is a crucial component of RANK signaling and osteoclastogenesis. Nat. Med. 11:394–99 138. Takayanagi H, Kim S, Koga T, Nishina H, Isshiki M, et al. 2002. Induction and activation of the transcription factor NFATc1 (NFAT2) integrate RANKL signaling in terminal differentiation of osteoclasts. Dev. Cell 3:889–901 139. Crabtree GR. 1999. Generic signals and speciﬁc outcomes: signaling through Ca2+ , calcineurin, and NF-AT. Cell 96:611–14 140. Takayanagi H, Kim S, Matsuo K, Suzuki H, Suzuki T, et al. 2002. RANKL maintains bone homeostasis through c-Fos-dependent induction of interferon-β. Nature 416:744– 49 141. Hayashi T, Kaneda T, Toyama Y, Kumegawa M, Hakeda Y. 2002. Regulation of receptor activator of NF-κB ligand-induced osteoclastogenesis by endogenous interferon-β (INF-β) and suppressors of cytokine signaling (SOCS). The possible counteracting role of SOCSs in IFN-β-inhibited osteoclast formation. J. Biol. Chem. 277:27880–86 142. Fox SW, Haque SJ, Lovibond AC, Chambers TJ. 2003. The possible role of TGF-βinduced suppressors of cytokine signaling expression in osteoclast/macrophage lineage commitment in vitro. J. Immunol. 170:3679–87 143. Ohishi M, Matsumura Y, Aki D, Mashima R, Taniguchi K, et al. 2005. Suppressors of cytokine signaling-1 and -3 regulate osteoclastogenesis in the presence of inﬂammatory cytokines. J. Immunol. 174:3024–31 144. Levy DE, Darnell JE Jr. 2002. Stats: transcriptional control and biological impact. Nat. Rev. Mol. Cell Biol. 3:651–62 145. Ramana CV, Gil MP, Schreiber RD, Stark GR. 2002. Stat1-dependent and -independent pathways in IFN-γ-dependent signaling. Trends Immunol. 23:96–101 146. Taniguchi T, Takaoka A. 2002. The interferon-α/β system in antiviral responses: a multimodal machinery of gene regulation by the IRF family of transcription factors. Curr. Opin. Immunol. 14:111–16 147. Kim S, Koga T, Isobe M, Kern BE, Yokochi T, et al. 2003. Stat1 functions as a cytoplasmic attenuator of Runx2 in the transcriptional program of osteoblast differentiation. Genes Dev. 17:1979–91 Walsh et al.

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148. Kadono Y, Okada F, Perchonock C, Jang HD, Lee SY, et al. 2005. Strength of TRAF6 signalling determines osteoclastogenesis. EMBO Rep. 6:171–76 149. Traverse S, Seedorf K, Paterson H, Marshall CJ, Cohen P, Ullrich A. 1994. EGF triggers neuronal differentiation of PC12 cells that overexpress the EGF receptor. Curr. Biol. 4:694–701 150. Vaudry D, Stork PJ, Lazarovici P, Eiden LE. 2002. Signaling pathways for PC12 cell differentiation: making the right connections. Science 296:1648–49

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Annual Review of Immunology Volume 24, 2006

Annu. Rev. Immunol. 2006.24:33-63. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:33-63. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaqu´ın Madrenas The FOCIS Center for Clinical Immunology and Immunotherapeutics, Robarts Research Institute, and Department of Microbiology and Immunology and Department of Medicine, University of Western Ontario, London, Ontario, Canada, N6A 5K8; email: [email protected]

Annu. Rev. Immunol. 2006. 24:65–97 First published online as a Review in Advance on November 8, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090535 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0065$20.00

Key Words costimulation, T cell activation, immunological synapse, PP2A, B7, signal plasticity

Abstract Within the paradigm of the two-signal model of lymphocyte activation, the interest in costimulation has witnessed a remarkable emergence in the past few years with the discovery of a large array of molecules that can serve this role, including some with an inhibitory function. Interest has been further enhanced by the realization of these molecules’ potential as targets to modulate clinical immune responses. Although the therapeutic translation of mechanistic knowledge in costimulatory molecules has been relatively straightforward, the capacity to target their inhibitory counterparts has remained limited. This limited capacity is particularly apparent in the case of the cytotoxic T lymphocyte-associated antigen-4 (CTLA-4), a major negative regulator of T cell responses. Because there have been several previous comprehensive reviews on the function of this molecule, we focus here on the physiological implications of its structural features. Such an exercise may ultimately help us to design immunotherapeutic agents that target CTLA-4.

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INTRODUCTION

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CTLA-4: cytotoxic T lymphocyteassociated antigen-4

Cytotoxic T lymphocyte-associated antigen4 (CTLA-4), also known as CD152, was initially described during a search for cytotoxic genes using subtractive hybridization (1). It is a type 1 transmembrane glycoprotein of the immunoglobin superfamily (1), 223 amino acids (aa) in length, with a 35 aa signal peptide (1, 2), and it is found as a covalent homodimer of 41–43 kDa (3, 4). The extracellular architecture of CTLA-4 is characterized by a single IgV-like domain containing the B71 (CD80)/B7-2 (CD86) ligand-binding site (1, 2, 4–6). Homodimerization of CTLA-4 is mediated by cysteine-dependent bonding at position 122 in the stalk region and by Nglycosylation at positions 78 and 110 (4, 7). The cytoplasmic portion of CTLA-4 is 36 aa in length and lacks any intrinsic enzymatic activity (8). It contains a lysine-rich motif located proximal to the membrane (9), two tyrosine residues at positions 165 and 182, and a proline-rich region starting at position 169 (8). Altogether, these areas have been implicated in the association of CTLA-4 with a variety of signaling molecules modulating its function (9–12). The simplicity of this structure disguises its critical function as a negative regulator of T cell–mediated immune responses (13–16).

MOLECULAR GENETICS OF CTLA-4 The CTLA-4 gene is located on chromosome 2 in humans and on chromosome 1 in mice (2, 17). This gene consists of 4 exons: Exons 1 and 2 encode the extracellular portion of the molecule, with exon 1 containing the leader peptide sequence and exon 2 the ligand-binding site; exon 3 encodes the transmembrane region, and exon 4 codes for the cytoplasmic tail (2, 17) (Figure 1). The resulting transcript can undergo alternative splicing, which is different between human and mouse CTLA-4. For human CTLA-4, one can detect (a) full-length mRNA containing 66

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exons 1 to 4, (b) a transcript coding for soluble CTLA-4 (sCTLA-4) that does not contain exon 3, and (c) a transcript coding only for exons 1 and 4 (1, 2, 17–21). Mouse T cells can express an additional CTLA-4 transcript that is known as ligand-independent CTLA-4 (liCTLA-4), containing exons 1, 3, and 4 (18). The CTLA-4 gene is mainly expressed in T cells upon activation (22), although it is constitutively expressed in the CD4+ CD25+ regulatory T cell (Treg) subset (3, 22–24). Memory T cells have a relatively large pool of intracellular CTLA-4, which is rapidly expressed on the cell surface upon activation compared to resting naive T cells, and CTLA4 is retained on the surface of memory T cells for a considerably longer time than is the case on activated naive T cells (25). Interestingly, mouse resting memory T cells express higher levels of liCTLA-4, which is downregulated upon activation (19). CTLA-4 expression is also detectable at the RNA and protein levels in resting thymocytes (1, 26) and is upregulated in all thymocyte populations upon in vivo activation with anti-CD3 antibodies (Abs) (27). Further reports have also shown CTLA-4 gene expression in a variety of cells, including B cells, monocytes, granulocytes, CD34+ stem cells, placental ﬁbroblasts, and mouse embryonic cells and embryoid bodies (28–32). The precise function of CTLA-4 in non-T cells is unknown. However, signaling to these cells by CTLA-4 ligation or to other cells via B7 could have a so far unknown regulatory role. The transcriptional regulation of CTLA4 gene expression is only partially known. It is likely initiated about 335 bp upstream from the start codon (17, 23) and may be dependent on NFAT (nuclear factor of activated T cells) because modulation of NFAT levels correlates directly with CTLA-4 expression (33, 34). Furthermore, inhibition of NFAT activation by cyclosporine A causes a marked decrease in CTLA-4 gene transcription (34). Sequence analysis of the 5 upstream region of the CTLA-4 gene revealed several transcriptional regulatory elements, including sites for

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Figure 1 The CTLA-4 gene and its various splice variants. The CTLA-4 gene consists of four exons that can give rise to four different splice variants. Each exon codes for a domain in the protein as indicated in the top of the ﬁgure. Humans express the full-length CTLA-4, exon 1–4 CTLA-4, and soluble CTLA-4 splice variants, whereas mice additionally express the ligand-independent CTLA-4.

AP-1, STAT, GATA-1, NF-κB, Oct-1, and an IL-4 negative regulatory element (17, 22, 34). Also, CTLA-4 expression is upregulated by cAMP, suggesting a cAMP-dependent regulation of CTLA-4 gene transcription (35). In T cells, CTLA-4 mRNA is detected after 1 h of T cell receptor (TCR) engagement and peaks around 24–36 h after stimulation (3, 22, 23). The translational controls and machinery responsible for the stabilization of CTLA-4 mRNA have not been completely elucidated. Investigators believe that the 3 UTR of CTLA-4 plays a major role in its stability because the 3 UTR contains three AUUUA motifs implicated as binding sites for proteins that mediate RNA degradation and as such determine the half-life of the mRNA (34, 36). Of interest, soluble CTLA-4 mRNA has

a shorter half-life than full-length CTLA-4 mRNA (37). The stability of CTLA-4 mRNA has also been tied to TCR signaling and costimulation. Consistent with this, the half-life of CTLA-4 mRNA is 4.6 h under conditions of TCR signaling alone and increases to 8.9 h in the presence of CD28-mediated costimulation (22, 34, 38). Expression of CTLA-4 protein on the surface of T cells is not detected until 24–48 h after activation (22). This export event involves a signal peptide that ensures proper targeting to the rough endoplasmic reticulum and subsequent localization within the membrane, and the leader peptide is cleaved off between serine-35 and glutamine-36 (1, 2). Once at the surface, CTLA-4 can be either retained or rapidly internalized by endocytosis and targeted to lysosomes, depending on its www.annualreviews.org • CTLA- 4 Function

Costimulation: also known as “second signal,” costimulation refers to a group of biochemical events that, in addition to antigen receptor–dependent signals, are required for full activation of T and B lymphocytes

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T cell activation: changes in gene expression that lead to the differentiation of a T lymphocyte into an effector cell or a memory cell, resulting from signals emanating from cell surface receptors (e.g., TCR) Polymorphism: high degree of allelic variation for a given gene

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molecular interactions at the surface through mechanisms discussed below. Upon internalization, it is quickly degraded with a half-life of 2 h in activated T cells (38–41). Recycling in the absence of TCR ligation has not been formally demonstrated, but TCR signaling can induce CTLA-4 recycling from the lysosomes to the surface of the T cell, providing an alternative method to increase surface expression of CTLA-4 in a timely manner (41).

THE EVOLUTIONARY BIOLOGY OF CTLA-4 Soon after CTLA-4 was identiﬁed (1), it was cloned and mapped to the same chromosomal region as CD28 in both the mouse and human genome (1, 2, 23, 42). At the protein level, CTLA-4 and CD28 share approximately 30% identity (43). Owing to the genetic and molecular similarities between CD28 and CTLA-4, the evolution of CTLA-4 has been attributed to a duplication event of an ancient costimulatory gene (23, 42, 44). There is an increasing degree of conservation of CTLA-4 from the extracellular domain to the intracellular tail. The intracellular tail of CTLA-4 is 100% conserved among mammalian species, as is the cysteine (C) residue mediating covalent bonding between individual CTLA-4 chains, although there is greater variability in the extracellular V domain (17) (Figure 2). This suggests that there is a selective pressure to conserve the mechanisms that mediate CTLA-4 dimerization and signaling through its cytoplasmic tail. CTLA-4 is expressed in humans, primates, dogs, cats, cows, sheep, rats, and mice (1, 2, 45, 46). Recent reports of a putative CTLA-4-like molecule in chickens and in rainbow trout require functional characterization (NCBI accession numbers XM 421960.1 and AY789436.1, respectively) but may provide additional insights into the evolution of the CTLA-4 molecule and the selective pressure for its mechanisms of action (Figure 3). These two homologs have cytoplasmic tails lacking many of the features seen in the wellTeft

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characterized mammalian CTLA-4. Speciﬁcally, trout CTLA-4 lacks tyrosine residues in its tail, and chicken CTLA-4 contains only one of the conserved tyrosine residues in the YVKM motif. This is consistent with the idea that the tail of CTLA-4 may have evolved from these species to take advantage of the signaling pathways present in the T cell, in addition to a more ancestral mechanism of sequestration of costimulatory ligands (see below for details). Experimental data are required to determine if the magnitude of upregulation of CTLA-4 is comparable among species. In humans, CTLA-4 is not expressed on the surface of naive, resting T cells. Only after activation is it expressed on the surface at low levels (less than 10% of total CTLA-4) (38, 47). One can argue that, given the lack of conventional signaling domains in the cytoplasmic tail of trout and chicken CTLA-4, this molecule should be massively upregulated upon T cell activation in these two nonmammalian species to compete with CD28-dependent costimulation (48).

CTLA-4 POLYMORPHISMS Given the evolutionary conservation of CTLA-4 and its differential expression in T cell subsets, transcriptional or translational changes in its expression may have serious immunological consequences. This has justiﬁed the search for polymorphisms and their link with immune-mediated diseases. To date, four main polymorphisms of the CTLA-4 gene have been identiﬁed and studied in the context of autoimmune disorders. Single nucleotide polymorphisms have been noted in human CTLA-4 at positions –1722, –1661, and –318, all in the regulatory/promoter region, and at position +49 in exon 1, which results in the substitution of an alanine for a threonine in the signal sequence of CTLA-4 (49–53). The A49G polymorphism is the only polymorphism that changes the primary amino acid sequence of CTLA-4. In vitro studies of A49G CTLA-4 have shown that this mutant form

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Figure 2 Alignment of reported mammalian CTLA-4 protein sequences. Amino acids in red indicate conserved residues among mammalian species; amino acids in blue indicate those with conserved biochemical properties. Speciﬁc areas of potential functional relevance (all of them conserved across mammalian species) are within yellow boxes. The greatest divergence of sequence occurs at the N terminus, which codes for the extracellular portion of the molecule excluding the B7-binding region. The C-terminal cytoplasmic tail of the molecule is 100% conserved among mammals.

of CTLA-4 is aberrantly processed in the endoplasmic reticulum, leading to reduced surface expression (54). In contrast, the C-318T polymorphism has been associated with higher promoter activity and subsequently increased CTLA-4 expression (52). The other two polymorphisms, T-1772C and A-1661G, are less characterized, and further investigation is required to deﬁne their effects on CTLA-4 expression and association with immunological pathologies. In addition, a dinucleotide repeat polymorphism in the 3 UTR of CTLA-4 has been identiﬁed (55). Patients with longer AT repeats in the 3 UTR

have T cells that have higher proliferative responses when stimulated with anti-CD3/antiCD28 (56). The correlation between all these polymorphisms of CTLA-4 and susceptibility to autoimmune disorders has been contentious, with some reports indicating strong correlation and others disputing such a correlation. More importantly, how these polymorphisms affect CTLA-4 function is unclear. They may affect CTLA-4 processing and intracellular trafﬁcking, or they may affect oligomerization and surface retention through indirect effects on CTLA-4 posttranslation modiﬁcations. www.annualreviews.org • CTLA- 4 Function

Oligomerization: structurally organized arrangement of individual molecules of a protein beyond dimers

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CTLA-4 AS AN INHIBITORY RECEPTOR IN T CELLS: IN VIVO EVIDENCE

Figure 3 Dendrogram indicating the degree of homology between CTLA-4 of various species. Note that human CTLA-4 most closely resembles chimpanzee CTLA-4 and has the greatest divergence from rainbow trout CTLA-4.

An additional polymorphism detected in mice is linked to the expression of liCTLA4. The expression of this alternatively spliced form is dependent on a single nucleotide polymorphism at base 77 of exon 2 (18). In diabetes-resistant mice, base 77 of exon 2 is an adenine, whereas in diabetes-susceptible mice base 77 of exon 2 is a guanine (18). Ueda et al. (18) predict that this polymorphism works by modulating alternative splicing of the CTLA-4 transcript so that the guanine favors the inclusion of exon 2 in the ﬁnal transcript, whereas the adenine does not. For comparison, the human CTLA-4 base 77 of exon 2 is a thymine that also appears to favor the transcription/translation of full-length CTLA-4 versus the liCTLA4 splice variant, which may explain why liCTLA-4 has not been detected in humans (18). 70

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The structural similarities between CTLA-4 and CD28 suggested that the former was another costimulatory receptor. Early in vitro evidence, using anti-CTLA-4 monoclonal Abs, showed that ligation of CTLA-4 enhanced T cell proliferation and IL-2 production (57–59). Subsequently, it was demonstrated that crosslinking of CTLA-4 downregulated T cell proliferation and IL-2 production, whereas blockade of CTLA-4 with Fabs had an enhancing effect on T cell activation. Together, these data pointed toward a negative role for CTLA-4 in T cell activation (60, 61). Further evidence for CTLA-4 as an inhibitory receptor came from the phenotype of CTLA-4-deﬁcient mice (62, 63). These mice are born healthy. The thymus of CTLA-4deﬁcient mice remains normal in size, alterations in thymocyte populations are not detected until late stages of their life (62–64), and negative selection seems mostly normal (65), although CTLA-4 may ﬁne tune negative selection (66). However, 5–6 days after birth a large percentage of T cells become activated and inﬁltrate nonlymphoid tissues, resulting in the death of the mice by 3–4 weeks. Pathology in these mice reveals substantial tissue destruction with severe myocarditis and pancreatitis, lymphadenopathy and splenomegaly, with their peripheral lymphoid organs containing 5–10 times the normal amount of lymphocytes. These mice also exhibit elevated levels of serum immunoglobulin owing to an increase in B cell activation (63). CTLA-4-deﬁcient T cells display a phenotype characteristic of an activated T cell (CD69+ , CD25+ , CD44hi ). The increase in T cell activation is not restricted to a particular subset of cells, as the ratio of CD4+ to CD8+ T cells remains constant and the TCR repertoire is unaltered. CTLA-4−/− T cells are capable of spontaneous proliferation in vitro and secrete increased amounts

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of interferon-γ (IFN-γ), interleukin-4 (IL-4), and granulocyte-macrophage colony-stimulating factor (GM-CSF) upon TCR engagement compared with normal mice (62). Additionally, the number of T cells progressing through the cell cycle was 3–4 times higher in CTLA-4-deﬁcient mice compared with normal littermate controls, as measured by BrdU labeling (64). Such a robust proliferation of CTLA-4-deﬁcient T cells is not due to a defect in apoptosis, as these cells are susceptible to Fas-mediated apoptosis (65). Together, this phenotype suggests a generalized expansion of self-reactive T cells that are no longer held in check, implying that CTLA-4 is involved in negative regulation of T cell activation. Subsequent studies have corroborated this interpretation and shown that CTLA-4 may act by terminating cytokine production and proliferation (67). These results indicate the essential role of CTLA-4 in the negative regulation of T cell activation as well as its requirement for maintenance of T cell homeostasis. Experimental data generated from TCRtransgenic mice suggest that antigen-speciﬁc TCR signaling is required for the initiation of T cell activation in CTLA-4-deﬁcient mice (65). In addition to TCR stimulation, CTLA4-deﬁcient T cells require costimulation. Recombinant CTLA-4-Ig can effectively block CD28:B7 interactions by binding to both B7-1 and B7-2, thereby preventing CD28dependent costimulation (68). Treatment of CTLA-4−/− mice with daily injections of CTLA-4-Ig protected these animals from fatal lymphoproliferation and multi-organ inﬁltration in vivo (64, 69). The absolute numbers of CD4+ and CD8+ T cells remained normal, and these cells retained a naive phenotype (64). Isolated splenocytes and lymph node cells failed to proliferate spontaneously in vitro compared with cells isolated from isotype-treated CTLA-4-deﬁcient mice. Removal of CTLA-4-Ig treatment resulted in the rapid induction of T cell activation and the onset of the lymphoproliferative disease (69). These results rule out a defect in the TCR-

dependent signaling machinery as a cause of the phenotype of CTLA-4−/− mice. An observation that is still puzzling is that the CTLA-4 deﬁciency in T cells is not cell autonomous. Although lack of CTLA-4 correlates with disregulated T cell activation and homeostasis, reconstituted RAG-deﬁcient mice receiving equal numbers of wild-type T cells and CTLA-4-deﬁcient T cells do not develop lymphoproliferation and remain mostly healthy (70, 71). In the presence of CTLA-4-expressing T cells, CTLA-4−/− T cells undergo normal activation, expansion, and contraction when challenged with acute, chronic, and persistent infections (72). Together, these data suggest that although CTLA-4 may provide inhibitory signals in a cell autonomous fashion, additional factors (potentially immunosuppressive cytokines) provided by all or at least a subset of CTLA-4expressing T cells (potentially Tregs) can protect mice from the lymphoproliferative disorder seen in CTLA-4-deﬁcient mice (72, 73). In summary, most of the observations of CTLA-4 function point clearly toward a negative regulatory role on TCR-mediated T cell activation. Such an effect can be revealed by coligation of CTLA-4 with the TCR or, alternatively, by blockade of B7-dependent costimulation with CTLA-4. Either one of these strategies leads to a decrease in T cell responses as measured by in vitro readouts or in vivo functional outcomes.

T cell homeostasis: regulation of steady state numbers of T lymphocytes

CTLA-4 LIGANDS: B7-1 AND B7-2 The structural similarities and sequence homology between CTLA-4 and CD28 extend to shared natural ligands: the B7 molecules B7-1 (CD80) and B7-2 (CD86) (74–76). However, a key feature that may explain CTLA-4’s mechanism of action is that CTLA-4 binds B7-1 and B7-2 with greater afﬁnity and avidity compared with CD28 (68, 77). Although initial measurements suggested a 100-fold increased afﬁnity and avidity for CTLA-4:B7 interactions, more recent www.annualreviews.org • CTLA- 4 Function

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APC: antigen-presenting cell Immunological synapse (IS): an organized interface between a T lymphocyte and an APC, involving a central core in which engaged antigen receptors and costimulatory molecules are clustered, and a peripheral ring composed of engaged adhesion molecules

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data using surface plasmon resonance experiments revealed that the interactions between these molecules are 10-fold weaker owing to rapid dissociation rates. CTLA-4 binds B7-1 and B7-2 with dissociation constants (Kd) of 0.2 and 2.6 μM, respectively, whereas CD28 binds B7-1 and B7-2 with Kd values of 4 and 20 μM, respectively (78–80). The higher afﬁnity and avidity of CTLA-4 over CD28 for the B7 molecules suggest that inhibitory signals are dominant over costimulatory signals. As CTLA-4 is primarily an intracellular molecule and is maximally expressed 30- to 50-fold less than CD28 on the cell surface following activation, the stronger CTLA-4:B7 interaction compensates for the bias towards CD28 costimulation (59). The higher avidity of CTLA-4 for B7 is due to the structural topology of the CTLA-4 homodimer that allows for bivalent binding to B7 (81). In contrast, CD28 is only able to bind B7 monovalently (78, 82). An additional factor to consider is that the binding of CTLA-4 and of CD28 to both B7-1 and B7-2 occurs via the same MYPPPY motif located on exon 2 (83). Thus, the increased CTLA-4 afﬁnity in the context of the conservation of the MYPPPY sequence suggests that residues outside of the MYPPPY motif play a role in stabilizing interactions with B7 molecules (78). Most studies in the literature have considered the binding of CTLA-4 to B7-1 functionally equivalent to the binding to B7-2. However, recent data have revealed differences in the response to CTLA-4 ligation by each of these molecules individually. It is important to point out that B7-1 and B7-2, although similar, are distinct molecules that have unique proﬁles of expression. The expression patterns of the B7 ligands mimic that of CD28 and CTLA-4. B7-2 is constitutively expressed on resting antigen-presenting cells (APCs) and is upregulated rapidly after activation, reaching peak levels at 24 h (84–86). In contrast, B7-1 expression is induced after activation, peaking around 48–72 h (84). Researchers interpreted the differences in temporal regulation of these molecules by conTeft

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cluding that B7-2 is the predominant CD28 (costimulatory) ligand and B7-1 is the predominant CTLA-4 (inhibitory) ligand. In addition, the binding properties of B7-1 and B7-2 to CTLA-4 and CD28 are such that the ratio of CTLA-4 engagement to CD28 engagement in binding to B7-1 is approximately 20 to 1, compared with a ratio of 8 to 1 in binding to B7-2 (78, 80). In humans, because B7-1 is maximally expressed upon activation of APCs, whereas B7-2 is constitutively expressed, it follows that CTLA-4 interactions with B7-1 will be more prominent later in the activation process, mediating stronger inhibitory capacities upon CTLA-4 engagement (84–86). In addition, CD28 homodimers, but not CTLA-4 homodimers, are predicted to be monovalent, suggesting differences in how these receptors interact with their ligands (78, 87). Crystallographic data are consistent in showing that CTLA4 homodimers interact with alternating B7-1 homodimers, forming a lattice-like structure (81). In contrast, homodimerization of B7-2 is considered unlikely, thereby limiting the extent of a similar lattice with CTLA-4 (78, 82). At the cellular level, recent data show that B7-2 stabilizes CD28 at the immunological synapse (IS), whereas B7-1 preferentially recruits CTLA-4 to the synapse (88). On the basis of this type of data, the CTLA-4/CD28binding paradigm has been reﬁned to propose that B7-1 and B7-2 are individual ligands with potentially independent functions, as shown by their differential interactions with CTLA4 and CD28 during the timeline of an immune response. Such a paradigm helps explain the different effects of B7-1 versus B7-2 blockade on the progression of autoimmunity (89). Although in the past ﬁve years new members of the B7 family have been identiﬁed and shown to have costimulatory and coinhibitory function (reviewed in 43, 90), only B7-1 and B7-2 behave as true endogenous ligands of CTLA-4. However, the potential for additional ligands remains, based on data pointing to alternative forms of CTLA-4 with B7independent effects and to an inherent signal

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plasticity of CTLA-4 revealed by recombinant ligands. Speciﬁcally, Kuchroo’s group (19) has revealed the existence of a splice variant of CTLA-4 that lacks exon 2 and thus does not have a B7-binding domain. This liCTLA4 is expressed at higher levels in resting memory T cells and appears to be functional as an attenuator of T cell activation. Similar B7independent CTLA-4 functions have been revealed by Bluestone’s group (91). Additionally, our group has shown that one can convert CTLA-4 from inhibitor to activator of T cells with a recombinant, bispeciﬁc tandem ScFv molecule named 24:26, raising the possibility of an endogenous ligand having a similar role (see below) (92). Similar activating effects of CTLA-4 ligation have been reported by Liu’s group (57) and more recently by Nandi’s group (93). The idea of a B7-independent function of CTLA-4 is also suggested by data linking liCTLA-4 both to resistance to autoimmune diabetes (19) and to decreased T cell proliferation and IFN-γ production in response to APC/anti-CD3 stimulation (19, 91). Similar conclusions have been reached by looking at the effect of reconstitution of CTLA-4−/− mice with a mutant CTLA-4 lacking a functional B7-binding site. The mechanism by which these B7-independent forms may work is unclear, but it has been linked to decreased phosphorylation of TCR-ζ and decreased TCR signaling (19). In fact, in these studies liCTLA-4 was more potent at inhibiting TCR-ζ phosphorylation than was full-length CTLA-4. The increased signaling capacity is illustrated by the fact that diabetes-resistant mice have higher expression of liCTLA-4 in memory/regulatory T cells than do diabetessusceptible mice (19).

CTLA-4 DIMERIZATION AND HIGHER ORDER OLIGOMERIZATION The crystal structure of CTLA-4 bound to B7-1 suggests that the bivalent binding of CTLA-4 to different B7-1 dimers

would lead to the formation of a repeating lattice-like structure of alternating CTLA-4/ B7 molecules (81, 94–96). Such an arrangement is contingent on CTLA-4 dimerization. Because CTLA-4 function may be determined by dimerization and subsequent high-order CTLA-4 arrangement, one may assume that these structural characteristics would be tightly regulated. This seems to be the case. CTLA-4 dimerization in solution is mediated by intermolecular disulﬁde bond formation through the C residue at position 122 [located in the extracellular portion of CTLA-4 (4)]. The CTLA-4 dimer is further stabilized by the interface between each monomer through 4 hydrogen bonds, 48 van der Waals contacts, and 15 water-mediated hydrogen bonds (5). However, in vivo, regulation of CTLA-4 dimerization is more complex and involves a hierarchical process that includes, in addition to C122-dependent disulﬁde bonding, N-glycosylation at N78 and N110 and engagement to B7 (7, 97) (Figure 4). Interestingly, both the C122 responsible for intermolecular disulﬁde bonding and the N-glycosylation sites are evolutionarily conserved among species, suggesting that the formation of the B7-dependent, CTLA-4-dimer-based oligomers is important for the maintenance of CTLA-4 function. Such an arrangement may be important for the mechanism of T cell inactivation either by facilitating the initiation of CTLA4-mediated signaling or the sequestration of B7 by CTLA-4, or both (see below). It is important to note that the current paradigm for receptor-mediated signaling involves either ligand-induced conformational changes (98) or ligand-induced dimerization (99). However, CTLA-4 does not undergo any detectable conformation change upon B7 binding (81), and prior to ligation, it already exists as a nonfunctional covalent homodimer (4, 7), suggesting that other mechanisms of signal initiation may be involved. Upon interaction with the APC, CTLA-4 on the T cell surface accumulates at the IS (for reviews see 100, 101). Such redistribution www.annualreviews.org • CTLA- 4 Function

Plasticity: the ability to generate different or polar outcomes from the same functional unit (either molecule, cell, or organism)

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Figure 4 Hierarchical regulation of CTLA-4 dimerization and oligomer formation. Dimerization of CTLA-4 is primarily dependent on an intermolecular disulﬁde bond at cysteine 122 that is formed in the endoplasmic reticulum. Molecules lacking this disulﬁde bond undergo aberrant glycosylation at N78 and N110 in the Golgi that helps stabilize CTLA-4 dimers. CTLA-4 molecules deﬁcient in disulﬁde bonding and N-glycosylation can still dimerize upon engagement with B7 at the T cell surface. Together, this elaborate regulation suggests the importance of CTLA-4 oligomerization for its function.

involves increased partitioning of CTLA-4 to lipid rafts and is dependent on the strength of TCR signaling (100, 102, 103). Allison’s group (88) has shown that the recruitment of CTLA-4 to the IS is highly dependent on B7-1, with little contribution of B7-2. This contrasts with the recruitment of CD28 to the IS that is mostly dependent on B7-2 (88). However, in the absence of both B7-1 and B7-2, there is still low but signiﬁcant relocation of CTLA-4 to the IS (88), suggesting that CTLA-4 may still play a role in the negative regulation of the T cell activation in a B7independent fashion. However, whether this form oligomerizes when located at the IS is unclear.

MTOC: microtubule organizing center

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INTRACELLULAR TRAFFICKING OF CTLA-4 AND LOCALIZATION AT THE IMMUNOLOGICAL SYNAPSE One remarkable feature of CTLA-4 expression is that most CTLA-4 molecules remain intracellular in vesicles that localize close to the microtubule organizing center (MTOC) (37, 38). Only a small pool is expressed on the T cell surface, in a very dynamic way. This proﬁle of distribution is regulated through an interaction of the cytoplasmic tail of CTLA-4 with AP-1 during its intracellular trafﬁcking as the protein leaves the Golgi (40) and is regulated by phospholipase D and ADP ribosylation factor-1 (104) and, more importantly,

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Figure 5 Schematic representation of CTLA-4 export pathways to the T cell surface under resting conditions. Upon entering the Golgi, CTLA-4 interacts with AP-1, which mediates its trafﬁc to the cell surface, through a mechanism that may be regulated by phospholipase D (PLD) and ADP ribosylation factor-1 (ARF). CTLA-4 can also be directed to endosomes and lysosomes for storage or degradation. Surface CTLA-4 can interact with AP-2, when its cytoplasmic domain tyrosine residues are not phosphorylated, and this mediates its internalization. Whether it can undergo recycling is still unknown.

through its association with the μ2 subunit of the clathrin-associated adapter protein AP2 (12, 105, 106) (Figure 5). This latter interaction is dependent on the Y165 residue in the cytoplasmic domain, and is regulated by its phosphorylation, which prevents binding to the AP-2 adapter and blocks endocytosis (12, 106). Phosphorylation of this tyrosine residue requires Lck-dependent and ZAP-70-dependent TCR signals and leads to retention of CTLA-4 on the cell surface (107) (Figure 6). The interaction between a T cell and an APC displaying the appropriate antigenic

peptide leads to the formation of a distinct structure at the interface between both cells known as the IS (108–110). The MTOC relocalizes proximal to the IS, and the pool of CTLA-4 also relocates to this T cell:APC interface (102, 111, 112). At this point, surface expression of CTLA-4 is modulated by phosphorylation of the Y165 VKM motif (3, 12, 107), and the strength of TCR signaling dictates CTLA-4 surface expression (102), consistent with the observation that TCR signaling leads to tyrosine phosphorylation of CTLA-4 and its retention on the cell surface (107) (Figure 6). www.annualreviews.org • CTLA- 4 Function

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Figure 6 Schematic representation of CTLA-4 export pathways and retention at the plasma membrane under conditions of T cell activation. Upon activation of the T cell, CTLA-4 is expressed and its RNA stabilized. The resulting protein is exported to the surface through the Golgi in association with AP-1. In addition, upon TCR signaling surface CTLA-4 is tyrosine phosphorylated (circled P), preventing the binding of AP-2 to its cytoplasmic tail and its internalization. The net result is increased surface CTLA-4 expression and retention. In addition, activation may mediate the recycling of internalized CTLA-4 from endosomes or lysosomes back to the cell surface.

The IS includes an accumulation of membrane microdomains known as lipid rafts, both on T cells and APCs (113–115). Lipid rafts are cholesterol- and glycoprotein-enriched microdomains that are involved in signalosome assembly. Although the majority of CTLA-4 in T cells localizes in the detergent-soluble fraction or nonlipid raft fraction, a portion of CTLA-4 partitions within lipid raft microdomains (103, 116). Upon CTLA-4 activation, the amount of CTLA-4 within the lipid raft fraction increases as CTLA-4 localizes to the IS (103, 116). Such a proﬁle of distribution is necessary but does not seem 76

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sufﬁcient for T cell inactivation by CTLA-4. It is important to note that the partitioning of CTLA-4 into lipid rafts depends on its cytoplasmic tail and not on its transmembrane domain because tailless CTLA-4 partitions much less in lipid rafts despite being expressed on the surface at much higher levels (103).

MECHANISMS OF T CELL INACTIVATION BY CTLA-4 T cell inhibition by CTLA-4 is achieved through two mechanisms: competitive antagonism of CD28 signals and direct negative

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signaling (48). Engagement of CD28 molecules by B7 during early T cell responses initiates a costimulatory signaling cascade that is required for optimal T cell activation through the TCR. Such coordinated signaling leads to reorientation of intracellular CTLA-4 toward the site of TCR engagement, an increase in its surface expression, and clustering within lipid rafts at the IS (102, 112). At the IS, CTLA-4 is available to compete with CD28 for B7 engagement. Because CTLA-4 has greater avidity and afﬁnity for B7-1/B7-2 than does CD28 (48, 94, 117), CTLA-4 will likely sequester B7 molecules, thereby reducing the amount of CD28-dependent costimulation available to the cell. This mechanism does not require the cytoplasmic tail of CTLA-4, as has been shown by different groups (see 48, 118). B7 sequestration at the IS provides a simple explanation for the inhibitory function of CTLA-4 and is fully operational in vitro (48). However, its relevance and sufﬁciency in vivo is still debatable. First, it is contingent on CTLA-4 being expressed at high levels on the T cell surface compared to CD28 (requiring a 3:1 or higher ratio of CTLA-4:CD28) (48), levels not often detected on the surface. But one can argue that such levels could be achieved in relative terms owing to the focal accumulation of CTLA-4 at the IS. A second caveat is that mice expressing CTLA-4 molecules that lack the cytoplasmic tail (and are thus unable to signal) still show a lymphoproliferative phenotype, albeit a much less aggressive one, suggesting that antagonism of CD28 costimulation is not a sufﬁcient mechanism for CTLA-4 function in vivo (48, 119, 120). However, it is important to note that proper partitioning within lipid rafts at the IS requires the cytoplasmic domain of CTLA-4 (103). Thus, tailless CTLA-4 may not be able to properly provide competitive antagonism for CD28 signals. A third point is that the inhibitory function of CTLA-4 still occurs in the presence of nonlimiting CD28 costimulation. Notably, CTLA-4-mediated inactivation of T cells only occurs upon

TCR-CTLA-4 coligation (121). Thus, ligation of the TCR and CTLA-4 must occur in cis (both signals being provided by the same APC). In contrast, CD28-mediated costimulation can be provided in trans, suggesting that sequestration of B7 molecules at the IS may not be sufﬁcient to block costimulatory signals received by the T cell. A fourth point is that CTLA-4 has also been shown to inhibit some T cell responses in the absence of CD28 expression (120, 122). Based on these arguments, we lean toward the idea that antagonism of CD28-mediated costimulation is not the only mechanism of CTLA-4 function, but may be operational in vivo under some circumstances, leading not only to the inhibition of T cell activation but also to the induction of anergy owing to limited costimulation or to regulation of the activity of cyclin-dependent kinase 2 and p27kip1 (123–128). The second mechanism of CTLA-4mediated T cell inactivation is the delivery of an inhibitory signal through its cytoplasmic tail. The evidence for this mechanism derives from observations that antibody cocrosslinking of CTLA-4 and TCR in the presence of nonlimiting CD28 costimulation is sufﬁcient to induce cell-cycle arrest and inhibition of IL-2 (107, 129). In contrast to antagonism of CD28 costimulation, this mechanism is operational when CTLA-4 is expressed at low levels on the T cell surface. CTLA-4-mediated signaling is dependent on the presence of its cytoplasmic tail (48). The precise signals delivered by CTLA4 and the level of action remain uncertain (see 130). Three main models have been proposed: (a) that CTLA-4 inhibits the early events of TCR signaling, (b) that CTLA-4 inhibits events downstream of TCR and CD28 signaling, and (c) that CTLA-4 directly inhibits CD28-dependent signaling. The precise pathways that have been linked to these models are discussed below. Independently of which signaling pathway is used, CTLA4 signaling downregulates cytokine production by inhibiting the accumulation of AP-1, NF-κB, and NFAT in the nucleus of activated www.annualreviews.org • CTLA- 4 Function

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ERK: extracellular signal-regulated kinase SHP: SH2 domain-containing tyrosine phosphatase

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T cells (121, 131). CTLA-4 also regulates cell proliferation by inhibiting cyclin D3, cyclindependent kinases 4 and 6, and the degradation of cell-cycle inhibitor p27kip1 (132). In addition to direct effects on TCR/CD28 signaling, two additional processes during T cell activation may directly or indirectly be affected by CTLA-4 signaling. One is the regulation of cell surface antigen receptor and signaling molecule availability through Cbl-b. A recent study has suggested that CD28 and CTLA-4 control T cell activation and proliferation through the regulation of the E3 ubiquitin ligase Cbl-b (133). Cbl-b-knockout mice are prone to autoimmune diseases, demonstrating an important role for this molecule in the regulation of immune responses (134, 135). Interestingly, the effects of CD28 costimulation and CTLA-4 inhibition are uncoupled in Cbl-b-deﬁcient T cells, suggesting that the function of CD28 and CTLA-4 depends on the expression of Cblb. Cbl-b is known to regulate the activity of Vav (134–137). CD28 costimulation leads to the downregulation of Cbl-b at the transcriptional and post-transcriptional levels. B7 ligation of CTLA-4 results in the re-expression of Cbl-b and regulates its expression primarily at the transcriptional level. Further, CTLA4-deﬁcient T cells have signiﬁcantly reduced levels of Cbl-b (133). Therefore, CD28 and CTLA-4 may control the threshold for T cell activation by regulating the expression of Cbl-b. The other function that may be affected by CTLA-4 is cytoskeletal reorganization, impacting many diverse responses such as cell mobility and adherence, by regulating the activity of the small G protein Rap1. CD28 costimulation has been linked to inhibition of Rap1 activity, leading to enhanced levels of extracellular signal-regulated kinase (ERK) activation (138). A recent study (139) has shown that CTLA-4 has a reduced capacity to inhibit T cell responses in mice transgenic for Rap1Gap1, a negative regulator of Rap1. Furthermore, CTLA-4 was shown to activate Rap1, suggesting that CTLA-4 may act to Teft

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inhibit T cell activation by modulating the CD28-dependent activity of Rap1 (139).

MOLECULAR INTERACTIONS THROUGH THE CYTOPLASMIC TAIL OF CTLA-4: CLUES TO CTLA-4 SIGNALING The cytoplasmic domain of CTLA-4 is unique from other known receptors with inhibitory activity. The tail of CTLA-4 lacks intrinsic enzymatic activity and does not contain a bona ﬁde immune receptor tyrosinebased inhibitory motif (ITIM). However, it contains many other potential protein:protein interacting motifs. The initial studies to identify the molecular interactions of the cytoplasmic domain of CTLA-4 concentrated on two tyrosine residues located at positions 165 and 182 that could provide a putative SH2 domain-binding site. Both Y165 and Y182 can be phosphorylated in vitro by src kinases Lck, fyn, and lyn, as well as by resting lymphocyte kinase (Rlk) and by Janus kinase 2 ( Jak2) (106, 140–143). Such a potential SH2 domain-binding site in addition to the increase in tyrosine phosphorylation observed in CTLA-4-deﬁcient T cells, suggested that an SH2 domain-containing tyrosine phosphatase might mediate CTLA-4 signaling (144). This guided the identiﬁcation of SH2 domain-containing tyrosine phosphatase-2 (SHP-2) as a potential mediator through association to the Y165 VKM motif in the tail of CTLA-4 (105, 145). Phosphorylation of Y165 would recruit SHP-2 and downregulate early TCR signaling through decreasing tyrosine phosphorylation of TCRζ and LAT (linker for activation of T cells) and the Ras regulator p52SHC (140, 142, 145, 146). CTLA-4 crosslinking was also reported to decrease tyrosine phosphorylation of TCRζ and LAT, events that correlated with the formation of a multimolecular complex containing CTLA-4, SHP-2, and Lck (10). Formation of this complex did not require tyrosine phosphorylation of CTLA-4, however. Despite these results, the ability of SHP-2

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to mediate CTLA-4-dependent T cell inhibition remains controversial. CTLA-4 can inhibit signals from CD3-ε as well as from TCR-ζ (147). Mutation of Y165 or Y182 residues has revealed that neither of these residues is required for CTLA-4-mediated T cell inhibition (107, 148). The inhibitory effects of CTLA-4 are enhanced when the Y165 residue is mutated as a result of higher surface expression owing to its inability to be internalized (107). These results suggest that negative signaling through CTLA-4 is likely mediated through phosphotyrosine-independent interactions. Even when it is phosphorylated, several studies have reported that CTLA4 does not directly associate with SHP-2 or the SH2 domains of SHP-2 (149, 150), and coimmunoprecipitation of SHP-2 with CTLA-4 has not been reproduced in different systems, including models using human T cells (107, 118, 151). In addition, SHP-2 enhances rather than inhibits Ras- and mitogen-activated protein kinase (MAPK)dependent signaling (152). It is important to note, though, that there is consistency in the detection of phosphatase activity associated to the cytoplasmic regions of CTLA-4, although not associated to SHP-2 but rather to SHP-1 (153) or protein phosphatase 2A (PP2A) (9, 154). An alternative body of work points to CTLA-4 signals working downstream of the TCR. This proposal was initially based on the observation of the lack of effect of antibodymediated ligation of CTLA-4 on phosphorylation of TCR-ζ and ZAP-70 (107, 151, 155). These studies concluded that CTLA4 decreases the level of ERK and Jun-NH2terminal kinase ( JNK) activity, leading to inhibition of IL-2 production (155). More recently, using primary murine T cells, Rudd’s group (156) has shown that ligation of CTLA4 with B7-1/B7-2 could differentially regulate members of the MAPK family by inhibiting ERK activity while maintaining JNK activity. Several molecules have been linked to the inhibitory effect of CTLA-4 on ERK-1/-2 ac-

tivation. One is Rap1, whose activation would lead to inhibition of ERK-1/-2 in a Rasindependent manner (157, 158). No information is currently available on the pathway linking CTLA-4 to Rap1. Another molecule is the serine/threonine phosphatase PP2A, reported independently by Thompson’s and Kuchroo’s and our group using yeast twohybrid screening (9, 154). PP2A accounts for close to 1% of all cellular proteins and provides the majority of serine/threonine phosphatase activity within eukaryotic cells (159). PP2A is a trimeric holoenzyme composed of a core enzyme and a regulatory B subunit (PP2AB). The core enzyme is made of a catalytic subunit (PP2AC) and a scaffolding subunit (PP2AA) (159–161). PP2AA is composed of 15 tandem repeats of 39–41 aa that conform to the HEAT (Huntington/elongation/A subunit/TOR) motif. The repeats are composed of two antiparallel helices connected by a short intrarepeat loop; the 15 repeats stack together to form an elongated, left-handed, rod-like architecture that resembles a hook (162). The intrarepeat loops are highly hydrophobic, providing a suitable surface for protein:protein interactions. Mutational analysis of PP2AA identiﬁed that PP2AB binds the intrarepeat loops of HEAT motifs 1–10 in the N terminus, whereas PP2AC binds repeats 11–15 in the C terminus (163, 164). In its interaction with CTLA-4, the A subunit of PP2A binds the lysine-rich motif located at lysine residues 152, 155, and 156 of the juxtamembrane region of the CTLA4 tail, and the C subunit binds the Y165 residue (9, 154). The binding sites for the A and C subunits within the tail of CTLA-4 are 14 aa apart, which correlates exactly with the length of each alpha-helix within PP2AA. Putting this information together, we propose a model of CTLA-4:PP2A interaction in which the CTLA-4 homodimer has a PP2A trimer bound to each tail. PP2AA would bind the lysine-rich motif on the juxtamembrane portion of CTLA-4 and adopt an extended conformation from which PP2AC and PP2AB would hang, the former binding the tyrosine www.annualreviews.org • CTLA- 4 Function

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Figure 7 A model for the interaction between CTLA-4 and the serine/threonine phosphatase PP2A. The A and C subunits of the trimeric PP2A interact with the cytoplasmic domain of CTLA-4 on regions centered at lysine residues 152, 155, and 156 and tyrosine residue 165. The length of the two amphipatic helices of PP2A is 14 aa, the same distance between the ﬁrst lysine residue and tyrosine residue 165. Numbers of the HEAT domains of PP2AA are indicated. The regulatory PP2AB subunit would be accessible to interact with other molecules. The aa highlighted in yellow represent putative PP2A contact residues with CTLA-4.

165 on that tail and the latter being able to interact with other molecules (Figure 7). What is the functional relevance of the association of PP2A to CTLA-4? Newly synthesized CTLA-4 associates with PP2A, and under these conditions the inhibitory function of CTLA-4 is inactive (9). Upon TCR-CTLA-4 coligation, the A subunit is tyrosine phosphorylated, and PP2A dissociates from CTLA-4 (Figure 8). This correlates with T cell inactivation. Mutations of the lysine-rich motif that render CTLA-4 unable to bind PP2A enhance its inhibitory function, implying that 80

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PP2A when associated with CTLA-4 blocks negative signaling through this receptor (9). Interestingly, PP2A is also able to associate with CD28 (154). The association between PP2A and CD28 is also present in resting conditions, and under these conditions PP2A appears to be enzymatically active. Upon T cell activation, CD28 is tyrosine phosphorylated by Lck, and this correlates with its dissociation from PP2A (154). Thus, PP2A seems to act as a negative regulator of the function of membrane receptors to which it associates.

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Figure 8 Mechanisms of CTLA-4-mediated T cell inactivation: crosstalk between CTLA-4 and CD28. Following T cell activation, CD28 and CTLA-4 are sequentially (t1 and t2 ) phosphorylated in a Lck/ZAP-70-dependent manner by different kinases. Phosphorylated CD28 activates PKB/Akt in a PI3K-dependent manner, leading to downregulation of the E3 ubiquitin ligase Cbl-b and the small G protein Rap1. Engagement of CTLA-4 with B7 correlates with dissociation of PP2A from its tail in a phosphorylation-dependent manner (circled P). This dissociation may cause inhibition of the PKB/Akt signaling pathway and/or enhance the activity of Cbl-b and Rap1. Together, these events may mediate the inhibition of downstream signaling events. The symmetry between CD28 and CTLA-4 signaling interactions suggests that these two receptors may regulate each other through sequestration/blockade of their respective functions.

The mechanism of receptor inhibition by PP2A (e.g., CTLA-4, CD28) and its precise role in CTLA-4-mediated signaling is unknown. PP2A can inactivate protein kinase B (PKB)/Akt (165, 166), a major component of the CD28 signaling pathway dependent on phosphatidylinositol 3-kinase (PI3K) (167–169). Thus, we favor a model in which, when CTLA-4 is expressed but not ligated,

PP2A is bound to CTLA-4 and is not able to inactivate Akt. Under these conditions, CTLA-4 is not inhibitory. This may be relevant in memory T cells, in which CTLA4 is expressed under resting conditions and may control the threshold of activation (25, 170). However, upon ligation of CTLA-4 with the TCR, PP2A would dissociate from CTLA-4 and be free to inactivate Akt, thus www.annualreviews.org • CTLA- 4 Function

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mediating the inhibitory function of CTLA4 (Figure 8). Consistent with this model, genomic analysis of TCR-CTLA-4 coligation has shown that the effect of CTLA-4mediated T cell inactivation is to downregulate CD28-dependent gene transcription (171). Because PP2A, PI3K, and Grb2 all bind the YMNM motif of CD28, the dissociation and potential inactivation of PP2A may be necessary to induce CD28-mediated costimulation by allowing the binding of PI3K or Grb2. Consistent with this model are the observations that okadaic acid, a potent inhibitor of PP2A, can partially inhibit IL-2 production upon TCR/CD28 ligation, that overexpression of inactive PP2A can increase IL-2 production (154), and that T cell activation through CD28 is enhanced in CTLA-4deﬁcient T cells (172). Therefore, CTLA-4 may inhibit T cell activation by downregulating CD28 signaling. This downregulation may occur through transfer of PP2A from CTLA-4 to CD28 or, alternatively, by direct phosphatase activity of PP2A on downstream steps of CD28 signaling such as Akt. According to the model outlined above, there is a remarkable symmetry in the signaling pathways used by CD28 and by CTLA-4. PI3K associates with the YVKM motif in the cytoplasmic tail of CTLA-4 (11, 141). Also, the SH2 binding domain of the p85 subunit of PI3K associates with a similar YMNM motif in CD28 when phosphorylated (173). The interaction between PI3K and CD28 leads to the recruitment and activation of Akt (PKB), a regulator of many downstream signaling events (reviewed in 174). The ability of both of these receptors to interact with PI3K suggests that CTLA-4 could potentially exchange PP2A and PI3K, sequestering PI3K and thereby reducing its availability to mediate CD28-dependent costimulation. This process may not be the only way this interaction works because CTLA-4 in intracellular vesicles may be phosphorylated by src kinases, allowing association with PI3K (175). The cytoplasmic tail of CTLA-4 also contains a potential docking site for SH3

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domain-containing proteins. This is centered at a proline-rich (PPTEP) motif starting at residue 169. No evidence for a direct SH3 domain-mediated protein interaction has been reported to date. Furthermore, mutation of these proline residues did not affect the inhibitory function of CTLA-4 (48), calling into question a functional role for a conventional interaction with this motif. In summary, the precise nature of the signals transmitted through CTLA-4 remains controversial. Based on the experimental data available, we propose that CTLA-4 and CD28 share a set of mechanisms that regulate themselves and counterregulate each other. Within this framework, the data point to the serine/threonine phosphatase PP2A as the most likely player of this role, inhibiting the function of the receptor to which it is associated through an unknown mechanism, and at the same time enhancing signaling by the other receptor through a Lck/PI3K/Aktdependent manner. Interestingly, current evidence also implies that the mechanism of action of CTLA-4 is distinct from that of the two other negative regulatory receptors reported in T cells: PD-1 (programmed death 1) and BTLA (B and T lymphocyte attenuator) (176, 177; M.L. Baroja, B.M. Carreno, and J. Madrenas, unpublished observations). In contrast to CTLA-4, both PD-1 and BTLA are expressed as a monomer on the cell surface, suggesting that these two receptors are not governed by the same structural constraints as CTLA-4 to exert their inhibitory activities (178), and both of them use SHP-1 and SHP-2 (177, 179, 180).

SIGNALING PLASTICITY OF CTLA-4 Although most of the evidence establishes that CTLA-4 is a key negative regulator of T cell responses, emerging data suggest that it has an inherent functional plasticity. This plasticity is illustrated by the capacity of CTLA-4 to act as a positive regulator or as a negative regulator of T cell activation, depending on the

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conditions of engagement and on the signals it transduces to the T cell. Such plasticity was implied by the very early experiments showing that CTLA-4 and CD28 could act synergistically to enhance T cell proliferation (59). Later, interactions between B7-1 and CTLA4 were shown to enhance T cell clonal expansion in the absence of CD28 expression (57). The potential of B7-1 acting on a second costimulatory molecule under these conditions was unlikely because intact or Fab CTLA-4 mAbs completely blocked T cell expansion. Additionally, ligation of CTLA-4 with mutant B7-1 that had lost its ability to bind CD28 also resulted in T cell clonal expansion. As suggested by Yang Liu, results based on the use of CTLA-4 blocking Abs may be misinterpreted because these Abs could be agonists rather than antagonists of CTLA-4 function (181). Further evidence for an activating signal through CTLA-4 comes from studies on thymocyte development, in which CTLA4 seemed to contribute to the activation of double-positive thymocytes because CTLA-4 blockade resulted in the inhibition of doublepositive thymocyte deletion (182). In addition, increased surface expression of CTLA-4 in Th2 effector cells renders these cells resistant to activation-induced cell death (183). This seems to occur by recruitment of PI3K and the activation of Akt, leading to the suppression of FasL expression and the induction of the survival factor Bcl-2. These studies further suggest that CTLA-4 may indeed have the ability to induce a positive signal to enhance T cell responses but do not demonstrate CTLA-4 signaling in the absence of other confounding factors. Direct evidence of the signal plasticity of CTLA-4 was obtained with a recombinant ligand of this receptor. This ligand is a bispeciﬁc tandem single chain Fv (ScFv) molecule directed against two different human-speciﬁc epitopes of CTLA-4 recognized by antiCTLA-4 24 and 26 Abs (thus named 24:26) (92). Remarkably, 24:26 by itself activates CTLA-4-expressing primary human T cells.

Although 24:26 effectively blocks the interaction with B7, its effect is independent of B7 and occurs without concomitant TCR signaling, ruling out blockade of CTLA-4 as a mechanism of this paradoxical response. The effects of 24:26-mediated T cell activation are dependent on the expression of the TCR/CD3 complex and Lck and can be further enhanced by CD28 costimulation. The ability of CTLA-4 to deliver an activating signal upon engagement with 24:26 is the result of CTLA-4 triggering a Lck-dependent, TCR-ζ-dependent signaling cascade with phosphorylation of TCRζ and LAT, and activation of ZAP-70, PLC-γ1, ERK-1/-2, and Ca2+ ﬂuxing (92). This signal transduction event is likely triggered by ligand-induced stabilization of the PP2A:CTLA-4 association, which then activates bystander Lck, in contrast to CTLA-4 ligation with B7, which triggers dissociation of PP2A from CTLA-4 (92) (Figure 9). Consistent with this, CTLA-4 mutants that are unable to bind PP2A do not respond to 24:26, whereas okadaic acid blocks the IL-2 response to 24:26 in a dose-dependent manner. The realization of the signaling plasticity of CTLA-4 puts forth a new paradigm on how CTLA-4 can regulate immune responses. According to this paradigm, PP2A is the critical regulator of CTLA-4 function both as an inhibitor and as an activator (9, 92). In conditions of TCR-CTLA-4 coligation, PP2A is phosphorylated and subsequently dissociates from the tail of CTLA-4, leading to T cell inhibition, whereas engagement of CTLA-4 with 24:26 stabilizes the association of PP2A, leading to T cell activation. We are currently investigating the functional importance of ligand-dependent differential association of PP2A to CTLA-4. So far, three factors appear to be important for the signaling plasticity of CTLA-4. First, the bivalency of CTLA-4 homodimers on the cell surface is important for the oligomerization of CTLA-4. Second, the ligand-dependent oligomerization of CTLA-4 seems to play a critical role in the ability of CTLA-4 to www.annualreviews.org • CTLA- 4 Function

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Figure 9 Signaling plasticity of CTLA-4 correlates with its differential association with PP2A. TCR-CD28 coligation results in T cell activation and induction of CTLA-4 expression. Twenty-four to 48 h after activation, CTLA-4 is expressed on the T cell surface associated with PP2A, which blocks its inhibitory function. Upon TCR-CTLA-4 coligation, PP2A is phosphorylated (circled P) and dissociates from CTLA-4. This correlates with CTLA-4 negative signaling and inhibition of T cell activation. Alternatively, CTLA-4 ligation with 24:26, by itself, stabilizes the association between PP2A and CTLA-4, and this correlates with CTLA-4-mediated T cell activation in a Lck-dependent, TCR-ζ-dependent manner.

associate with signaling molecules. Lastly, the differential association of CTLA-4 with PP2A is key in regulating the function of CTLA-4 as either an inhibitor or activator of T cell responses. Preliminary evidence indicates that the ability of CTLA-4 to act as an inhibitor or an activator of T cells seems to correlate with the formation of structurally distinct oligomers on the cell surface in response to engagement with B7 or with this recombinant ligand (W.A. Teft and J. Madrenas, unpublished observations). The existence of an endogenous ligand with analogous function to 84

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the response seen with 24:26 is currently under investigation. Moreover, such plasticity is important when considering the development of novel therapeutic agents to enhance immunity (e.g., adjuvants). One should note that the plasticity of CTLA-4 signaling refers to the ability of CTLA-4 to deliver a positive or a negative signal, and it should not be confused with the ability to modulate a polyclonal immune response by CTLA-4 blockade. This blockade can cause immune deviation, can have differential effects on CD4+ T cells versus CD8+

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T cells, and can affect Tregs. Immune deviation is based on the representation of highafﬁnity T cell clones that would otherwise be inactivated owing to their more rapid upregulation of CTLA-4. Such an effect may translate into immune deviation from Th2 to Th1 responses (73) that can be achieved by direct modulation of cytokine signaling (184). Th1 and Th2 subsets have similar kinetics of CTLA-4 upregulation upon activation and respond similarly to CTLA-4 blockade. However, Th2 cells express signiﬁcantly higher levels of CTLA-4 than do Th1 cells, although the ratio of intracellular to surface expression is constant between both subsets (47, 183). CTLA-4 seems to downregulate predominantly Th2 cell differentiation (185, 186). Therefore, lack of CTLA-4 function correlates with Th2 cell differentiation (187, 188), and blockade of CTLA-4 function resulted in the polarization of the naive CD4+ T cells to Th1 cells (73, 189). One needs to keep in mind that the timing of modulation of CTLA-4 function and the strength of the TCR signal may determine the functional outcome (184, 190–192). For example, CTLA-4 blockade in the presence of strong TCR signals preferentially results in enhanced Th2 immune responses and loss of Th1 responses, whereas T cells receiving weak TCR signals and CTLA-4 blockade can expand both Th1 and Th2 populations (73). Another differential effect of blockade of CTLA-4 can result from differential sensitivity of CD4+ T cells versus CD8+ T cells (193). For example, the effect of CTLA-4 on

CD8+ T cells seems to be of lesser magnitude because CTLA-4−/− , MHC class I–restricted TCR transgenic mice remain healthy for a signiﬁcantly longer time than CTLA-4−/− mice, although they eventually develop a lymphoproliferative disease that appears to be mediated by a small remaining CD4+ T cell subset (65, 194). Furthermore, CD8+ T cells from CTLA-4-deﬁcient mice expressing a transgenic lymphocytic choriomeningitis virus (LCMV)-speciﬁc TCR are able to mount an immune response similar to normal mice upon LCMV infection in vivo (195). Under these types of circumstances involving polyclonal immune responses, CTLA-4 works as a threshold modulator favoring the expansion of some clones over others. Finally, CTLA-4 may also modulate the function of Tregs or APCs through B7. In some studies, in vitro blockade of CTLA-4 on Tregs abrogated the inhibition of proliferation of CTLA-4-deﬁcient responding T cells in a TGF-β-independent manner (196). In contrast, Tregs from CTLA-4-deﬁcient mice were still able to suppress responder T cells in vitro in the presence or absence of CTLA-4 blockade. In human Tregs, CTLA-4 expression has been linked to enhanced suppressor activity and higher expression of FoxP3 compared with CD4+ CD25+ CTLA-4− Tregs, and blockade of CTLA-4 in this subset resulted in a signiﬁcant but incomplete loss of Treg suppressor activity (197). CTLA-4 may also mediate the suppression of T cells through a direct interaction between CTLA4 on Tregs and B7 molecules on DCs to induce tryptophan catabolism (198–200).

SUMMARY POINTS 1. CTLA-4 is an activation-induced homodimeric glycoprotein receptor on T cells and interacts with the B7-1 (CD80)/B7-2 (CD86) ligands on the surface of an APC. 2. When engaged by B7, CTLA-4 plays a key role as a negative regulator of T cell activation, leading to downregulation of T cell responses and to the preservation of T cell homeostasis and peripheral tolerance.

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3. The mechanism of T cell inactivation by CTLA-4 involves antagonism of CD28dependent costimulation and direct negative signaling through its cytoplasmic tail. Both mechanisms may be operational through a hierarchical regulation of CTLA-4 oligomerization within lipid rafts at the immunological synapse. 4. The pathway involved in CTLA-4 signaling remains uncertain. Emerging data indicate that it is primarily regulated by the serine/threonine phosphatase PP2A acting downstream of early TCR and CD28 signaling, either by direct competition with PI3K or by inhibition of the PKB/Akt pathway.

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5. CTLA-4 has an intrinsic plasticity for signaling. This concept refers to the ability of CTLA-4 to actively deliver, by itself, a negative signal or a positive signal to a T cell, depending on the ligand it engages and its association/dissociation to PP2A. When engaged by B7, it dissociates from PP2A and acts as an inactivator of T cells. When engaged by recombinant ligands (e.g., 24:26), its association to PP2A is stabilized and it acts as an activator of T cells. Such plasticity may have important implications in the development of immunotherapeutic agents targeting CTLA-4.

ACKNOWLEDGMENTS We thank our colleagues working in the ﬁeld of CTLA-4, in particular Drs. M.L. Baroja, B.M. Carreno, M. Collins, V. Kuchroo, and V. Ling, and the members of the Madrenas laboratory for many very useful discussions. We apologize in advance to those investigators whose work on CTLA-4 could not be explicitly referenced here owing to space limitations. This work was supported by operating grants from the Canadian Institutes of Health Research (CIHR), the Kidney Foundation of Canada, and the London Health Sciences Center Multi-Organ Transplant Program. Wendy A. Teft holds a CIHR doctoral award, Mark G. Kirchhof holds a MD/PhD studentship from CIHR, and Joaqu´ın Madrenas holds a Canada Research Chair in Transplantation and Immunobiology. We dedicate this review article to the memory of Dr. Cynthia A. Chambers.

LITERATURE CITED 1. Brunet JF, Denizot F, Luciani MF, Roux-Dosseto M, Suzan M, et al. 1987. A new member of the immunoglobulin superfamily—CTLA-4. Nature 328:267–70 2. Dariavach P, Mattei MG, Golstein P, Lefranc MP. 1988. Human Ig superfamily CTLA4 gene: chromosomal localization and identity of protein sequence between murine and human CTLA-4 cytoplasmic domains. Eur. J. Immunol. 18:1901–5 3. Lindsten T, Lee KP, Harris ES, Petryniak B, Craighead N, et al. 1993. Characterization of CTLA-4 structure and expression on human T cells. J. Immunol. 151:3489–99 4. Linsley PS, Nadler SG, Bajorath J, Peach R, Leung HT, et al. 1995. Binding stoichiometry of the cytotoxic T lymphocyte-associated molecule-4 (CTLA-4). A disulﬁde-linked homodimer binds two CD86 molecules. J. Biol. Chem. 270:15417–24 5. Ostrov DA, Shi W, Schwartz JC, Almo SC, Nathenson SG. 2000. Structure of murine CTLA-4 and its role in modulating T cell responsiveness. Science 290:816–19 86

Teft

·

Kirchhof

·

Madrenas

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ANRV270-IY24-03

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13 February 2006

11:54

6. Metzler WJ, Bajorath J, Fenderson W, Shaw SY, Constantine KL, et al. 1997. Solution structure of human CTLA-4 and delineation of a CD80/CD86 binding site conserved in CD28. Nat. Struct. Biol. 4:527–31 7. Darlington PJ, Kirchhof MG, Criado G, Sondhi J, Madrenas J. 2005. Hierarchical regulation of CTLA-4 dimer-based lattice formation and its biological relevance for T cell inactivation. J. Immunol. 175:996–1004 8. Baroja ML, Darlington PJ, Carreno BM, Madrenas J. 2000. Inhibition of T cell activation by CTLA-4: truths and red herrings. Mod. Asp. Immunobiol. 1:169–73 9. Baroja ML, Vijayakrishnan L, Bettelli E, Darlington PJ, Chau TA, et al. 2002. Inhibition of CTLA-4 function by the regulatory subunit of serine/threonine phosphatase 2A. J. Immunol. 168:5070–78 10. Lee KM, Chuang E, Grifﬁn M, Khattri R, Hong DK, et al. 1998. Molecular basis of T cell inactivation by CTLA-4. Science 282:2263–66 11. Schneider H, Prasad KV, Shoelson SE, Rudd CE. 1995. CTLA-4 binding to the lipid kinase phosphatidylinositol 3-kinase in T cells. J. Exp. Med. 181:351–55 12. Shiratori T, Miyatake S, Ohno H, Nakaseko C, Isono K, et al. 1997. Tyrosine phosphorylation controls internalization of CTLA-4 by regulating its interaction with clathrinassociated adaptor complex AP-2. Immunity 6:583–89 13. Chambers CA, Kuhns MS, Egen JG, Allison JP. 2001. CTLA-4-mediated inhibition in regulation of T cell responses: mechanisms and manipulation in tumor immunotherapy. Annu. Rev. Immunol. 19:565–94 14. Salomon B, Bluestone JA. 2001. Complexities of CD28/B7: CTLA-4 costimulatory pathways in autoimmunity and transplantation. Annu. Rev. Immunol. 19:225–52 15. Rudd CE, Martin M, Schneider H. 2002. CTLA-4 negative signaling via lipid rafts: a new perspective. Sci. STKE 2002:PE18 16. Alegre ML, Frauwirth KA, Thompson CB. 2001. T-cell regulation by CD28 and CTLA-4. Nat. Rev. Immunol. 1:220–28 17. Ling V, Wu PW, Finnerty HF, Sharpe AH, Gray GS, Collins M. 1999. Complete sequence determination of the mouse and human CTLA4 gene loci: cross-species DNA sequence similarity beyond exon borders. Genomics 60:341–55 18. Ueda H, Howson JM, Esposito L, Heward J, Snook H, et al. 2003. Association of the T-cell regulatory gene CTLA4 with susceptibility to autoimmune disease. Nature 423:506–11 19. Vijayakrishnan L, Slavik JM, Illes Z, Greenwald RJ, Rainbow D, et al. 2004. An autoimmune disease-associated CTLA-4 splice variant lacking the B7 binding domain signals negatively in T cells. Immunity 20:563–75 20. Magistrelli G, Jeannin P, Herbault N, Benoit DC, Gauchat JF, et al. 1999. A soluble form of CTLA-4 generated by alternative splicing is expressed by nonstimulated human T cells. Eur. J. Immunol. 29:3596–602 21. Oaks MK, Hallett KM, Penwell RT, Stauber EC, Warren SJ, Tector AJ. 2000. A native soluble form of CTLA-4. Cell. Immunol. 201:144–53 22. Perkins D, Wang Z, Donovan C, He H, Mark D, et al. 1996. Regulation of CTLA-4 expression during T cell activation. J. Immunol. 156:4154–59 23. Harper K, Balzano C, Rouvier E, Mattei MG, Luciani MF, Golstein P. 1991. CTLA-4 and CD28 activated lymphocyte molecules are closely related in both mouse and human as to sequence, message expression, gene structure, and chromosomal location. J. Immunol. 147:1037–44 24. Takahashi T, Tagami T, Yamazaki S, Uede T, Shimizu J, et al. 2000. Immunologic selftolerance maintained by CD25+ CD4+ regulatory T cells constitutively expressing cytotoxic T lymphocyte-associated antigen 4. J. Exp. Med. 192:303–10 www.annualreviews.org • CTLA- 4 Function

87

ARI

13 February 2006

11:54

25. Jago CB, Yates J, Camara NO, Lechler RI, Lombardi G. 2004. Differential expression of CTLA-4 among T cell subsets. Clin. Exp. Immunol. 136:463–71 26. Wagner DH Jr, Hagman J, Linsley PS, Hodsdon W, Freed JH, Newell MK. 1996. Rescue of thymocytes from glucocorticoid-induced cell death mediated by CD28/CTLA-4 costimulatory interactions with B7-1/B7-2. J. Exp. Med. 184:1631–38 27. Cilio CM, Daws MR, Malashicheva A, Sentman CL, Holmberg D. 1998. Cytotoxic T lymphocyte antigen 4 is induced in the thymus upon in vivo activation and its blockade prevents anti-CD3-mediated depletion of thymocytes. J. Exp. Med. 188:1239–46 28. Pioli C, Gatta L, Ubaldi V, Doria G. 2000. Inhibition of IgG1 and IgE production by stimulation of the B cell CTLA-4 receptor. J. Immunol. 165:5530–36 29. Pistillo MP, Tazzari PL, Palmisano GL, Pierri I, Bolognesi A, et al. 2003. CTLA-4 is not restricted to the lymphoid cell lineage and can function as a target molecule for apoptosis induction of leukemic cells. Blood 101:202–9 30. Kaufman KA, Bowen JA, Tsai AF, Bluestone JA, Hunt JS, Ober C. 1999. The CTLA-4 gene is expressed in placental ﬁbroblasts. Mol. Hum. Reprod. 5:84–87 31. Wang XB, Giscombe R, Yan Z, Heiden T, Xu D, Lefvert AK. 2002. Expression of CTLA-4 by human monocytes. Scand. J. Immunol. 55:53–60 32. Ling V, Munroe RC, Murphy EA, Gray GS. 1998. Embryonic stem cells and embryoid bodies express lymphocyte costimulatory molecules. Exp. Cell Res. 241:55–65 33. Miller RE, Fayen JD, Mohammad SF, Stein K, Kadereit S, et al. 2002. Reduced CTLA4 protein and messenger RNA expression in umbilical cord blood T lymphocytes. Exp. Hematol. 30:738–44 34. Finn PW, He H, Wang Y, Wang Z, Guan G, et al. 1997. Synergistic induction of CTLA-4 expression by costimulation with TCR plus CD28 signals mediated by increased transcription and messenger ribonucleic acid stability. J. Immunol. 158:4074–81 35. Vendetti S, Riccomi A, Sacchi A, Gatta L, Pioli C, De Magistris MT. 2002. Cyclic adenosine 5’-monophosphate and calcium induce CD152 (CTLA-4) up-regulation in resting CD4+ T lymphocytes. J. Immunol. 169:6231–35 36. Shaw G, Kamen R. 1986. A conserved AU sequence from the 3 untranslated region of GM-CSF mRNA mediates selective mRNA degradation. Cell 46:659–67 37. Wang XB, Zheng CY, Giscombe R, Lefvert AK. 2001. Regulation of surface and intracellular expression of CTLA-4 on human peripheral T cells. Scand. J. Immunol. 54:453–58 38. Alegre ML, Noel PJ, Eisfelder BJ, Chuang E, Clark MR, et al. 1996. Regulation of surface and intracellular expression of CTLA4 on mouse T cells. J. Immunol. 157:4762–70 39. Chuang E, Alegre ML, Duckett CS, Noel PJ, Vander Heiden MG, Thompson CB. 1997. Interaction of CTLA-4 with the clathrin-associated protein AP50 results in ligandindependent endocytosis that limits cell surface expression. J. Immunol. 159:144–51 40. Schneider H, Martin M, Agarraberes FA, Yin L, Rapoport I, et al. 1999. Cytolytic T lymphocyte-associated antigen-4 and the TCRζ/CD3 complex, but not CD28, interact with clathrin adaptor complexes AP-1 and AP-2. J. Immunol. 163:1868–79 41. Iida T, Ohno H, Nakaseko C, Sakuma M, Takeda-Ezaki M, et al. 2000. Regulation of cell surface expression of CTLA-4 by secretion of CTLA-4-containing lysosomes upon activation of CD4+ T cells. J. Immunol. 165:5062–68 42. Lafage-Pochitaloff M, Costello R, Couez D, Simonetti J, Mannoni P, et al. 1990. Human CD28 and CTLA-4 Ig superfamily genes are located on chromosome 2 at bands q33-q34. Immunogenetics 31:198–201 43. Carreno BM, Collins M. 2002. The B7 family of ligands and its receptors: new pathways for costimulation and inhibition of immune responses. Annu. Rev. Immunol. 20:29–53

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ANRV270-IY24-03

ARI

13 February 2006

11:54

44. Howard TA, Rochelle JM, Seldin MF. 1991. CD28 and CTLA-4, two related members of the Ig supergene family, are tightly linked on proximal mouse chromosome 1. Immunogenetics 33:74–76 45. Choi IS, Hash SM, Collisson EW. 2000. Molecular cloning and expression of feline CD28 and CTLA-4 cDNA. Vet. Immunol. Immunopathol. 76:45–59 46. Palmisano GL, Tazzari PL, Cozzi E, Bolognesi A, Polito L, et al. 2004. Expression of CTLA-4 in nonhuman primate lymphocytes and its use as a potential target for speciﬁc immunotoxin-mediated apoptosis: results of in vitro studies. Clin. Exp. Immunol. 135:259– 66 47. Alegre ML, Shiels H, Thompson CB, Gajewski TF. 1998. Expression and function of CTLA-4 in Th1 and Th2 cells. J. Immunol. 161:3347–56 48. Carreno BM, Bennett F, Chau TA, Ling V, Luxenberg D, et al. 2000. CTLA-4 (CD152) can inhibit T cell activation by two different mechanisms depending on its level of cell surface expression. J. Immunol. 165:1352–56 49. Nistico L, Buzzetti R, Pritchard LE, Van der Auwera B, Giovannini C, et al. 1996. The CTLA-4 gene region of chromosome 2q33 is linked to, and associated with, type 1 diabetes. Belgian Diabetes Registry. Hum. Mol. Genet. 5:1075–80 50. Deichmann K, Heinzmann A, Bruggenolte E, Forster J, Kuehr J. 1996. An Mse I RFLP in the human CTLA4 promotor. Biochem. Biophys. Res. Commun. 225:817–18 51. Donner H, Rau H, Walﬁsh PG, Braun J, Siegmund T, et al. 1997. CTLA4 alanine-17 confers genetic susceptibility to Graves’ disease and to type 1 diabetes mellitus. J. Clin. Endocrinol. Metab. 82:143–46 52. Wang XB, Zhao X, Giscombe R, Lefvert AK. 2002. A CTLA-4 gene polymorphism at position -318 in the promoter region affects the expression of protein. Genes Immun. 3:233–34 53. Kristiansen OP, Larsen ZM, Pociot F. 2000. CTLA-4 in autoimmune diseases—a general susceptibility gene to autoimmunity? Genes Immun. 1:170–84 54. Anjos S, Nguyen A, Ounissi-Benkalha H, Tessier MC, Polychronakos C. 2002. A common autoimmunity predisposing signal peptide variant of the cytotoxic T-lymphocyte antigen 4 results in inefﬁcient glycosylation of the susceptibility allele. J. Biol. Chem. 277:46478–86 55. Huang D, Giscombe R, Zhou Y, Pirskanen R, Lefvert AK. 2000. Dinucleotide repeat expansion in the CTLA-4 gene leads to T cell hyper-reactivity via the CD28 pathway in myasthenia gravis. J. Neuroimmunol. 105:69–77 56. Wang XB, Kakoulidou M, Giscombe R, Qiu Q, Huang D, et al. 2002. Abnormal expression of CTLA-4 by T cells from patients with myasthenia gravis: effect of an AT-rich gene sequence. J. Neuroimmunol. 130:224–32 57. Wu Y, Guo Y, Huang A, Zheng P, Liu Y. 1997. CTLA-4-B7 interaction is sufﬁcient to costimulate T cell clonal expansion. J. Exp. Med. 185:1327–35 58. Damle NK, Klussman K, Leytze G, Myrdal S, Aruffo A, et al. 1994. Costimulation of T lymphocytes with integrin ligands intercellular adhesion molecule-1 or vascular cell adhesion molecule-1 induces functional expression of CTLA-4, a second receptor for B7. J. Immunol. 152:2686–97 59. Linsley PS, Greene JL, Tan P, Bradshaw J, Ledbetter JA, et al. 1992. Coexpression and functional cooperation of CTLA-4 and CD28 on activated T lymphocytes. J. Exp. Med. 176:1595–604 60. Walunas TL, Lenschow DJ, Bakker CY, Linsley PS, Freeman GJ, et al. 1994. CTLA-4 can function as a negative regulator of T cell activation. Immunity 1:405–13 61. Krummel MF, Allison JP. 1995. CD28 and CTLA-4 have opposing effects on the response of T cells to stimulation. J. Exp. Med. 182:459–65 www.annualreviews.org • CTLA- 4 Function

89

ARI

13 February 2006

11:54

62. Tivol EA, Borriello F, Schweitzer AN, Lynch WP, Bluestone JA, Sharpe AH. 1995. Loss of CTLA-4 leads to massive lymphoproliferation and fatal multiorgan tissue destruction, revealing a critical negative regulatory role of CTLA-4. Immunity 3:541–47 63. Waterhouse P, Penninger JM, Timms E, Wakeham A, Shahinian A, et al. 1995. Lymphoproliferative disorders with early lethality in mice deﬁcient in CTLA-4. Science 270:985– 88 64. Chambers CA, Cado D, Truong T, Allison JP. 1997. Thymocyte development is normal in CTLA-4-deﬁcient mice. Proc. Natl. Acad. Sci. USA 94:9296–301 65. Waterhouse P, Bachmann MF, Penninger JM, Ohashi PS, Mak TW. 1997. Normal thymic selection, normal viability and decreased lymphoproliferation in T cell receptor-transgenic CTLA-4-deﬁcient mice. Eur. J. Immunol. 27:1887–92 66. Takahashi S, Kataoka H, Hara S, Yokosuka T, Takase K, et al. 2005. In vivo overexpression of CTLA-4 suppresses lymphoproliferative diseases and thymic negative selection. Eur. J. Immunol. 35:399–407 67. Allison JP, Krummel MF. 1995. The yin and yang of T cell costimulation. Science 270:932– 33 68. Linsley PS, Brady W, Urnes M, Grosmaire LS, Damle NK, Ledbetter JA. 1991. CTLA-4 is a second receptor for the B cell activation antigen B7. J. Exp. Med. 174:561–69 69. Tivol EA, Boyd SD, McKeon S, Borriello F, Nickerson P, et al. 1997. CTLA4Ig prevents lymphoproliferation and fatal multiorgan tissue destruction in CTLA-4-deﬁcient mice. J. Immunol. 158:5091–94 70. Bachmann MF, Kohler G, Ecabert B, Mak TW, Kopf M. 1999. Cutting edge: Lymphoproliferative disease in the absence of CTLA-4 is not T cell autonomous. J. Immunol. 163:1128–31 71. Tivol EA, Gorski J. 2002. Re-establishing peripheral tolerance in the absence of CTLA-4: complementation by wild-type T cells points to an indirect role for CTLA-4. J. Immunol. 169:1852–58 72. Bachmann MF, Gallimore A, Jones E, Ecabert B, Acha-Orbea H, Kopf M. 2001. Normal pathogen-speciﬁc immune responses mounted by CTLA-4-deﬁcient T cells: a paradigm reconsidered. Eur. J. Immunol. 31:450–58 73. Anderson DE, Bieganowska KD, Bar-Or A, Oliveira EM, Carreno B, et al. 2000. Paradoxical inhibition of T-cell function in response to CTLA-4 blockade; heterogeneity within the human T-cell population. Nat. Med. 6:211–14 74. Linsley PS, Clark EA, Ledbetter JA. 1990. T-cell antigen CD28 mediates adhesion with B cells by interacting with activation antigen B7/BB-1. Proc. Natl. Acad. Sci. USA 87:5031– 35 75. Freeman GJ, Borriello F, Hodes RJ, Reiser H, Gribben JG, et al. 1993. Murine B7-2, an alternative CTLA4 counter-receptor that costimulates T cell proliferation and interleukin 2 production. J. Exp. Med. 178:2185–92 76. Freeman GJ, Gribben JG, Boussiotis VA, Ng JW, Restivo VA Jr, et al. 1993. Cloning of B7-2: a CTLA-4 counter-receptor that costimulates human T cell proliferation. Science 262:909–11 77. Linsley PS, Greene JL, Brady W, Bajorath J, Ledbetter JA, Peach R. 1994. Human B7-1 (CD80) and B7-2 (CD86) bind with similar avidities but distinct kinetics to CD28 and CTLA-4 receptors. Immunity 1:793–801 78. Collins AV, Brodie DW, Gilbert RJ, Iaboni A, Manso-Sancho R, et al. 2002. The interaction properties of costimulatory molecules revisited. Immunity 17:201–10

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ANRV270-IY24-03

ARI

13 February 2006

11:54

79. van der Merwe PA, Bodian DL, Daenke S, Linsley P, Davis SJ. 1997. CD80 (B7-1) binds both CD28 and CTLA-4 with a low afﬁnity and very fast kinetics. J. Exp. Med. 185:393– 403 80. van der Merwe PA, Davis SJ. 2003. Molecular interactions mediating T cell antigen recognition. Annu. Rev. Immunol. 21:659–84 81. Stamper CC, Zhang Y, Tobin JF, Erbe DV, Ikemizu S, et al. 2001. Crystal structure of the B7-1/CTLA-4 complex that inhibits human immune responses. Nature 410:608–11 82. Zhang X, Schwartz JC, Almo SC, Nathenson SG. 2003. Crystal structure of the receptorbinding domain of human B7-2: insights into organization and signaling. Proc. Natl. Acad. Sci. USA 100:2586–91 83. Balzano C, Buonavista N, Rouvier E, Golstein P. 1992. CTLA-4 and CD28: similar proteins, neighbouring genes. Int. J. Cancer Suppl. 7:28–32 84. Lenschow DJ, Su GH, Zuckerman LA, Nabavi N, Jellis CL, et al. 1993. Expression and functional signiﬁcance of an additional ligand for CTLA-4. Proc. Natl. Acad. Sci. USA 90:11054–58 85. Hathcock KS, Laszlo G, Pucillo C, Linsley P, Hodes RJ. 1994. Comparative analysis of B7-1 and B7-2 costimulatory ligands: expression and function. J. Exp. Med. 180:631–40 86. Lenschow DJ, Walunas TL, Bluestone JA. 1996. CD28/B7 system of T cell costimulation. Annu. Rev. Immunol. 14:233–58 87. Evans EJ, Esnouf RM, Manso-Sancho R, Gilbert RJ, James JR, et al. 2005. Crystal structure of a soluble CD28-Fab complex. Nat. Immunol. 6:271–79 88. Pentcheva-Hoang T, Egen JG, Wojnoonski K, Allison JP. 2004. B7-1 and B7-2 selectively recruit CTLA-4 and CD28 to the immunological synapse. Immunity 21:401–13 89. Kuchroo VK, Das MP, Brown JA, Ranger AM, Zamvil SS, et al. 1995. B7-1 and B7-2 costimulatory molecules activate differentially the Th1/Th2 developmental pathways: application to autoimmune disease therapy. Cell 80:707–18 90. Chen L. 2004. Co-inhibitory molecules of the B7-CD28 family in the control of T-cell immunity. Nat. Rev. Immunol. 4:336–47 91. Chikuma S, Abbas AK, Bluestone JA. 2005. B7-independent inhibition of T cells by CTLA-4. J. Immunol. 175:177–81 92. Madrenas J, Chau LA, Teft WA, Wu PW, Jussif J, et al. 2004. Conversion of CTLA-4 from inhibitor to activator of T cells with a bispeciﬁc tandem single-chain Fv ligand. J. Immunol. 172:5948–56 93. Mukherjee S, Ahmed A, Nandi D. 2005. CTLA4-CD80/CD86 interactions on primary mouse CD4+ T cells integrate signal-strength information to modulate activation with Concanavalin A. J. Leukoc. Biol. 78:144–57 94. Greene JL, Leytze GM, Emswiler J, Peach R, Bajorath J, et al. 1996. Covalent dimerization of CD28/CTLA-4 and oligomerization of CD80/CD86 regulate T cell costimulatory interactions. J. Biol. Chem. 271:26762–71 95. Schwartz JC, Zhang X, Fedorov AA, Nathenson SG, Almo SC. 2001. Structural basis for co-stimulation by the human CTLA-4/B7-2 complex. Nature 410:604–8 96. Schwartz JC, Zhang X, Nathenson SG, Almo SC. 2002. Structural mechanisms of costimulation. Nat. Immunol. 3:427–34 97. Chun T, Choi HJ, Chung YH. 2004. Two different forms of human CTLA-4 proteins following peripheral T cell activation. Immunol. Lett. 91:213–20 98. Luo RZ, Beniac DR, Fernandes A, Yip CC, Ottensmeyer FP. 1999. Quaternary structure of the insulin-insulin receptor complex. Science 285:1077–80 www.annualreviews.org • CTLA- 4 Function

91

ARI

13 February 2006

11:54

99. Cunningham BC, Ultsch M, De Vos AM, Mulkerrin MG, Clauser KR, Wells JA. 1991. Dimerization of the extracellular domain of the human growth hormone receptor by a single hormone molecule. Science 254:821–25 100. Baroja ML, Madrenas J. 2003. Viewpoint: therapeutic implications of CTLA-4 compartmentalization. Am. J. Transplant. 3:919–26 101. Chikuma S, Bluestone JA. 2003. CTLA-4 and tolerance: the biochemical point of view. Immunol. Res. 28:241–53 102. Egen JG, Allison JP. 2002. Cytotoxic T lymphocyte antigen-4 accumulation in the immunological synapse is regulated by TCR signal strength. Immunity 16:23–35 103. Darlington PJ, Baroja ML, Chau TA, Siu E, Ling V, et al. 2002. Surface cytotoxic T lymphocyte-associated antigen 4 partitions within lipid rafts and relocates to the immunological synapse under conditions of inhibition of T cell activation. J. Exp. Med. 195:1337–47 104. Mead KI, Zheng Y, Manzotti CN, Perry LC, Liu MK, et al. 2005. Exocytosis of CTLA-4 is dependent on phospholipase D and ADP ribosylation factor-1 and stimulated during activation of regulatory T cells. J. Immunol. 174:4803–11 105. Zhang Y, Allison JP. 1997. Interaction of CTLA-4 with AP50, a clathrin-coated pit adaptor protein. Proc. Natl. Acad. Sci. USA 94:9273–78 106. Bradshaw JD, Lu P, Leytze G, Rodgers J, Schieven GL, et al. 1997. Interaction of the cytoplasmic tail of CTLA-4 (CD152) with a clathrin-associated protein is negatively regulated by tyrosine phosphorylation. Biochemistry 36:15975–82 107. Baroja ML, Luxenberg D, Chau T, Ling V, Strathdee CA, et al. 2000. The inhibitory function of CTLA-4 does not require its tyrosine phosphorylation. J. Immunol. 164:49–55 108. Davis DM, Dustin ML. 2004. What is the importance of the immunological synapse? Trends Immunol. 25:323–27 109. Jacobelli J, Andres PG, Boisvert J, Krummel MF. 2004. New views of the immunological synapse: variations in assembly and function. Curr. Opin. Immunol. 16:345–52 110. Dustin ML. 2003. Coordination of T cell activation and migration through formation of the immunological synapse. Ann. NY Acad. Sci. 987:51–59 111. Kupfer A, Swain SL, Singer SJ. 1987. The speciﬁc direct interaction of helper T cells and antigen-presenting B cells. II. Reorientation of the microtubule organizing center and reorganization of the membrane-associated cytoskeleton inside the bound helper T cells. J. Exp. Med. 165:1565–80 112. Linsley PS, Bradshaw J, Greene J, Peach R, Bennett KL, Mittler RS. 1996. Intracellular trafﬁcking of CTLA-4 and focal localization towards sites of TCR engagement. Immunity 4:535–43 113. Taner SB, Onfelt B, Pirinen NJ, McCann FE, Magee AI, Davis DM. 2004. Control of immune responses by trafﬁcking cell surface proteins, vesicles and lipid rafts to and from the immunological synapse. Trafﬁc 5:651–61 114. Vogt AB, Spindeldreher S, Kropshofer H. 2002. Clustering of MHC-peptide complexes prior to their engagement in the immunological synapse: lipid raft and tetraspan microdomains. Immunol. Rev. 189:136–51 115. Miceli MC, Moran M, Chung CD, Patel VP, Low T, Zinnanti W. 2001. Co-stimulation and counter-stimulation: lipid raft clustering controls TCR signaling and functional outcomes. Semin. Immunol. 13:115–28 116. Chikuma S, Imboden JB, Bluestone JA. 2003. Negative regulation of T cell receptor-lipid raft interaction by cytotoxic T lymphocyte-associated antigen 4. J. Exp. Med. 197:129–35

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117. Leung HT, Bradshaw J, Cleaveland JS, Linsley PS. 1995. Cytotoxic T lymphocyteassociated molecule-4, a high-avidity receptor for CD80 and CD86, contains an intracellular localization motif in its cytoplasmic tail. J. Biol. Chem. 270:25107–14 118. Cinek T, Sadra A, Imboden JB. 2000. Cutting edge: tyrosine-independent transmission of inhibitory signals by CTLA-4. J. Immunol. 164:5–8 119. Masteller EL, Chuang E, Mullen AC, Reiner SL, Thompson CB. 2000. Structural analysis of CTLA-4 function in vivo. J. Immunol. 164:5319–27 120. Fallarino F, Fields PE, Gajewski TF. 1998. B7-1 engagement of cytotoxic T lymphocyte antigen 4 inhibits T cell activation in the absence of CD28. J. Exp. Med. 188:205–10 121. Fraser JH, Rincon M, McCoy KD, Le Gros G. 1999. CTLA4 ligation attenuates AP-1, NFAT and NF-κB activity in activated T cells. Eur. J. Immunol. 29:838–44 122. Lin H, Rathmell JC, Gray GS, Thompson CB, Leiden JM, Alegre ML. 1998. Cytotoxic T lymphocyte antigen 4 (CTLA4) blockade accelerates the acute rejection of cardiac allografts in CD28-deﬁcient mice: CTLA4 can function independently of CD28. J. Exp. Med. 188:199–204 123. Greenwald RJ, Boussiotis VA, Lorsbach RB, Abbas AK, Sharpe AH. 2001. CTLA-4 regulates induction of anergy in vivo. Immunity 14:145–55 124. Wells AD, Walsh MC, Bluestone JA, Turka LA. 2001. Signaling through CD28 and CTLA-4 controls two distinct forms of T cell anergy. J. Clin. Invest. 108:895–903 125. Vanasek TL, Khoruts A, Zell T, Mueller DL. 2001. Antagonistic roles for CTLA-4 and the mammalian target of rapamycin in the regulation of clonal anergy: enhanced cell cycle progression promotes recall antigen responsiveness. J. Immunol. 167:5636–44 126. Eagar TN, Karandikar NJ, Bluestone JA, Miller SD. 2002. The role of CTLA-4 in induction and maintenance of peripheral T cell tolerance. Eur. J. Immunol. 32:972–81 127. Eagar TN, Turley DM, Padilla J, Karandikar NJ, Tan L, et al. 2004. CTLA-4 regulates expansion and differentiation of Th1 cells following induction of peripheral T cell tolerance. J. Immunol. 172:7442–50 128. Schwartz RH. 2003. T cell anergy. Annu. Rev. Immunol. 21:305–34 129. Krummel MF, Allison JP. 1996. CTLA-4 engagement inhibits IL-2 accumulation and cell cycle progression upon activation of resting T cells. J. Exp. Med. 183:2533–40 130. Rudd CE, Schneider H. 2003. Unifying concepts in CD28, ICOS and CTLA4 co-receptor signalling. Nat. Rev. Immunol.. 3:544–56 131. Olsson C, Riesbeck K, Dohlsten M, Michaelsson E. 1999. CTLA-4 ligation suppresses CD28-induced NF-κB and AP-1 activity in mouse T cell blasts. J. Biol. Chem. 274:14400– 5 132. Brunner MC, Chambers CA, Chan FK, Hanke J, Winoto A, Allison JP. 1999. CTLA-4mediated inhibition of early events of T cell proliferation. J. Immunol. 162:5813–20 133. Li D, Gal I, Vermes C, Alegre ML, Chong AS, et al. 2004. Cutting edge: Cbl-b: one of the key molecules tuning CD28- and CTLA-4-mediated T cell costimulation. J. Immunol. 173:7135–39 134. Bachmaier K, Krawczyk C, Kozieradzki I, Kong YY, Sasaki T, et al. 2000. Negative regulation of lymphocyte activation and autoimmunity by the molecular adaptor Cbl-b. Nature 403:211–16 135. Chiang YJ, Kole HK, Brown K, Naramura M, Fukuhara S, et al. 2000. Cbl-b regulates the CD28 dependence of T-cell activation. Nature 403:216–20 136. Michel F, Mangino G, Attal-Bonnefoy G, Tuosto L, Alcover A, et al. 2000. CD28 utilizes Vav-1 to enhance TCR-proximal signaling and NF-AT activation. J. Immunol. 165:3820– 29 www.annualreviews.org • CTLA- 4 Function

93

ARI

13 February 2006

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137. Salojin KV, Zhang J, Delovitch TL. 1999. TCR and CD28 are coupled via ZAP-70 to the activation of the Vav/Rac-1-/PAK-1/p38 MAPK signaling pathway. J. Immunol. 163:844– 53 138. Carey KD, Dillon TJ, Schmitt JM, Baird AM, Holdorf AD, et al. 2000. CD28 and the tyrosine kinase lck stimulate mitogen-activated protein kinase activity in T cells via inhibition of the small G protein Rap1. Mol. Cell. Biol. 20:8409–19 139. Dillon TJ, Carey KD, Wetzel SA, Parker DC, Stork PJ. 2005. Regulation of the small GTPase Rap1 and extracellular signal-regulated kinases by the costimulatory molecule CTLA-4. Mol. Cell. Biol. 25:4117–28 140. Miyatake S, Nakaseko C, Umemori H, Yamamoto T, Saito T. 1998. Src family tyrosine kinases associate with and phosphorylate CTLA-4 (CD152). Biochem. Biophys. Res. Commun. 249:444–48 141. Schneider H, Schwartzberg PL, Rudd CE. 1998. Resting lymphocyte kinase (Rlk/Txk) phosphorylates the YVKM motif and regulates PI 3-kinase binding to T-cell antigen CTLA-4. Biochem. Biophys. Res. Commun. 252:14–19 142. Chuang E, Lee KM, Robbins MD, Duerr JM, Alegre ML, et al. 1999. Regulation of cytotoxic T lymphocyte-associated molecule-4 by Src kinases. J. Immunol. 162:1270–77 143. Chikuma S, Murakami M, Tanaka K, Uede T. 2000. Janus kinase 2 is associated with a box 1-like motif and phosphorylates a critical tyrosine residue in the cytoplasmic region of cytotoxic T lymphocyte associated molecule-4. J. Cell. Biochem. 78:241–50 144. Chambers CA, Allison JP. 1996. The role of tyrosine phosphorylation and PTP-1C in CTLA-4 signal transduction. Eur. J. Immunol. 26:3224–29 145. Marengere LE, Waterhouse P, Duncan GS, Mittrucker HW, Feng GS, Mak TW. 1996. Regulation of T cell receptor signaling by tyrosine phosphatase SYP association with CTLA-4. Science 272:1170–73 146. Yi LA, Hajialiasgar S, Chuang E. 2004. Tyrosine-mediated inhibitory signals contribute to CTLA-4 function in vivo. Int. Immunol. 16:539–47 147. Siu E, Carreno BM, Madrenas J. 2003. TCR subunit speciﬁcity of CTLA-4-mediated signaling. J. Leukoc. Biol. 74:1102–7 148. Nakaseko C, Miyatake S, Iida T, Hara S, Abe R, et al. 1999. Cytotoxic T lymphocyte antigen 4 (CTLA-4) engagement delivers an inhibitory signal through the membraneproximal region in the absence of the tyrosine motif in the cytoplasmic tail. J. Exp. Med. 190:765–74 149. Tonks NK, Neel BG. 1996. From form to function: signaling by protein tyrosine phosphatases. Cell 87:365–68 150. Schneider H, Rudd CE. 2000. Tyrosine phosphatase SHP-2 binding to CTLA-4: absence of direct YVKM/YFIP motif recognition. Biochem. Biophys. Res. Commun. 269:279–83 151. Schneider H, da Rocha Dias S, Hu H, Rudd CE. 2001. A regulatory role for cytoplasmic YVKM motif in CTLA-4 inhibition of TCR signaling. Eur. J. Immunol. 31:2042–50 152. Frearson JA, Alexander DR. 1998. The phosphotyrosine phosphatase SHP-2 participates in a multimeric signaling complex and regulates T cell receptor (TCR) coupling to the Ras/mitogen-activated protein kinase (MAPK) pathway in Jurkat T cells. J. Exp. Med. 187:1417–26 153. Guntermann C, Alexander DR. 2002. CTLA-4 suppresses proximal TCR signaling in resting human CD4+ T cells by inhibiting ZAP-70 Tyr319 phosphorylation: a potential role for tyrosine phosphatases. J. Immunol. 168:4420–29 154. Chuang E, Fisher TS, Morgan RW, Robbins MD, Duerr JM, et al. 2000. The CD28 and CTLA-4 receptors associate with the serine/threonine phosphatase PP2A. Immunity 13:313–22

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ANRV270-IY24-03

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155. Calvo CR, Amsen D, Kruisbeek AM. 1997. Cytotoxic T lymphocyte antigen 4 (CTLA-4) interferes with extracellular signal-regulated kinase (ERK) and Jun NH2-terminal kinase (JNK) activation, but does not affect phosphorylation of T cell receptor ζ and ZAP70. J. Exp. Med. 186:1645–53 156. Schneider H, Mandelbrot DA, Greenwald RJ, Ng F, Lechler R, et al. 2002. Cutting edge: CTLA-4 (CD152) differentially regulates mitogen-activated protein kinases (extracellular signal-regulated kinase and c-Jun N-terminal kinase) in CD4+ T cells from receptor/ligand-deﬁcient mice. J. Immunol. 169:3475–79 157. Bos JL, de Bruyn K, Enserink J, Kuiperij B, Rangarajan S, et al. 2003. The role of Rap1 in integrin-mediated cell adhesion. Biochem. Soc. Trans. 31:83–86 158. Stork PJ, Schmitt JM. 2002. Crosstalk between cAMP and MAP kinase signaling in the regulation of cell proliferation. Trends Cell Biol. 12:258–66 159. Sontag E. 2001. Protein phosphatase 2A: the Trojan Horse of cellular signaling. Cell. Signal. 13:7–16 160. Lechward K, Awotunde OS, Swiatek W, Muszynska G. 2001. Protein phosphatase 2A: variety of forms and diversity of functions. Acta Biochim. Pol. 48:921–33 161. Janssens V, Goris J. 2001. Protein phosphatase 2A: a highly regulated family of serine/threonine phosphatases implicated in cell growth and signalling. Biochem. J. 353:417– 39 162. Groves MR, Hanlon N, Turowski P, Hemmings BA, Barford D. 1999. The structure of the protein phosphatase 2A PR65/A subunit reveals the conformation of its 15 tandemly repeated HEAT motifs. Cell 96:99–110 163. Ruediger R, Hentz M, Fait J, Mumby M, Walter G. 1994. Molecular model of the A subunit of protein phosphatase 2A: interaction with other subunits and tumor antigens. J. Virol. 68:123–29 164. Kremmer E, Ohst K, Kiefer J, Brewis N, Walter G. 1997. Separation of PP2A core enzyme and holoenzyme with monoclonal antibodies against the regulatory A subunit: abundant expression of both forms in cells. Mol. Cell. Biol. 17:1692–701 165. Cazzolli R, Carpenter L, Biden TJ, Schmitz-Peiffer C. 2001. A role for protein phosphatase 2A-like activity, but not atypical protein kinase Cζ, in the inhibition of protein kinase B/Akt and glycogen synthesis by palmitate. Diabetes 50:2210–18 166. Resjo S, Goransson O, Harndahl L, Zolnierowicz S, Manganiello V, Degerman E. 2002. Protein phosphatase 2A is the main phosphatase involved in the regulation of protein kinase B in rat adipocytes. Cell. Signal. 14:231–38 167. Kane LP, Andres PG, Howland KC, Abbas AK, Weiss A. 2001. Akt provides the CD28 costimulatory signal for up-regulation of IL-2 and IFN-γ but not TH2 cytokines. Nat. Immunol. 2:37–44 168. Cantrell D. 2002. Protein kinase B (Akt) regulation and function in T lymphocytes. Semin. Immunol. 14:19–26 169. Appleman LJ, van Puijenbroek AA, Shu KM, Nadler LM, Boussiotis VA. 2002. CD28 costimulation mediates down-regulation of p27kip1 and cell cycle progression by activation of the PI3K/PKB signaling pathway in primary human T cells. J. Immunol. 168:2729–36 170. Metz DP, Farber DL, Taylor T, Bottomly K. 1998. Differential role of CTLA-4 in regulation of resting memory versus naive CD4 T cell activation. J. Immunol. 161:5855– 61 171. Riley JL, Mao M, Kobayashi S, Biery M, Burchard J, et al. 2002. Modulation of TCRinduced transcriptional proﬁles by ligation of CD28, ICOS, and CTLA-4 receptors. Proc. Natl. Acad. Sci. USA 99:11790–95 www.annualreviews.org • CTLA- 4 Function

95

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172. Mandelbrot DA, McAdam AJ, Sharpe AH. 1999. B7-1 or B7-2 is required to produce the lymphoproliferative phenotype in mice lacking cytotoxic T lymphocyte-associated antigen 4 (CTLA-4). J. Exp. Med. 189:435–40 173. Stein PH, Fraser JD, Weiss A. 1994. The cytoplasmic domain of CD28 is both necessary and sufﬁcient for costimulation of interleukin-2 secretion and association with phosphatidylinositol 3 -kinase. Mol. Cell. Biol. 14:3392–402 174. Deane JA, Fruman DA. 2004. Phosphoinositide 3-kinase: diverse roles in immune cell activation. Annu. Rev. Immunol. 22:563–98 175. Hu H, Rudd CE, Schneider H. 2001. Src kinases Fyn and Lck facilitate the accumulation of phosphorylated CTLA-4 and its association with PI-3 kinase in intracellular compartments of T-cells. Biochem. Biophys. Res. Commun. 288:573–78 176. Freeman GJ, Long AJ, Iwai Y, Bourque K, Chernova T, et al. 2000. Engagement of the PD-1 immunoinhibitory receptor by a novel B7 family member leads to negative regulation of lymphocyte activation. J. Exp. Med. 192:1027–34 177. Watanabe N, Gavrieli M, Sedy JR, Yang J, Fallarino F, et al. 2003. BTLA is a lymphocyte inhibitory receptor with similarities to CTLA-4 and PD-1. Nat. Immunol. 4:670–79 178. Zhang X, Schwartz JC, Guo X, Bhatia S, Cao E, et al. 2004. Structural and functional analysis of the costimulatory receptor programmed death-1. Immunity 20:337–47 179. Shlapatska LM, Mikhalap SV, Berdova AG, Zelensky OM, Yun TJ, et al. 2001. CD150 association with either the SH2-containing inositol phosphatase or the SH2-containing protein tyrosine phosphatase is regulated by the adaptor protein SH2D1A. J. Immunol. 166:5480–87 180. Okazaki T, Maeda A, Nishimura H, Kurosaki T, Honjo T. 2001. PD-1 immunoreceptor inhibits B cell receptor-mediated signaling by recruiting src homology 2-domaincontaining tyrosine phosphatase 2 to phosphotyrosine. Proc. Natl. Acad. Sci. USA 98: 13866–71 181. Liu Y. 1997. Is CTLA-4 a negative regulator for T-cell activation? Immunol. Today 18:569– 72 182. Kwon H, Jun HS, Khil LY, Yoon JW. 2004. Role of CTLA-4 in the activation of singleand double-positive thymocytes. J. Immunol. 173:6645–53 183. Pandiyan P, Gartner D, Soezeri O, Radbruch A, Schulze-Osthoff K, Brunner-Weinzierl MC. 2004. CD152 (CTLA-4) determines the unequal resistance of Th1 and Th2 cells against activation-induced cell death by a mechanism requiring PI3 kinase function. J. Exp. Med. 199:831–42 184. Bour-Jordan H, Grogan JL, Tang Q, Auger JA, Locksley RM, Bluestone JA. 2003. CTLA4 regulates the requirement for cytokine-induced signals in T(H)2 lineage commitment. Nat. Immunol. 4:182–88 185. Khattri R, Auger JA, Grifﬁn MD, Sharpe AH, Bluestone JA. 1999. Lymphoproliferative disorder in CTLA-4 knockout mice is characterized by CD28-regulated activation of Th2 responses. J. Immunol. 162:5784–91 186. Oosterwegel MA, Mandelbrot DA, Boyd SD, Lorsbach RB, Jarrett DY, et al. 1999. The role of CTLA-4 in regulating Th2 differentiation. J. Immunol. 163:2634–39 187. Constant S, Pfeiffer C, Woodard A, Pasqualini T, Bottomly K. 1995. Extent of T cell receptor ligation can determine the functional differentiation of naive CD4+ T cells. J. Exp. Med. 182:1591–96 188. Hosken NA, Shibuya K, Heath AW, Murphy KM, O’Garra A. 1995. The effect of antigen dose on CD4+ T helper cell phenotype development in a T cell receptor-αβ-transgenic model. J. Exp. Med. 182:1579–84

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189. Kato T, Nariuchi H. 2000. Polarization of naive CD4+ T cells toward the Th1 subset by CTLA-4 costimulation. J. Immunol. 164:3554–62 190. Perrin PJ, Maldonado JH, Davis TA, June CH, Racke MK. 1996. CTLA-4 blockade enhances clinical disease and cytokine production during experimental allergic encephalomyelitis. J. Immunol. 157:1333–36 191. Hurwitz AA, Sullivan TJ, Krummel MF, Sobel RA, Allison JP. 1997. Speciﬁc blockade of CTLA-4/B7 interactions results in exacerbated clinical and histologic disease in an actively-induced model of experimental allergic encephalomyelitis. J. Neuroimmunol. 73:57–62 192. Karandikar NJ, Vanderlugt CL, Walunas TL, Miller SD, Bluestone JA. 1996. CTLA-4: a negative regulator of autoimmune disease. J. Exp. Med. 184:783–88 193. Chambers CA, Kuhns MS, Allison JP. 1999. Cytotoxic T lymphocyte antigen-4 (CTLA4) regulates primary and secondary peptide-speciﬁc CD4+ T cell responses. Proc. Natl. Acad. Sci. USA 96:8603–8 194. Chambers CA, Sullivan TJ, Truong T, Allison JP. 1998. Secondary but not primary T cell responses are enhanced in CTLA-4-deﬁcient CD8+ T cells. Eur. J. Immunol. 28:3137–43 195. Bachmann MF, Waterhouse P, Speiser DE, McKall-Faienza K, Mak TW, Ohashi PS. 1998. Normal responsiveness of CTLA-4-deﬁcient anti-viral cytotoxic T cells. J. Immunol. 160:95–100 196. Tang Q, Boden EK, Henriksen KJ, Bour-Jordan H, Bi M, Bluestone JA. 2004. Distinct roles of CTLA-4 and TGF-β in CD4+ CD25+ regulatory T cell function. Eur. J. Immunol. 34:2996–3005 197. Birebent B, Lorho R, Lechartier H, de Guibert S, Alizadeh M, et al. 2004. Suppressive properties of human CD4+ CD25+ regulatory T cells are dependent on CTLA-4 expression. Eur. J. Immunol. 34:3485–96 198. Grohmann U, Orabona C, Fallarino F, Vacca C, Calcinaro F, et al. 2002. CTLA-4-Ig regulates tryptophan catabolism in vivo. Nat. Immunol. 3:1097–101 199. Grohmann U, Fallarino F, Puccetti P. 2003. Tolerance, DCs and tryptophan: much ado about IDO. Trends Immunol. 24:242–48 200. Fallarino F, Grohmann U, Hwang KW, Orabona C, Vacca C, et al. 2003. Modulation of tryptophan catabolism by regulatory T cells. Nat. Immunol. 4:1206–12

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Contents

Annual Review of Immunology Volume 24, 2006

Annu. Rev. Immunol. 2006.24:65-97. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:65-97. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li,1 Yisong Y. Wan,1 Shomyseh Sanjabi,1 Anna-Karin L. Robertson,1 and Richard A. Flavell1,2 1

Section of Immunobiology, 2 Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, Connecticut 06520; email: [email protected], [email protected], [email protected], [email protected], richard.ﬂ[email protected]

Annu. Rev. Immunol. 2006. 24:99–146 First published online as a Review in Advance on November 8, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090737 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0099$20.00

Key Words TGF-β, Smad, tolerance, inﬂammation, immunopathology

Abstract Transforming growth factor-β (TGF-β) is a potent regulatory cytokine with diverse effects on hemopoietic cells. The pivotal function of TGF-β in the immune system is to maintain tolerance via the regulation of lymphocyte proliferation, differentiation, and survival. In addition, TGF-β controls the initiation and resolution of inﬂammatory responses through the regulation of chemotaxis, activation, and survival of lymphocytes, natural killer cells, dendritic cells, macrophages, mast cells, and granulocytes. The regulatory activity of TGF-β is modulated by the cell differentiation state and by the presence of inﬂammatory cytokines and costimulatory molecules. Collectively, TGF-β inhibits the development of immunopathology to self or nonharmful antigens without compromising immune responses to pathogens. This review highlights the ﬁndings that have advanced our understanding of TGF-β in the immune system and in disease.

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INTRODUCTION

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Tolerance: a physiological state in which the immune system does not mount destructive responses to antigens Regulatory T cells (Tregs): T cell subsets that possess immune suppression activities that are essential for maintaining self-tolerance and for controlling pathological immune responses.

The immune system has evolved to mount robust responses to pathogens while maintaining tolerance to self or innocuous antigens such as those in commensal bacteria. Multiple mechanisms operate to ensure this normal immunological function. Deletion of highafﬁnity self-reactive T and B cell clones during their development creates a peripheral lymphocyte repertoire that preferentially recognizes nonself-antigens (1, 2). Innate immune recognition of and activation by pathogenassociated molecules further directs immune responses to foreign antigens (3). Nonetheless, no absolute demarcation exists for these selection- or recognition-based mechanisms. There is clear evidence that self-reactive lymphocytes are present in the periphery of normal individuals (4). Self components can also

activate innate immune receptors and sometimes precipitate the development of autoimmune diseases (5, 6). Recently, regulatory T cells (Tregs) and cytokines have been recognized as essential components of peripheral tolerance mechanisms (7, 8). Transforming growth factor-β (TGF-β) is one such regulatory cytokine with a pivotal role in regulating immune responses (9, 10) (Figure 1). TGF-βs are regulatory molecules with pleiotropic effects on cell proliferation, differentiation, migration, and survival that affect multiple biological processes, including development, carcinogenesis, ﬁbrosis, wound healing, and immune responses (11). The TGF-β system originated about one billion years ago, before the divergence of arthropods from vertebrates, and has been progressively adapted to regulate newly emerged

Figure 1 TGF-β regulation of tolerance and immunity. TGF-β signaling is coupled to other environmental changes, allowing it to act as a maintainer of peripheral tolerance and immune homeostasis during steady state in a healthy host. There are times when it is essential to initiate immune responses to provide protection against foreign pathogens. TGF-β is a key regulator of the immune responses, as it controls the threshold of activation and aids in chemotaxis. By regulating the function and survival of activated leukocytes and the healing process, TGF-β also helps to resolve the immune response. An immune response can result in the development of immunological memory, which provides long-term protection. The role of TGF-β in the generation and maintenance of memory cells is an area that requires more investigation. Some of the processes affected by TGF-β during each phase of the immune response are listed. 100

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systems in higher organisms (12). The ﬁrst observation of TGF-β regulation of immune cell functions was made two decades ago (13). In the 1990s, generation and analysis of TGFβ1−/− mice established a central role for this cytokine in inhibiting inﬂammation and autoimmune diseases and has fostered growing interest in this cytokine in the immune system (14, 15). Identiﬁcation of TGF-β receptors and Smad factors as mediators of receptor signaling pathways has also shed light on the molecular mechanisms of TGF-β regulation of immune responses (16, 17). In the 2000s, development of mouse models with cell type– speciﬁc inactivation of TGF-β signaling has started to reveal the regulatory network of the TGF-β pathway in vivo (18–22). In this review, we discuss the multifaceted roles of TGF-β in the immune system with an emphasis on its regulation of peripheral leukocyte functions.

TGF-β EXPRESSION AND ACTIVATION TGF-βs belong to the TGF-β superfamily, with additional members including bone morphogenetic proteins, activins, and growth differentiation factors (23). There are three homologous TGF-β isoforms in mammals, TGF-β1, TGF-β2, and TGF-β3, encoded by different genes (24). TGF-β1 is the predominant isoform expressed in the immune system, but all three isoforms have similar properties in vitro. TGF-β is synthesized as a prepro-TGF-β precursor. The pre region contains a signal peptide, and pro-TGF-β is processed in the Golgi by a furin-like peptidase (25), where the N-terminus of the immature protein, the propeptide, is removed. A homodimer of this new protein, called the latencyassociated protein (LAP), is noncovalently associated with a homodimer of mature TGFβ.This complex is called latent TGF-β or the small latent complex (SLC). The SLC can be secreted as such or in association with latentTGF-β-binding protein (LTBP) as a large

TGF-β AND WOUND HEALING Tissue damage often occurs during pathological immune responses due to inﬂammation, where neutrophils enter the wound site to remove pathogens and damaged tissue through phagocytosis, and macrophages (Ms) appear to continue this process and secrete more growth factors, including TGF-β. Following pathogen clearance, ﬁbroblasts migrate in to deposit collagen that is cross-linked and organized to form an extracellular matrix during the ﬁnal healing phase. For these highly organized processes to occur, numerous cell-signaling events are required, and TGF-β is an important one. As a ﬁbrotic factor, TGF-β facilitates wound healing during resolution of immune responses and many other wound healing processes, e.g., incisional and excisional procedure, punch, ulcers, and bone damage. Discussion of the role of TGF-β in wound healing is beyond the scope of this review. It is nevertheless a critical regulator for immune resolution.

latent complex (LLC). LTBP plays an important role in targeting TGF-β to the extracellular matrix (26). TGF-β cannot bind to its receptors in its latent form but needs to be liberated from the constraints of LAP and LTBP. In vitro, this can be achieved using extremes of pH, heat, or several proteases (26). In vivo, the mechanisms for activation are less clear, but several models have been proposed, including proteolytic activation by transglutaminase, conformational change of LAP through physical interaction with thrombospondin, and mechanical traction via αvβ6 integrins on epithelial cells (27–31). Little is known about the transcriptional regulation of TGF-β1. The TGF-β1 promoter contains activator protein 1 (AP-1) binding sites where c-jun and c-fos induced by TGF-β1 itself bind to and stimulate TGF-β1 production (32–34). A recent study has shown that the AP-1 activators c-Jun-NH2-terminal kinase 1 and 2 (JNK1 and JNK2) inhibit TGF-β1 expression in ﬁbroblasts (35), suggesting that AP-1 may play a dual role in regulating TGF-β1 transcription. Cytokines can also modulate TGF-β1 expression in

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Smad: a family of intracellular transcription factors activated by the TGF-β family of receptors through phosphorylation. The nomenclature is a combination of the orthologs identiﬁed in C. elegans (Sma) and Drosophila (Mad) LAP: latency-associated protein SLC: small latent complex LTBP: latent-TGFβ-binding protein LLC: large latent complex

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ALK: activin receptor like kinase

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SBE: Smad-binding element

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immune cells (see below), although the mechanisms, in most cases, are uncharacterized. In many cases, TGF-β1 message levels do not correlate with the amount of secreted protein, suggesting post-transcriptional and/or post-translational regulation of TGF-β1 expression (36). The most abundant TGF-β1 transcript has a long 5 untranslated region that negatively regulates TGF-β1 translation, the molecular mechanism of which is still not completely understood (37). The regulation of TGF-β2 and TGF-β3 expression is distinct from that of TGF-β1 and is mostly under the control of developmental or hormonal signals (38).

TGF-β RECEPTOR AND SIGNALING The TGF-β superfamily mediates its biological functions via binding to type I and II transmembrane serine/threonine kinase receptors. Although more than 35 TGF family members have been identiﬁed, only ﬁve type I (activin receptor like kinase, ALK family) and seven type II receptors have been reported (23). TGF-β mediates its functions mostly via ALK5 and TGF-β receptor II (TGFβRII) (Figure 2). The active forms of TGF-β initially engage TGFβRII. TGF-β1 and TGFβ3 bind to TGFβRII with high afﬁnity, but strong binding of TGF-β2 to this receptor only occurs in the presence of membranebound betaglycan, also known as TGFβRIII (39). Although ALK5 is not required for the initial binding of TGF-β, it is necessary for the signaling. Active TGF-β dimer binds to the tetrameric ALK5 and TGFβRII receptor complex to initiate cell signaling (40). In endothelial cells, TGF-β can use an alternative type I receptor, ALK1, for signaling, but its involvement in immune cell functions remains unknown (41). TGF-β family signaling is best characterized as a linear pathway where ligand binding brings the type II receptor near and then activates the type I receptor, which subsequently phosphorylates intracellular Smad 102

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proteins whose nuclear localization is required for the transcriptional regulation of target genes (40, 42) (Figure 2). The eight vertebrate Smads identiﬁed so far are grouped into three categories: ﬁve receptor-associated Smads (R-Smad1, 2, 3, 5, and 8), one common Smad (Co-Smad4), and two inhibitory Smads (I-Smad6 and 7). R-Smads are sequestered in the cytoplasm in the absence of signaling. Upon phosphorylation by ALK5, R-Smad2 and 3 associate with Co-Smad4 and translocate into the nucleus to bind to SBE (Smad-binding element) (43–46). Although Co-Smad4 facilitates nuclear translocation of R-Smads, R-Smad can translocate into the nucleus without Co-Smad4 (42, 47). Unlike R-Smads, I-Smad7 is not phosphorylated upon TGF-β activation (48). Instead of being a transmitter for TGF-β signaling, I-Smad7 suppresses TGF-β signaling through at least two mechanisms: (a) competing with R-Smads for the binding to ALK5 and (b) recruiting Smurf-containing E3 ubiquitinase complexes to degrade ALK5 (49, 50) (Figure 2). Activated Smad protein complexes bind to DNA weakly, and their highafﬁnity DNA binding is achieved by associating with a large number of other transcription factors (51–53). To control gene expression, Smad complexes recruit coactivators that contain histone-acetyl transferase (HAT) activity, e.g., CBP/p300; or histone-deacetylase (HDAC) activity–containing corepressors, e.g., Sno/Ski, to activate or repress target genes, respectively (52, 54, 55) (Figure 2). Studies using cells that are deﬁcient in Co-Smad4 or that express dominant-negative R-Smads or mutated ALK5 that is defective in signaling support the existence of Smad-independent TGF-β signaling pathways (56, 57) (Figure 2). Indeed, rapid activation of Ras-Erk, TAK-MKK4-JNK, TAKMKK3/6-p38, Rho-Rac-cdc42 MAPK, and PI3K-Akt pathways were observed when cells were treated with TGF-β, although detailed mechanisms are not yet fully understood (54). Furthermore, cross-talk between MAPKs and Smad proteins appears to modulate TGF-β

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Figure 2 TGF-β transduces signals through Smad-dependent and Smad-independent pathways. Dimeric TGF-β becomes activated following dissociation from LAP and LTBP and binds to and activates tetrameric TGF-β receptor complex consisting of ALK5 and TGFβRII. Activated ALK5 phosphorylates Smad2 and Smad3, which translocate into the nucleus in a complex with Smad4. Smad complex binds to a target promoter in association with other transcription factors (TFs) and regulates gene expression via recruiting HAT or HDAC. One of TGF-β’s targets is Smad7, which downregulates TGF-β signaling by competing with Smad2 and Smad3 for ALK5 binding and by degrading ALK5 through recruiting Smurf-containing E3 ubiquitin ligase complexes. In addition to Smads, PI3K and various MAPKs are also activated by TGF-β. Ras-ERK also cross-talks with Smads. Furthermore, activated TGF-β receptors modulate the function of FKBP12, TRIP-1, PP2A, and eIFs by direct association.

responses (58–60). In addition, TGF-β receptors activate FKBP12, TRIP-1, and PP2A and regulate certain translation initiation complexes, such as eIF2a and eIF3 through direct protein binding (61–63). The signiﬁcance of these pathways in regulating immune cell functions remains to be established.

TGF-β REGULATION OF IMMUNE CELL FUNCTION TGF-β and T Lymphocytes TGF-β exerts the greatest impact on T lymphocytes, as is evident in mice with a T cell– speciﬁc blockade of TGF-β signaling (18).

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Figure 3 TGF-β regulation of T cell responses. TGF-β blocks T cell proliferation by inhibiting IL-2 production via Smad3 and by downregulating the expression of cyclinD2 and E, CDK4, and c-myc. TGF-β inhibits differentiation of Th1 and Th2 cells through suppressing the expression or function of T-bet/Stat4 and GATA-3/NFAT. Mechanisms involved in TGF-β inhibition of CTL differentiation are not well understood, although inhibition of c-myc and T-bet expression is suggested. TGF-β induces FoxP3 expression and the generation of Tregs. TGF-β also prevents T cell activation-induced cell death (AICD) through inhibiting c-myc-mediated FasL expression and other unknown mechanisms.

cki: cycle-dependent kinase inhibitor

TGF-β affects T cell proliferation, differentiation, and survival (64, 65) (Figure 3). The differentiation states of target cells and the presence of additional regulatory signals, including costimulatory molecules and inﬂammatory cytokines, inﬂuence TGF-β regulation of T cell activity. The ﬁnal outcome of TGF-β on T cells is therefore context dependent. TGF-β also induces the FoxP3expressing CD4+ CD25+ Tregs and can therefore indirectly inﬂuence T cell activation (Figure 3). TGF-β regulation of T cell proliferation. A crucial function for TGF-β in T cell biology was ﬁrst demonstrated by its antiproliferation activity on T cells in vitro (13). TGF-β uses multiple pathways to inhibit T cell proliferation, including inhibiting the expression of the T cell mitogenic cytokine interleukin (IL)-2 which is most likely via suppression of IL-2 transcription. Functional analysis of cis-regulatory elements of the murine IL-2 promoter region showed that TGF-β blocks

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IL-2 transcription via inhibition of the activity of an octamer-binding enhancer (66). Smad3−/− murine T cells are resistant to TGF-β-induced inhibition of IL-2 expression (67). Interestingly, one SBE was mapped to the –105 region upstream of the human IL-2 promoter, through which Smads cooperate with the transcriptional corepressor Tob to inhibit IL-2 transcription (68). Inclusion of IL-2 in T cell culture relieves TGF-β inhibition of T cell proliferation in a dose-dependent manner (13). Nevertheless, even with the highest dose of IL-2, TGF-βinduced reduction of T cell proliferation was observed (13). TGF-β inhibits proliferation of various cell types through mechanisms directly targeting cell cycle regulators, including upregulation of cyclin-dependent kinase inhibitors (cki) p15, p21, or p27, and downregulation of c-myc (69–72). A recent study showed that TGF-β inhibits T cell proliferation in all three cki knockouts, demonstrating a dispensable or redundant role for the ckis in the TGF-β pathway in T cells (73).

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TGF-β blocks the expression of c-myc, cyclin D2, and cyclin E in T cell lines (74–76), as well as cdk4 expression in primary CD4+ T cells (73). The signiﬁcance of these molecules in TGF-β inhibition of T cell proliferation warrants further investigation. Smad3 is essential for TGF-β blockade of IL-2 production in both CD4+ and CD8+ T cells (77). Nevertheless, although TGF-β inhibition of T cell receptor (TCR)-mediated CD4+ T cell proliferation is abrogated in Smad3−/− T cells, its inhibition of CD8+ T cell proliferation remains intact (77). These observations revealed a Smad3- and IL-2-independent antiproliferative pathway in CD8+ T cells, the molecular mechanism of which is currently unknown. TGF-β inhibition of T cell proliferation is inﬂuenced by the differentiation status of T cells and the integrated signals from cytokines and costimulatory molecules on T cells. Although TGF-β inhibits naive T cell proliferation, it has minimal effect on activated T cells, which correlates with reduced TGFβRII expression (78). Interestingly, IL-10 treatment enhances TGFβRII expression and restores TGF-β responsiveness on activated T cells (78). The presence of CD28 costimulation attenuates TGF-β inhibition of TCR-mediated naive CD4+ T cell proliferation (79). Although the expression of IL-2 is strongly inhibited by TGF-β even with CD28 stimulation, IL-2 is required for the proliferation of TCR/CD28/TGF-βstimulated naive T cells (79). It appears that CD28 costimulation synergizes with low IL-2 to drive T cell proliferation, which overrides the inhibitory effects of TGF-β. It is conceivable that TGF-β inhibition of naive T cell activation in the absence of costimulation blunts T cell responses to self-antigens under steady state, whereas reversal of TGF-β inhibition by strong costimulatory signals, which are often associated with infection, limits the suppressive activity of TGF-β during a normal immune response. Thymocytes isolated from TGF-β1deﬁcient mice hyperproliferated upon stim-

ulation (80), which might contribute to the development of their autoimmune phenotype. In transgenic mice, the expression of a dominant-negative mutant of TGFβRII under the CD2 promoter speciﬁcally inhibits TGF-β signaling in T cells (19). These mice develop a progressive CD8+ T cell lymphoproliferative disorder that later transforms into lymphoma (81). Recently, an important function for Smad3 in inhibiting leukemogenesis was revealed: A compound mutation of p27 cki and Smad3 in mice results in the development of T cell lymphoma (82). Interestingly, Smad3 protein is also absent or reduced in several cases of human T cell acute lymphoblastic leukemia (82). Thus, TGF-β inhibition of T cell proliferation appears to be an important mechanism to maintain T cell homeostasis and to prevent lymphoproliferative disorder.

TCR: T cell receptor

TGF-β regulation of helper T (Th) cell differentiation. Activation of naive CD4+ T cells under polarization conditions leads to their differentiation into helper T cells (83). T helper 1 (Th1) cells express interferon-γ (IFN-γ) and lymphotoxin-α (LT-α), which mobilize the cellular arm of the immune system to combat intracellular pathogens. Th2 cells secrete cytokines, including IL-4, IL-13, and IL-5, which are essential for optimal antibody production and elimination of extracellular pathogens. Dysregulated CD4+ Th cells can induce immunopathology. Excessive Th1 responses are associated with various autoimmune and inﬂammatory disorders, whereas enhanced Th2 cytokine production is involved in atopic diseases, including allergic asthma. TGF-β is a potent regulator of effector T cell differentiation, and it generally inhibits the acquisition of Th cell functions (64). TGF-β-mediated inhibition of Th cell differentiation occurs even in the presence of IL-2, in which case T cell proliferation is largely unaffected (84). These observations demonstrate that TGF-β can target both proliferation and differentiation programs in T cells independently.

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Effector T cell differentiation is orchestrated by signals from TCR, costimulatory molecules, and cytokine receptors (83). These integrated signals induce the expression of lineage-commitment transcription factors that drive Th cell differentiation. Transcription factors T-bet and Stat4 specify Th1 cell fate, whereas GATA-3 and Stat6 induce Th2 cell differentiation (83). Several reports show that TGF-β prevents the development of Th2 cells by inhibiting the expression of GATA-3 (85, 86). However, these studies do not reveal whether TGF-β directly inhibits GATA-3 expression. One recent study showed that TGF-β interferes with TCR and costimulatory receptor signaling pathways by inhibiting the activation of the Tec kinase Itk and the calcium inﬂux elicited by TCR/CD28 stimulation in CD4+ T cells (87). Impaired calcium signaling compromises nuclear translocation of nuclear factor of activated T cells (NFAT) transcription factors and the expression of GATA-3 (87). TGF-β blockade of Th1 cell differentiation is associated with reduced IL-12 receptor β2 (IL-12Rβ2) and T-bet expression (88, 89). T-bet is required for the induction of IL12Rβ2 (90). Therefore, reduced IL-12Rβ2 levels upon TGF-β treatment is likely due to its inhibition of T-bet expression, which is dependent on the IFN-γ/Stat1 pathway (90). T-bet also induces IFN-γ expression (91) and provides a positive feedback loop to stabilize Th1 cell fate. Whether TGF-β primarily targets IFN-γ or T-bet to inhibit Th1 cell differentiation is unclear. TGF-β also inhibits the expression of Stat4 (92), a transcription factor activated by IL-12. It appears that Stat4 downregulation blocks IFN-γ expression at priming, whereas reduced T-bet expression inhibits IFN-γ production at recall responses (92). Thus, TGF-β can target multiple signaling pathways to regulate Th1 cell differentiation. Similar to T cell proliferation, TGF-β regulation of Th cell differentiation is inﬂuenced by cell differentiation state and other signals. Fully differentiated Th2 cells are re-

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fractory to TGF-β inhibition of cytokine production, whereas IFN-γ expression in differentiated Th1 cells is inhibited by TGF-β (93). Nevertheless, early IFN-γ production by Th1 cells attenuated the inhibitory effects of TGF-β, as IFN-γ neutralization leads to a complete blockade of Th1 cell differentiation (84). IFN-γ is known to induce Smad7 expression and consequent inhibition of TGF-β signaling (94). Whether the same mechanism operates in Th1 cells remains to be tested. Whereas TGF-β inhibits the production of most effector cytokines in CD4+ T cells, ectopic expression of TGF-β1 in T cells results in enhanced production of the antiinﬂammatory cytokine IL-10 (95). This effect is more profound under Th1 conditions and is associated with TGF-β activation of the IL-10 promoter via Smad4 (95). These ﬁndings are consistent with the observation that these two suppressive cytokines are coexpressed at sites of inﬂammation (96). TGF-β also induces the expression of IL-9 in CD4+ T cells (97), an important cytokine involved in the Th2 responses that induce mast cells. The in vivo signiﬁcance of this pathway remains unknown. The prominent inhibitory activity of TGF-β on Th cell differentiation in vitro is supported by the in vivo phenotypes of TGF-β1−/− mice and mice with T cell– speciﬁc blockade of TGF-β signaling. TGFβ1-deﬁcient mice in a mixed genetic background develop a multifocal inﬂammatory disease associated with increased inﬂammatory cytokine production (14, 15). Similar CD4+ T cell activation and differentiation was observed when TGF-β1−/− mice were backcrossed onto the BALB/c background (98). Signiﬁcantly, depletion of CD4+ T cells or crossing TGF-β1−/− mice onto an MHC class II–null background inhibits inﬂammation (99), demonstrating an essential role for CD4+ T cells in promoting disease development. Whether T cells themselves are direct targets of TGF-β1 is not revealed from these studies because TGF-β1 can modulate the activity of multiple cell types. Expression

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of a dominant-negative form of TGFβRII from the CD4 promoter (CD4-DNR) blocks TGF-β signaling in T cells (18). These mice develop an autoimmune inﬂammatory phenotype associated with CD4+ T cell differentiation into Th1 or Th2 cells (18). Another study demonstrated that transgenic expression of Smad7 in T cells attenuates TGF-β signaling in T cells and renders mice hyperresponsive to antigen-induced airway inﬂammation associated with enhanced Th2 cell differentiation (20). These observations reveal that TGF-β is an essential regulator of Th1 and Th2 cell responses in vivo. TGF-β regulation of cytotoxic T lymphocyte differentiation. CD8+ T cells represent an important arm of adaptive immunity to intracellular pathogens and tumors. Dysregulated CD8+ T cells mediate pathogenesis in various autoimmune diseases. CD8+ T cells exert their effector functions through production of inﬂammatory cytokines such as IFN-γ and LT-α, and acquisition of cytolytic activities mediated by perforin/granzyme and death receptor pathways. Compared with CD4+ T cells, naive CD8+ T cells acquire effector T cell functions more readily upon antigenic stimulation (100) and therefore require stringent regulation. In addition to proliferation, TGF-β potently inhibits CD8+ T cell differentiation. Early studies showed that CD8+ T cells activated in the presence of TGF-β do not acquire cytotoxic T lymphocyte (CTL) functions (101), which is likely due to TGF-β inhibition of perforin expression in activated CD8+ T cells (102). TGF-β also inhibits Fas ligand (FasL) expression in T cell lines (75), thereby affecting the death receptor cytotoxic pathway of CD8+ T cells. As in Th1 cells, the presence of TGF-β greatly attenuates IFN-γ expression in CD8+ T cells (103, 104). Thus, TGF-β inhibits the expression of multiple effector molecules of CTLs. Although not well understood, several molecular mechanisms involved in TGF-β-mediated inhibition of CTL effector functions have been

suggested. IFN-γ expression and CTL differentiation is regulated by two T-box transcription factors, eomesodermin and T-bet (105). Similar to Th1 cells, TGF-β inhibition of IFN-γ expression in CD8+ T cells is associated with reduced T-bet expression (104). TGF-β inhibition of FasL expression appears to be a consequence of blockade of cmyc expression (75). In addition, TGF-β inhibition of perforin expression was observed upon activation of resting T cells (102), implying perforin as a direct TGF-β target. TGFβ regulation of CD8+ T cell differentiation is context dependent. 4-1BB is a costimulatory molecule induced in activated CD4+ and CD8+ T cells (106). 4-1BB costimulation reverses TGF-β inhibition of CTL differentiation (107), which is further enhanced by IL-12 but reduced by IL-4 (107). Therefore, TGF-β inhibition of CD8+ T cell differentiation is modulated by costimulatory receptor and cytokine signaling pathways. TGF-β regulation of CD8+ T cell differentiation has been further illuminated by in vivo studies with TGF-β1−/− mice and CD4-DNR mice. Elimination of CD8+ T cells alleviated the inﬂammatory disease developed in TGF-β1−/− mice (108). The CD4 promoter used in CD4-DNR mice lacks the CD8 silencer (18); therefore, TGF-β signaling in both CD4+ and CD8+ T cells is blocked in these mice. CD8+ T cells in this model spontaneously acquire an activated phenotype and produce effector cytokines (18). Remarkably, CD4-DNR mice also mount a strong antitumor response (109), associated with the expansion and enhanced activities of tumorspeciﬁc CTLs (109). Thus, TGF-β essentially regulates CD8+ T cell differentiation in vivo.

DNR: dominant-negative TGF-β receptor II CTL: cytotoxic T lymphocytes

TGF-β regulation of Tregs. The existence of T cell subsets that actively suppress immune responses and maintain immunological tolerance was documented in the 1970s (110). These suppressor T cells are now termed regulatory T cells (Tregs). The bestcharacterized Tregs are the naturally occurring Tregs that are CD4+ CD25+ and

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Activation-induced cell death (AICD): T cells proliferate in response to antigen stimulation and undergo apoptosis to eliminate excess numbers of activated T cells

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comprise 5%–10% of peripheral CD4+ T cells (7). FoxP3, a forkhead family transcription factor, is a lineage speciﬁcation factor for Tregs (111). Tregs are generated during thymic development as well as in the periphery during homeostatic proliferation (7, 112, 113). In addition to Tregs, mature T cells are able to acquire immunoregulatory functions upon activation, such as CD4+ Th3 cells generated in oral tolerance models (114). The role of TGF-β in the development, maintenance, and induction of Tregs has attracted much attention recently. Investigators have shown that TGF-β1 can convert CD4+ CD25− T cells to Tregs in vitro (115– 119). Overexpression of TGF-β1 in the islets of the pancreas also expands the Treg population that protects nonobese diabetic (NOD) mice from type 1 diabetes (120). In both cases, the induction of regulatory activities correlates with increased expression of FoxP3. In fact, only FoxP3-expressing T cells possess suppressive activities (119). The function of TGF-β in Treg generation/maintenance under physiological conditions remains controversial and poorly understood. Blockade of TGF-β signaling in Tregs inhibits their proliferation in mice treated with dextran sulfate, a drug that induces colitis. These Tregs also fail to protect mice from dextran sulfate–induced colitis (121). In another report, peripheral but not thymic Treg numbers were found reduced in 8- to 10-day old TGF-β1−/− mice (122), which suggests a function for endogenous TGF-β1 in the maintenance of the peripheral Treg population. However, an earlier study showed no defect in Treg development in TGF-β1−/− mice (123). In another recent report, thymic Tregs from CD4-DNR mice retain their ability to inhibit colitis, and TGF-β1−/− TCR transgenic T cells possess a normal number of FoxP3-expressing Tregs (124). The reasons for these discrepancies are unknown but may relate to the different experimental systems and mouse genetic backgrounds used in these studies. In addition, CD25 was used as a marker for Tregs. Because CD25 is also exLi et al.

pressed by activated T cells, Tregs identiﬁed by CD25 staining might be contaminated by effector T cells. With the development of GFP- and RFP-FoxP3-knockin mice (119, 125), these potential complications can be circumvented. TGF-β regulation of T cell survival. Acquired immune response to foreign antigens is a multistep process that involves clonal expansion of antigen-speciﬁc T cells followed by contraction; most of these are eliminated by apoptosis, but some activated T cells develop into memory T cells. There are two predominant apoptotic pathways in T cells: the intrinsic pathway that is regulated by the proand antiapoptotic Bcl-2 family members; and the death receptor pathway, including CD95 (APO-1/Fas), that induces apoptosis via direct caspase activation (126, 127). TGF-β promotes the survival of T cells during T cell expansion and differentiation. TGF-β inhibition of T cell activation (see above) is inﬂuenced by the T cell differentiation status and integrated signals from cytokines and costimulatory molecules. Therefore, in the absence of CD28 costimulation, TGF-β inhibits TCR-stimulated proliferation of naive T cells. However, in the presence of CD28, TGF-β inhibits T cell apoptosis and promotes T cell expansion (79, 128). TGF-β1 also promotes the survival of T cells with memory/effector phenotypes (78, 79, 93, 129, 130). Additionally, IL-2 and TGF-β synergize to block activation-induced cell death (AICD) in both Th1 and Th2 CD4+ T cells (129, 131–133). The mechanism of TGF-β-mediated T cell survival is not well understood; however, several studies have shown that TGF-β inhibits Fas-induced T cell apoptosis (134, 135). The antiapoptotic effect of TGF-β was associated with reduced c-myc expression that resulted in reduced levels of FasL (75). Together, these studies demonstrate that TGF-β can promote T cell survival and effector T cell expansion. The ﬁrst in vivo demonstration of TGFβ’s role in T cell survival came from the

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analysis of TGF-β1−/− mice (136). Spontaneous apoptosis was found in both thymic and peripheral T cells in the absence of TGF-β1 (136). When TGF-β1−/− mice were treated with anti-CD3 antibody, massive thymic and peripheral T cell apoptosis were observed (136). Addition of TGF-β to T cell culture does not correct the increased cell death in TGF-β1−/− cells, and the apoptotic phenotype was not found in Smad3−/− mice. On the basis of these observations, investigators suggested an antiapoptotic role for intracellular TGF-β1 (136). In another study, splenic T cells from TGF-β1−/− mice show elevated IFN-γ in response to TCR stimulation and undergo enhanced apoptosis (137). IFNγ precipitates T cell AICD by inducing caspase 8 expression (138). Therefore, increased T cell apoptosis in TGF-β1−/− mice may also be a consequence of enhanced IFN-γ production. It remains to be determined whether TGF-β regulation of T cell survival represents an independent pathway or is secondary to its regulation of T cell activation.

T cell production of TGF-β. Among the three family members of TGF-β, TGF-β1 is the predominant isoform expressed in leukocytes (9). Early studies showed that activated T cells produce TGF-β1 (13). Later studies of oral tolerance models revealed a subset of CD4+ T cells, named Th3 cells, that express TGF-β1 as an effector cytokine (114). TGF-β1 produced by Th3 cells has been proposed to inhibit Th1 and Th2 cell differentiation and provide help for B cells in IgA production (139). Despite considerable study, the mechanisms by which the recently identiﬁed CD4+ CD25+ Tregs suppress immune responses remain poorly understood. Some Tregs express surface TGF-β1, which investigators have suggested mediates their suppressor function (140–142). However, TGF-β1−/− Tregs still inhibit proliferation of responder T cells in vitro, indicating that TGF-β1 synthesis by Tregs is not required for suppression under these culture conditions

(143). The protective activity of Tregs against inﬂammatory bowel disease (IBD) induced by transfer of CD4+ CD45RBhigh T cells into SCID mice is reversed by the treatment of the recipients with anti-TGF-β antibody (144). However, in an autoimmune gastritis model, Treg-mediated protection is not affected by the same treatment (143). Even in the same colitis model, TGF-β1−/− Tregs were shown to be protective or inactive depending on experimental systems (123, 124, 145). Therefore, the function of Treg-produced TGF-β1 remains controversial. To determine a deﬁnitive function of T cell–produced TGF-β1 in vivo, we have generated a strain of TGF-β1 conditional knockout mice. Mice that lack TGF-β1 selectively in T cells develop an autoimmune inﬂammatory disease characterized by multiorgan leukocytic inﬁltration and circulating autoreactive antibodies, which is associated with spontaneous T cell activation and effector T cell differentiation. TGF-β1 produced by Tregs, as well as by naive/activated T cells, is required to protect mice from IBD (M.O. Li and R.A. Flavell, unpublished). These observations reveal a speciﬁc function for T cell–produced TGF-β1 in suppressing T cell activation and inﬂammatory disease. Cell signaling pathways involved in TGFβ1 production in T cells remain largely uncharacterized. Engagement of the inhibitory receptor CTLA-4 was described as inducing TGF-β1 production (146–148), although this was disputed in another report (149). CD69 is an early leukocyte activation molecule. Cross-linking of CD69 induces TGF-β1 production in both CD4+ and CD8+ T cells (150). Interestingly, CD69−/− mice develop exaggerated collagen-induced arthritis (151), which is normally inhibited by TGF-β1. Th2 polarization conditions enhance and Th1 conditions inhibit TGF-β1 production in vitro (152). Consistent with this, CD4+ T cells from T-bet-deﬁcient mice produce more TGF-β1 than wild-type T cells (153). In addition, T cells produce TGF-β1 when

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Figure 4 TGF-β regulation of B cell responses. TGF-β inhibits B cell proliferation through induction of Id3, p27, and p21 and inhibition of c-myc, calcyclin, and ATM. TGF-β blocks IL-4-induced Stat6 activation via SOCS1 and SOCS3 and B cell receptor–induced Src activation through CD72 and SHIP-1. TGF-β promotes IgA class switching through Smad3/4 and CBFα3-mediated transactivation of germline α-promoter. TGF-β also induces apoptosis in immature and resting B cells through induction of Id3 and Bim, and inhibition of NF-κB and c-myc.

Class switching: a process in which the heavy chain of an antibody is switched from the IgM isotype to IgG, IgA, or IgE isotypes

they undergo apoptosis (154), which might downregulate immune responses to apoptotic cell-associated antigens.

TGF-β and B Lymphocytes TGF-β is an important regulator of B lymphocyte activity, exempliﬁed by the phenotype of mice with a B cell–speciﬁc blockade of TGF-β signaling (21). TGF-β inhibits B cell proliferation, induces apoptosis of immature or resting B cells, and blocks B cell activation and class switching to most isotypes except for IgA (Figure 4). Thus, similar to T cells, TGF-β has both inhibitory and stimulatory effects on B cells (155, 156). TGF-β regulation of B cell proliferation. TGF-β inhibits proliferation of B lymphocyte progenitors, which is mediated partly by the induction of TGF-β target gene Id3, a helixloop-helix transcription factor (157). Overexpression of Smad7 reverses TGF-β-induced Id3 expression, suggesting a requirement of the Smad pathway in this regulation (157).

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TGF-β also potently inhibits proliferation of mature B cells in vitro, which is associated with cell cycle blockade at G1/S (158–160). TGF-β represses cyclin A expression and inactivates cdk2, possibly via the upregulation of the cki p27 (161, 162). Recently, global gene changes regulated by TGF-β in primary B cells have been characterized. TGF-β induces the expression of the cki p21 and suppresses the expression of c-myc, calcyclin, and ATM (163), which might also collectively induce growth arrest. Studies with Smad3−/− B cells reveal that TGF-β inhibition of B cell proliferation is Smad3 independent (16, 17). The signaling pathway that leads to the inhibition of mature B cell proliferation remains to be identiﬁed. Mature splenic B cells from conditional B cell TGFβRIIknockout mice show increased BrdU labeling (21), revealing an important function for TGF-β in inhibiting B cell proliferation in vivo. As in T cells, TGF-β regulation of B cell proliferation is modulated by other signaling pathways. CD40, a member of the tumor

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necrosis factor (TNF) receptor superfamily, plays an important role in promoting B cell survival and immunoglobulin (Ig) class switching. Engagement of CD40 induces Smad7 expression and protects B cells from TGF-β-induced growth inhibition (164).

attenuating B cell responses to low-afﬁnity/ -avidity antigens. Interestingly, anti-DNA antibody titers also increase in these mice (21), suggesting that TGF-β is an important regulator of B cell tolerance to self-antigens in vivo.

TGF-β regulation of B cell differentiation. TGF-β is an important regulator of B cell activation and differentiation. Early studies showed that exogenously administrated TGF-β inhibits the expression of both light chains and the secreted form of the heavy chain (165, 166). TGF-β also blocks class switching to most IgG isotypes (158). In contrast, TGF-β promotes switching to IgA and IgG2b in mouse B cells and IgA in human B cells in vitro (167–171). However, although TGF-β signaling in B cells is essential for IgA switching in mice (see below), it is dispensable for IgG2b production (21). These in vivo studies reveal a general inhibitory function for TGF-β on antibody production with the exception of IgA. IL-4 produced by Th2 cells is an important mitogenic and survival factor for B cells and promotes isotype switching to IgG1 and IgE via the Jak/Stat pathway. Signiﬁcantly, TGFβ attenuates IL-4-induced Stat6 phosphorylation in B cells (163), which correlates with enhanced expression of SOCS1 and SOCS3 (163), negative regulators of the Jak/Stat pathway. TGF-β also induces the expression of CD72 and inositol-pyrophosphate phosphatase SHIP-1 (163). The CD72/SHIP-1 complex plays an important role in antagonizing the activity of Src kinases activated upon B cell receptor stimulation. Therefore, TGFβ targets both the cytokine and B cell receptor pathways to regulate B cell activation and differentiation. Consistent with TGF-β inhibition of the production of most antibody isotypes, conditional inactivation of TGF-β signaling in B cells results in elevated serum Ig in mice (21). These mice also develop enhanced IgG3 responses to a normally weak antigen (21), revealing an important function for TGF-β in

TGF-β regulation of IgA production. Secretory IgA is important for the prevention of microbial infection in mucosa (172). TGF-β has a unique role in promoting the differentiation of IgA-secreting plasma cells. Upon optimal antigen and cytokine stimulation, the antiproliferative activity of TGF-β subsides, and TGF-β strongly promotes IgA production (173). TGF-β-induced IgA class switching is associated with increased transcription of germline α-transcripts (174). The identiﬁcation of a TGF-β-responsive tandem repeat sequence in the mouse germline α-promoter led to the characterization of transcription factors involved in this regulation (175). Two Smad-binding sites and two core binding factor elements are necessary and, when dimerized, sufﬁcient for the TGF-β response (175). In vitro studies showed that Smad3/4 and CBFα3 [also known as acute myeloid leukemia (AML)-2 or Runx3] bind to these sites, respectively (176– 178). The Smad3/4 and CBFα3 complexes may further recruit the transcriptional coactivator p300 to augment transcription of the germline α-promoter (179, 180). However, Smad3−/− mice have normal levels of serum IgA (16, 17). The mechanism of Smad3independent regulation of IgA production in vivo is unknown. Mice with a blockade of TGF-β signaling in B cells are almost devoid of serum IgA (21). The TGF-β pathway is also required for the IgA response to mucosally administered antigens (orally or intranasally) (181). TGF-β1−/− mice have IgA only at reduced levels (182). Maternally derived TGF-β1 could be responsible for this partial phenotype (183). Alternatively, TGF-β2 or TGF-β3 may also be involved in promoting IgA switching.

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TGF-β regulation of B cell survival. TGF-β induces apoptosis in immature B cells and resting B cells (184, 185). TGF-βmediated apoptosis in the mouse immature B cell line, WEHI 231, is associated with decreased expression of c-myc (186). TGF-β also induces NF-κB inhibitor IκBα and inhibits NF-κB activation (186). Ectopic expression of the c-Rel subunit of NF-κB or cmyc overrides apoptosis induced by TGF-β1 (186), revealing the functional importance of these two targets. In WEHI 231 cells, TGF-β also induces expression of the BH3-only protein Bim (187), which is a proapoptotic member of the Bcl-2 family. It remains to be established whether downregulation of NF-κB and c-myc causes induction of Bim or whether they represent separate apoptotic pathways. TGF-β induces apoptosis in B lymphocyte progenitors, which is mediated by TGF-β target gene Id3 (157). Interestingly, both c-myc and Id3 regulate B cell proliferation. Therefore, the antiproliferation pathway and the apoptotic pathway may overlap signiﬁcantly. Mice with a B cell blockade of TGF-β signaling have increased peritoneal B-1 cells (21), a unique B cell subset responsible for producing natural Igs. However, B-1 cells show reduced BrdU labeling (21), suggesting that increased B-1 cell pools are mediated by enhanced survival of these cells. The B cell receptor repertoire of B-1 cells recognizes shared microbial structures and/or selfantigens (188, 189). TGF-β signaling may therefore be involved in B cell negative selection to eliminate high-afﬁnity self-reactive B-1 cell clones.

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B cell production of TGF-β. Resting human B cells express TGF-β1 mRNA but secrete low amounts of TGF-β1 protein (158). Upon stimulation with Staphylococcus aureus Cowan, B cells produce high amounts of TGF-β1 protein with little alteration of TGF-β1 mRNA (158). Polyclonal stimulation of B cells with LPS or anti-IgM antibody also induces TGF-β1 production. Neutralization of TGF-β1 from LPS-stimulated 112

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B cells results in a signiﬁcant decrease of IgG2a, IgG3, IgG1, and IgE secretion, but not IgM secretion (190). Restoration of TGFβ1 revealed that low doses of TGF-β1 promote, whereas high doses inhibit, antibody secretion (190). These observations suggest that autocrine TGF-β1 may stimulate B cells to secrete antibody. The signiﬁcance of this pathway remains unknown in vivo. Stimulation of human B cells with anti-CD40 antibody induced TGF-β production and IgA class switching, which was ablated by anti-TGF-β antibody (191), suggesting that B cell–produced autocrine TGF-β may promote IgA class switching. Studies of autoimmune MRL/lpr mice have revealed that a fraction of B cell– produced TGF-β1 associates with IgG autoantibodies (192). The antibody-bound TGF-β1 is consistently more active than the uncomplexed TGF-β1 in vivo and is responsible for the increased susceptibility of these mice to bacterial infection (192). In light of the potent suppressive function of TGF-β1 in autoimmunity (see below), increased TGFβ1 levels in MRL/lpr mice might represent a negative feedback mechanism to attenuate disease development.

TGF-β and Natural Killer Cells Natural killer (NK) cells are lymphoid cells that participate in innate immunity and in early defense against a variety of infections and cancers. NK cells express cell surface activating and inhibitory receptors that allow them to respond to microbial products, cytokines, stress signals, and inducible molecules expressed in cancer cells (193, 194). Thus, without requiring prior activation or immunization, NK cells recognize and kill infected or tumorigenic cells and rapidly produce chemokines and cytokines. IFN-γ produced by NK cells is considered essential for stimulation of Th1 responses (22, 195). TGFβ is a potent inhibitor of NK cell functions via attenuating its cytolytic activity and IFNγ production (196–199). The importance of

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TGF-β signaling in NK cells is supported by a recent study of transgenic mice in which TGF-β signaling is blocked in both NK cells and dendritic cells (DCs) (22). Interestingly, a selective inhibitory function of TGF-β in NK cell homeostasis was also observed (22). TGF-β therefore controls the homeostasis of NK cells and suppresses their cytokine production and cytolytic activity.

and the downregulation of NKp30 results in inhibition of DC killing by TGF-β1treated NK cells (206). Downmodulation of NKG2D, which is associated with elevated levels of TGF-β1, was also observed in cancer patients (207). Thus, by regulating the expression of activating receptors on NK cells, TGF-β can inﬂuence their cytolytic functions.

TGF-β regulation of cytokine production by NK cells. TGF-β is a potent antagonist of IL-12-induced IFN-γ expression in NK cells (199, 200), and its expression coincides with the downregulation of NK cell responses during viral infection (201). TGF-β also inhibits IFN-γ production by human NK cells stimulated with IL-2 (202, 203). A recent study further demonstrated an essential role for TGF-β in inhibiting NK IFN-γ expression in vivo (22), also showing that NK cells rather than DCs are the major innate sources of IFN-γ. Stimulation of TGF-β-resistant NK cells results in higher IFN-γ production, which promotes Th1 differentiation (22). The mechanisms whereby TGF-β antagonizes NK cell production of IFN-γ are unknown. TGF-β downregulates the expression of IFN-α receptor and the α chain of the IL-2 receptor on human NK cells (198, 202), which might be involved in TGF-β inhibition of cytokine signaling pathways in NK cells.

NK cell production of TGF-β. NK cells have been shown to constitutively produce active and latent TGF-β1 (208). In addition, anti-CD2 antibody stimulation strongly enhances CD3− CD56+ NK cell production of active TGF-β1 (208). Stimulation of NK cells with anti-CD69 antibody induces TGF-β1 production (150). Signiﬁcantly, CD69−/− mice mount enhanced NK responses to tumors (150). Thus, NK cells may represent an important in vivo source of TGF-β1 to inhibit immune responses.

DCs are professional antigen-presenting cells (APCs) with an essential role in immunity and tolerance (209). TGF-β1 is required to support the development of Langerhans cells (LCs), resident DCs present within epithelial cells in the epidermis. TGF-β also regulates the maturation of differentiated DCs and DCmediated T cell responses (210).

TGF-β regulation of cytolytic function of NK cells. The ﬁrst demonstration of TGFβ inhibition of NK cell cytolytic function, in which TGF-β suppressed cytolytic activity of IFN-α-stimulated NK cells, was made two decades ago (198). Targeted killing by NK cells depends on the engagement of activating receptors and coreceptors on NK cells, among which the NK cell–speciﬁc NKp46, NKp30, and NKp44, collectively termed natural cytotoxicity receptors (204), and NKG2D (205) appear to play a major role in NK cell– mediated cytotoxicity. TGF-β1 inhibits expression of NKp30 and NKG2D receptors,

TGF-β regulation of Langerhans cell development. In vitro, TGF-β1 is required for the differentiation of LCs from hematopoietic precursor cells in serum-free medium supplemented with GM-CSF, TNF-α, and stem cell factor (211). In the absence of TGF-β1, monocytes are generated (211). The addition of TGF-β1 to GM-CSF- and IL-4supplemented cultures of human monocytes induces the expression of LC markers such as E-cadherin and CD1a (212). These studies and others (213, 214) establish that TGF-β1 promotes LC generation from monocytes. TGF-β1 stimulation of LC development

TGF-β and Dendritic Cells

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from hematopoietic progenitor cells is also associated with protection of DC progenitor cells from apoptosis (215). TGF-β1 therefore regulates both cell differentiation and cell survival during LC development. The importance of TGF-β1 in LC development is underscored by the observation that TGF-β1−/− mice lack LCs (216). Transfer of TGF-β1-deﬁcient bone marrow cells into lethally irradiated recipients results in the differentiation of donor LCs (217), demonstrating that paracrine sources of TGF-β1 are sufﬁcient to support LC development. Conversely, reconstitution of SCID TGF-β1deﬁcient mice with TGF-β1 heterozygous bone marrow also leads to the generation of LCs (218). These observations suggest that TGF-β1 functions via both paracrine and autocrine mechanisms to induce LC differentiation. Id2, a helix-loop-helix transcription factor, is strongly upregulated during differentiation of DCs, including LCs (219). TGF-β1 induces Id2 expression in DCs (219). Signiﬁcantly, Id2−/− mice lack LCs (219), suggesting that Id2 is an essential TGF-β1 target in promoting LC differentiation.

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TGF-β regulation of DC maturation and function. DCs differentiated from human hematopoietic progenitor cells or monocytes in the presence of TGF-β1 express predominantly intracellular MHC class II, low levels of CD1d, and costimulatory molecules including CD80, CD83, and CD86 (220– 222), features characteristic of in vivo LCs. These observations are consistent with murine studies in which TGF-β1 inhibits maturation of DCs differentiated from bone marrow cells with GM-CSF (223), demonstrating that TGF-β1 promotes the generation of DCs with an immature phenotype. In astrocytes, TGF-β1 inhibits the transcription of class II transactivator (CIITA) and consequently MHC class II expression, which depends on the presence of Smad3 (224). Whether TGF-β uses the same pathway to regulate MHC class II expression in DCs remains to be tested. 114

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TGF-β also regulates the antigenpresentation function of differentiated DCs in vitro. Stimulation of DCs with bacterial components, inﬂammatory cytokines, or costimulatory receptors upregulates MHC class II and costimulatory molecules and cytokines such as IL-12, which induce T cell activation and differentiation. Inclusion of TGF-β in LPS-stimulated DC culture inhibits the expression of MHC class II and costimulatory molecules, which attenuates the antigenpresentation function of DCs (221). In addition, DC maturation and IL-12 production induced by inﬂammatory cytokines IL-1 and TNF-α are inhibited by TGF-β (221). In contrast, CD40 ligand or anti-CD40 antibody-induced DC maturation or cytokine production is not affected by TGF-β (221, 225). CD40 ligand represents a key T helper signal for DC maturation. Lack of TGF-β inhibition of CD40 ligand–induced DC maturation implies that TGF-β1 primarily suppresses noncognate DC activation. DC production of TGF-β. Immunostaining of epidermal sections revealed that TGFβ1 is strongly associated with LCs (226). DCs differentiated from human CD34+ cord blood cells also express high levels of TGFβ1 mRNA (227). In mouse, TGF-β1 is highly expressed in bone marrow–derived CD11c+ CD86− immature DCs (228). The function of DC-produced TGF-β1 remains unknown. Under steady state, presentation of antigens by immature DCs results in T cell unresponsiveness (229). It is conceivable that DC TGF-β1 may act on T cells in a paracrine manner to induce T cell tolerance. DC TGFβ1 may also be involved in the maintenance of the immature state of DCs via an autocrine pathway. To this end, it is interesting to note that TGF-β1 is expressed at low levels in bone marrow–derived CD11c+ CD86+ mature DCs (228).

TGF-β and Macrophages Macrophages (Ms) are professional phagocytes with a critical role in clearing

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apoptotic cells (230). They also phagocytose microbes and produce inﬂammatory mediators upon microbial challenge. In addition, Ms present antigens to T cells and regulate adaptive immunity. TGF-β regulation of the monocyte/M lineage appears to depend on the differentiation stage of the cells. Generally, TGF-β stimulates cells at the resting state (monocytes), whereas activated cells (Ms) are inhibited (231). TGF-β regulation of monocyte recruitment and production of inflammatory mediators. TGF-β recruits monocytes to the site of injury or inﬂammation via multiple mechanisms: It acts as a chemoattractant for monocytes (232–234); it induces adhesion molecules, including LFA-1 and the ﬁbronectin receptor on monocytes, enabling their attachment to extracellular matrix (235, 236); and it induces matrix metalloproteinases (MMPs), which can dissolve vascular membranes and facilitate monocyte transmigration (235). In addition, TGF-β potentiates inﬂammation through induction of IL-1, IL-6, and leukotriene C4 synthase in monocytes (232, 237, 238). These observations reveal a proinﬂammatory function for TGF-β on monocytes. TGF-β regulation of macrophage phagocytosis. Once monocytes differentiate into Ms, TGF-β functions mostly as an inhibitory molecule. Ms express surface receptors involved in the phagocytosis of bacteria as well as of aged or apoptotic cells. CD36 and SR-A are two such scavenger receptors, the expression of which is downregulated by TGF-β (239, 240). The ability of Ms to phagocytose microbes is greatly enhanced by IgG opsonization. Expression of two of the IgG receptors, FcγRI and FcγRIII, and of the common γ-subunit is downregulated by TGF-β in murine myeloid cells and in human THP-1 cells, which results in reduced phagocytosis of IgG-coated particles (241).

TGF-β regulation of macrophage activation. Stimulation of Ms with IFN-γ and/or microbial products such as LPS induces their activation. In vitro, TGF-β inhibits the expression of several LPS-induced inﬂammatory mediators such as TNF-α and MMP12 as well as chemokines including MIP-1α and MIP-2 (242, 243). Reactive oxygen and nitrogen species produced by activated Ms are important mediators against invading microbes. TGF-β downregulates the production of nitric oxide (NO) and superoxide ion and inhibits the expression of inducible NO synthase (iNOS) in activated Ms (244, 245). These inhibitory activities of TGF-β on Ms may resolve inﬂammation and prevent the development of immunopathology. Toll-like receptors (TLRs) are innate immune receptors involved in the recognition of microbes (246). LPS is recognized by TLR4 with the assistance of CD14. TGF-β inhibits CD14 expression in LPSstimulated Ms and may therefore attenuate TLR4 signaling (247). MyD88 is a key adapter molecule downstream of TLRs. TGF-β inhibits MyD88-dependent TLR signaling pathways in Ms by promoting its degradation (248). Overexpression of Smad3 blocks LPS-induced iNOS and MMP-12 promoter activity in Ms, whereas a dominantnegative mutant of Smad3 alleviates the TGF-β inhibition, suggesting that Smad3 is a critical mediator of TGF-β inhibition of M activation (243). TGF-β regulation of macrophage antigen presentation. IFN-γ-induced expression of MHC class II molecules in Ms is inhibited by TGF-β via the attenuation of CIITA (249). In addition, TGF-β inhibits expression of the costimulatory molecule CD40 and the inﬂammatory cytokine IL-12p40, which collectively results in the inhibition of the antigenpresentation function of Ms (250, 251). This inhibition may play an important role in resolving an ongoing immune response by diminishing secondary stimulation of T cells at the site of infection.

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PMN: polymorphonuclear neutrophils

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Macrophage production of TGF-β. It has been reported that phagocytosis of apoptotic cells leads to TGF-β1 secretion, which inhibits the production of inﬂammatory cytokines and chemokines in Ms including IL1β, TNF-α, GM-CSF, and IL-8 (252, 253). Exposure of phosphatidylserine on the outer leaﬂet of the plasma membrane represents a key signal for the recognition of apoptotic cells by Ms (254). Liposomes containing phosphatidylserine induce TGF-β1 in Ms in vitro (255), and instillation of apoptotic cells or phosphatidylserine-containing liposomes enhances TGF-β1 production associated with reduced inﬂammation in mice, illustrating the in vivo relevance for this pathway (256, 257). Infection of Ms with certain microbes triggers the production of TGF-β1, which modulates the host responses (see below). TGF-β1 production in Ms is additionally regulated by other cytokines: IL-9, a Th2 cytokine, promotes TGF-β1 production in LPS-stimulated human peripheral blood monocytes (258); and the p40 subunit of IL-12 inhibits TGF-β1 production in Ms, as p40−/− Ms secrete a large amount of TGF-β1 (259).

TGF-β and Mast Cells Mast cells are recognized mostly for their role in allergy and asthma, but they are also important in host defense against pathogens and in autoimmune diseases. They act mainly through the release of effector molecules such as histamine, proteases, and TNF-α. TGF-β has a stimulatory effect on the initial phase of mast cell–dependent immune responses and may also regulate their functions at later phases. TGF-β regulation of mast cell recruitment and activation. TGF-β is a potent chemoattractant for mast cells (260, 261). In murine bone marrow–derived mast cells, TGF-β induces integrin α7 expression, which facilitates their intraepithelial

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migration through binding to laminin-1 (262). Mast cells are activated mainly through the cross-binding of IgE to FcεR. TGF-β attenuates the expression of FcεR receptor in cultured human and mouse mast cells by blocking protein translation (263). Data regarding the effects of TGF-β on mast cell proliferation, histamine release, and the secretion of TNFα and other inﬂammatory mediators are contradictory, possibly due to heterogeneity of mast cells derived from different species and anatomical sites (264–268). Mast cell production of TGF-β. Early studies showed that mast cells or cell lines produce TGF-β1, which induces collagen synthesis in ﬁbroblasts (269, 270). The Th2 cytokine IL-9 also induces TGF-β1 production from peritoneal mast cells or D36 mast cells (264, 268). Mast cells produce chymase-1 that activates TGF-β1. Both molecules are stored in secretory granules, which implies that mast cell stimulation and degranulation may result in both the secretion and the activation of TGF-β1 (271).

TGF-β and Granulocytes Polymorphonuclear neutrophils (PMN) are professional phagocytes essential for ﬁrstline defense against microorganisms (272). Together with proteases liberated from the granules, reactive oxygen species derived from superoxide (which is generated after PMN activation) is used to kill ingested microbes. Eosinophils are granulocytes known for their role in immune responses to infection with helminthic parasitic worms (273). They secrete inﬂammatory mediators, including lipid mediators, cationic proteins, cytokines, chemokines, and components of the oxidative burst, all of which can also cause tissue damage in inﬂammatory diseases (274). Many of the eosinophil-speciﬁc mediators act directly on mast cells and aggravate allergic inﬂammation.

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TGF-β regulation of granulocyte recruitment. TGF-β is a potent chemoattractant for PMN, acting at femtomolar levels (275, 276), and may play an important role in recruiting cells to sites of infection or injury. However, TNF-α-induced endothelial cell expression of IL-8, another PMN chemoattractant, is inhibited by TGF-β (277). TGF-β also downregulates the expression of Eselection and VCAM-1 in endothelial cells (278, 279) and inhibits PMN transmigration in TNF-α-activated endothelial monolayers (279). It appears that TGF-β acts initially to stimulate an immune response by recruiting PMN, but when inﬂammation is ongoing with secretion of proinﬂammatory mediators, TGF-β limits the inﬂux of leukocytes through other mechanisms. An alternative interpretation is that the chemotactic property of TGF-β is important for PMN movement in tissues, whereas it inhibits their migration across the endothelium (279). Smad3−/− PMN are impaired in their chemotactic responses toward TGF-β (16), which is associated with chronic mucosal infections in these mice. Therefore, TGF-β-mediated PMN chemotaxis may represent an important host defense mechanism. TGF-β has also been demonstrated to act as a chemoattractant for human eosinophils (280). The in vivo signiﬁcance of this pathway remains to be determined. TGF-β regulation of granulocyte function. TGF-β regulation of PMN activation and effector functions such as phagocytosis, degranulation, and respiratory burst is controversial and requires further investigation (276, 281, 282). IL-13 plays a crucial role in activation and survival of eosinophils, and surface expression of the IL-13Rα1 is enhanced by TGF-β in vitro (283). These observations suggest a stimulatory function for TGF-β on eosinophil-mediated inﬂammation. In contrast, TGF-β reduces the viability and the release of peroxidase in eosinophils isolated from peripheral human blood (284). These ﬁndings imply a complex

role for TGF-β in the regulation of eosinophil functions. Granulocyte production of TGF-β. PMN-derived TGF-β1 may play a role in synovial effusions (285). TGF-β1 produced by eosinophils may be involved in ﬁbrosis and the remodeling of asthmatic airways by inducing collagen and other extracellular matrix proteins by smooth muscle cells (SMCs) and ﬁbroblasts (274). IL-5 is an important cytokine for eosinophil differentiation, activation, and recruitment. IL-5 induces TGF-β1 production in eosinophils in vitro (286). Eosinophils from IL-5 transgenic mice concordantly secrete large amounts of TGFβ1 (286). Consistent with these ﬁndings, deﬁciency of IL-5 or IL-5R or treatment with anti-IL-5 antibody leads to a parallel reduction of eosinophils, TGF-β1, and ﬁbrotic lesions in models of airway remodeling (287, 288). These observations suggest that IL-5-induced TGF-β1 in eosinophils may play a critical role in promoting ﬁbrosis in asthma.

Chemotaxis: a phenomenon in which mobile cells direct their movement to the concentration gradient of chemicals. Chemotaxis is crucial for the recruitment of inﬂammatory cells to sites of infection or injury

TGF-β AND DISEASE TGF-β Regulation of Autoimmune Diseases Autoimmune diseases, including more than 70 chronic disorders affecting 3%–5% of the population, are characterized by loss of immunological tolerance to self-antigens. Studies of animal models and human patients have revealed a critical function for TGF-β in regulating leukocyte functions in autoimmune diseases (289). Systemic lupus erythematosus. Systemic lupus erythematosus (SLE) is a B and T cell autoimmune disorder characterized by B cell hyperactivity and activation of autoreactive T cells. Autoantibodies and immune complexes, together with pathogenic T cells, induce pathology in multiple organs, including skin, blood vessels, lung, joints, and kidney.

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Interestingly, TGF-β1−/− mice develop immunopathology resembling that of SLE, including formation of autoantibodies and IgG deposits in their renal glomeruli (14, 15, 290, 291). Consistent with this ﬁnding, decreased production of TGF-β1 was found in some SLE patients (292). Nonetheless, decreased production of active TGF-β in SLE patients does not correlate with disease severity (293). SLE is a multifactorial disease with genetic and environmental factors. MRL/lpr mice develop systemic autoimmune disease mimicking human lupus. It is paradoxical that these mice have elevated levels of circulating TGF-β1 (294). Increased TGFβ1 suppresses T cell and PMN functions, which may account for the susceptibility to infection of these mice (192, 294). Interestingly, overexpression of TGF-β1 in MRL/lpr mice inhibits the development of SLE (295). These observations suggest that TGF-β1 elaboration during evolution of autoimmunity in these mice may represent a negative feedback mechanism of the host to restore self-tolerance.

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Rheumatoid arthritis. Rheumatoid arthritis (RA) is a T cell–dependent, antibodymediated inﬂammatory disease that targets the joints. Collagen-induced arthritis (CIA) is an experimental model sharing several clinical features with RA (296). Regulation of RA by TGF-β1 was shown to be site and context dependent, owing to its pleiotropic activities. Systemic administration of TGFβ1 to mice inhibits CIA (297), whereas its local administration to joints induces synovitis and aggravates disease (234). Similarly, blocking endogenous TGF-β1 by the systemic injection of anti-TGF-β antibody exacerbates CIA in mice (297), whereas the local blockade of TGF-β1 ameliorates ongoing inﬂammation (298). TGF-β signaling in T cells regulates CIA because inhibition of TGF-β signaling in T cells renders mice more susceptible to the disease; this is associated with increased production of IFN-γ 118

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and TNF-α (299). Inhibition of T cell activation and differentiation by endogenous TGFβ is therefore required for the maintenance of joint integrity. CD69 is an early activation molecule expressed in different leukocyte subsets. CD69 cross-linking induces TGFβ1 production in T cells as well as in NK cells and Ms (151). Signiﬁcantly, CD69−/− mice develop severe CIA associated with decreased TGF-β1 production (151). These studies suggest that the CD69 pathway is required to inhibit CIA by regulating TGF-β1 production. Insulin-dependent (type 1) diabetes mellitus. Type 1 diabetes (T1DM) is an autoimmune disease with a progressive T cell– mediated destruction of insulin-producing β cells in the pancreatic islets. NOD mice develop spontaneous T1DM resembling human disease. Several laboratories have developed transgenic models to study TGF-β1 regulation of T1DM in NOD mice. Constitutive expression of TGF-β1 in β cells under the rat insulin promoter inhibits T1DM, which investigators propose is caused by immune deviation from Th1 to Th2 responses (300) and increased T cell apoptosis (301). Expression of TGF-β1 in α cells with the transgene under the control of rat glucagon promoter also blocks disease development (302). In addition, these mice are resistant to passive transfer of disease with diabetogenic T cells (302), which demonstrates that paracrine TGF-β1 is sufﬁcient to protect β cells from killing by CD4+ and/or CD8+ effector T cells. Overproduction of TGF-β1 in the islets induces ﬁbrosis and may complicate the analysis of disease development (301). Expression of TGF-β1 in a temporally controlled inducible system during either preinsulitis, or priming, or the destruction phase of the disease protects mice from T1DM (120). Signiﬁcantly, overproduction of TGF-β1 expands FoxP3expressing Tregs in the islets, which inhibits T1DM (120). These observations suggest that TGF-β may use multiple mechanisms to inhibit disease development.

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Endogenously produced TGF-β1 also regulates autoimmune T1DM. Treatment with nonmitogenic anti-CD3 antibody induces long-term remission of T1DM in NOD mice (303). Interestingly, the protective effect is abrogated by neutralizing anti-TGF-β antibody (303). T cells are likely a key source of TGF-β, as anti-CD3 antibody treatment stimulates CD4+ T cells to produce TGFβ1 (303). A humanized anti-CD3 antibody is currently in clinical trial for autoimmune T1DM (304). It will be interesting to investigate whether the same TGF-β1-dependent mechanism is also involved in inhibiting human disease. Multiple sclerosis. Multiple sclerosis (MS) is an autoimmune disorder affecting the central nervous system. Experimental autoimmune encephalomyelitis (EAE) serves as an experimental model for MS. Early studies showed that administration of exogenous TGF-β1 to mice or treatment of myelin basic protein (MBP)-speciﬁc T cells with TGF-β is able to prevent or inhibit EAE (305–307). Increased expression of TGF-β1 mRNA or protein is associated with remission of the disease (308, 309). Administration of neutralizing antibody to TGF-β also enhances the clinical severity of the disease (310). Together, these studies suggest that endogenous TGF-β1 is involved in protection from EAE. The sources and targets of TGF-β1 in the regulation of EAE remain to be deﬁned. TGF-β1 is associated with the induction of oral tolerance to EAE antigens. Oral administration of high doses of MBP results in clonal deletion or anergy, whereas treatment with low doses of MBP induces suppressive T cells (114). These T cells, named Th3 cells, secrete high amounts of TGF-β1 and protect mice from EAE (114). Anti-TGFβ1 antibody treatment abrogates the protection (114), suggesting that TGF-β1 functions as an effector cytokine of Th3 cells. Nasal administration of MBP also induces TGFβ1-producing Tregs in rats (311), which prevents the relapse of EAE. These observations

demonstrate a critical function for TGF-β1 in the induction of tolerance to antigens administrated via the mucosal route.

TGF-β Regulation of Tumor Immunity Cancers result from accumulated genetic and epigenetic changes in tumor cells, which confer growth advantages. At least three stages are involved in cancer development: (a) induction, the accumulation of mutations induced by carcinogens; (b) promotion, a process triggered by mechanical or chemical injury that is not mutagenic by itself; and (c) progression, the invasive growth that results in aggressive metastasis (312, 313). Most tumor cells are clonal in origin, and during cancer progression new subpopulations arise with alterations in their morphology, hormone dependence, migration property, and proteome composition (314, 315). Consequently, most tumors acquire new antigens. Their invasive activity also fosters an inﬂammatory environment, which potentiates antitumor immune responses involving both the innate arm (NK cells, PMN, and Ms) and the adaptive arm (CD8+ and CD4+ T cells) of the immune system (316, 317). During their evolution, tumors that produce increased TGF-β or promote TGFβ production by surrounding cells not only advance tumor progression but also allow tumors to evade immune surveillance (318– 320). A recent study showed that RNAi inhibition of TGF-β expression in glioma cells restores NKG2D expression in NK and CD8+ T cells and that the expression of the NKG2D ligand MICA in tumor cells is associated with decreased tumor growth in mice (321). On the other hand, overexpression of TGFβ1 in a highly immunogenic murine tumor suppresses the antitumor immune response (322). These observations demonstrate that TGF-β produced by tumor cells is involved in their immune evasion. Studies with CD4-DNR mice reveal that TGF-β is required for suppressing T cell–mediated tumor

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rejection (109). These mice were resistant to challenge with two highly aggressive tumors, EL-4 and B16-F10 (109). TGF-β signaling in CD8+ but not in CD4+ T cells was essential for tumor immune evasion (109). Tregs have been shown to promote tumorigenesis (323), and abrogation of Tregs resulted in enhanced antitumor immunity (324–326). Interestingly, the inhibitory function of Tregs requires TGF-β signaling in CD8+ T cells (327). In addition to the inhibition of effector T cell functions, TGF-β may promote tumor growth through modulating Tregs. Most tumor cells produce large amounts of TGF-β that may induce Treg differentiation. In support of this notion, a recent study demonstrated that FoxP3-expressing Tregs increase in the peripheral blood of tumorbearing patients (328) and decrease following therapy (329). It will therefore be interesting to study whether TGF-β signaling is involved in the generation of tumor-promoted Tregs.

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TGF-β Regulation of Atherosclerosis Atherosclerosis is an inﬂammatory disease that is a response to inﬁltration, retention, and oxidation of low-density lipoprotein (LDL) in the arterial intima beneath the endothelial cell layer (330). Lipid-laden M-derived foam cells as well as collagen-producing SMCs comprise the bulk of the lesion. Inﬂammatory cells, including activated T cells and APCs such as Ms and mature DCs, affect both the size and vulnerability of the lesion. Vulnerable plaques with little collagen, high lipid content, and a high density of inﬂammatory cells are prone to rupture and may have detrimental consequences such as formation of thrombi, which may lead to myocardial infarction or stroke (330). IFN-γ, TNF-α, IL-18, and IL-12 secreted by inﬂammatory cells within the lesion promote the formation of vulnerable plaques (331, 332). Neutralization of TGF-β increased lesion vulnerability in mouse mod120

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els, supporting an antiatherosclerotic function of TGF-β (333, 334). Apolipoprotein E−/− mice bred with CD4-DNR mice exhibited a dramatic increase of large vulnerable lesions, despite decreased cholesterol levels in serum (335). IFN-γ mRNA increased more than 100-fold in the aortas, concomitant with less collagen, more T cells, and increased expression of MHC class II in the lesions (335). Reconstitution of LDL receptor−/− mice with bone marrow from CD2-DNR mice similarly led to the formation of vulnerable lesions but with minimal effects on the lesion size (336). Thus, T cells are central targets of the antiatherosclerotic property of TGFβ. Proatherogenic T cells likely drive atherogenesis by affecting endothelial cells, Ms, and SMCs. Inhibition of T cells by TGFβ dampens these atherogenic mechanisms (335). In addition, TGF-β may directly target endothelial cells, DCs, Ms, and SMCs to regulate atherogenesis. In endothelial cells, TGF-β inhibits expression of adhesion molecules and chemokines that partake in recruitment of leukocytes (278, 279). The expression of the scavenger receptors CD36 and SR-A in Ms are downregulated by TGF-β, and cholesterol efﬂux is increased through TGF-β regulation of ATP-binding cassette transporter-1 (ABC1) (239, 240, 337). Collectively, this may lead to reduced foam cell formation. Because TGF-β reduces antigen presentation by Ms and DCs, disease-promoting adaptive immune responses against proatherogenic antigens are likely inhibited by TGF-β. In addition, the formation of stable lesions may be promoted by TGF-β induction of collagen and TIMP (tissue inhibitor of matrix metalloproteinase) synthesis in SMCs (338, 339). Therefore, TGF-β may target multiple cell types to prevent the development of vulnerable atherosclerotic plaques. Interestingly, higher levels of TGF-β1 are present in human asymptomatic lesions than in lesions from symptomatic patients (340). Lower serum levels of active TGF-β1 have also been found in patients with advanced atherosclerosis

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(341), suggesting that TGF-β1 may regulate atherogenesis in human patients.

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TGF-β Regulation of Asthma Asthma is a complex inﬂammatory disease dependent on Th2-type immune responses and is characterized by airway hyperresponsiveness (AHS). In a murine model of asthma, overexpression of TGF-β1 in ovalbumin (OVA)-speciﬁc Th cells abolishes AHS and airway inﬂammation induced by OVAspeciﬁc Th2 cells (342). A recent study of TGF-β1 heterozygous mice, which produce lower levels of TGF-β1 than wild-type mice, showed exacerbated asthmatic immunopathology (343). These studies demonstrate that endogenous TGF-β1 is an important suppressor of asthma in mice. T cells appear to be a key target of TGF-β1 in this model. Transgenic expression of Smad7 under the control of a distal Lck promoter blocks TGF-β signaling in mature T cells (20). Signiﬁcantly, both OVA-induced AHS and airway inﬂammation is enhanced in the transgenic mice associated with high production of Th2 cytokines (20). Whether TGF-β regulates human asthma remains to be established. Levels of TGF-β1 in the airways of asthmatics were found higher than in normal controls (344). It is conceivable that elevated TGF-β1 in these patients may represent a negative feedback mechanism to control airway inﬂammation. Because TGFβ1 is important in healing, increased TGF-β1 production may be involved in the repair of asthmatic airways. On the other hand, TGFβ1 may also induce ﬁbrosis to exaggerate disease development (345).

TGF-β Regulation of Inflammatory Bowel Disease The inﬂammatory bowel diseases (IBD), comprising Crohn’s disease and ulcerative colitis, are chronic inﬂammatory diseases of the gastrointestinal tract (346). Development of IBD is often a consequence of a loss of tolerance to-

ward commensal bacterial ﬂora (346). Blockade of TGF-β signaling in T cells (CD4DNR) results in T cell activation and induction of IBD in mice (18), demonstrating an essential role for TGF-β signaling in T cells in inhibiting colitis. In a murine model of colitis, protection from IBD induced by transfer of CD4+ CD45RBhigh T cells into SCID mice by CD4+ CD45RBlow or CD4+ CD25+ Tregs is abrogated by administration of neutralizing anti-TGF-β antibody (144). Consistent with this, colitis induced by transfer of CD4+ CD45RBhigh T cells isolated from CD4-DNR mice escapes control by Tregs (124). The cellular sources of TGF-β1 in regulating colitis remain controversial. In a recent study, CD4+ CD25+ cells isolated from TGFβ1−/− mice failed to prevent colitis (145), suggesting an essential function for Treg TGF-β1. In another study, Tregs isolated from TCR transgenic TGF-β1−/− mice were capable of suppressing colitis (124). However, suppression was still abrogated by antiTGF-β antibody (124), demonstrating that TGF-β produced by cells other than Tregs is also involved in inhibiting colitis. In light of the mouse studies, it seems paradoxical that TGF-β1 levels are elevated in the gut of patients with IBD (347). Interestingly, T cells from these patients are refractory to the inhibitory activity of TGF-β1, which is associated with increased expression of Smad7 (348), a negative regulator of TGF-β signaling. These observations suggest that defective TGF-β signaling may be involved in human IBD.

TGF-β Regulation of Infectious Diseases Copious evidence suggests that TGF-β controls many interactions between pathogens and their hosts. TGF-β has the potential to inﬂuence all phases of pathogenesis and plays an important role in the regulation of immune responses to pathogens, inﬂammation, and wound healing. TGF-β is also a common

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molecule exploited by pathogens to facilitate entry, replication, and persistence in the host. In general, most M pathogens have evolved mechanisms to induce TGF-β production, and TGF-β production in turn suppresses the killing activity of Ms, enhances intracellular proliferation of the pathogen, and thus favors virulence. However, in some organisms, TGF-β plays a protective role in host resistance against infection. Therefore, the role of TGF-β in the resolution or exacerbation of infection depends on the nature of the immune response necessary to control parasite replication and disease.

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Parasites Trypanosome. Trypanosoma cruzi is one parasite that makes direct use of the host’s TGF-β signaling pathway. It uses TGF-β receptors I and II for successful entry into mammalian cells (349). Epithelial cells lacking TGF-β receptors are resistant to T. cruzi infection, and infectivity is restored following transfection of functional TGF-β receptors (349). T. cruzi can also induce expression of a TGF-β-responsive reporter gene, suggesting that the parasite might directly activate the TGF-β signaling pathway to facilitate entry into mammalian cells (349). Spleen cells from T. cruzi–infected mice have higher levels of bioactive TGF-β (350). TGF-β treatment of mouse and human Ms blocks IFNγ-mediated inhibition of parasite growth, and TGF-β-treated mice develop higher parasite loads and die faster than control mice (350). An alpha-fetoprotein homolog of T. cruzi can induce TGF-β from mouse splenocytes (351). The African trypanosome T. brucei, which does not invade host cells, might nevertheless possess a functional homolog of this TGF-β-activating moiety because this parasite has also been shown to release a factor that induces TGF-β mRNA expression (352). Leishmania. Leishmania can survive and replicate in the phagolysosome of Ms. Mice 122

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resistant to Leishmania major develop a Th1type response with high levels of IL-2 and IFN-γ, whereas susceptible strains of mice develop a Th2-like response with accompanying increased production of IL-4 and IL-10. NK-produced IFN-γ is responsible for activating Ms to express high levels of iNOS and toxic NO, a molecule with the potent ability to kill intracellular parasites (353). Treatment with anti-IFN-γ abrogates the resistance to L. major in the C3H/HeN mouse strain, whereas administration of IFN-γ offers protection in the susceptible BALB/c strain (354, 355), suggesting that IFN-γ is the most important cytokine for resistance to L. major. Virulent strains of Leishmania have developed mechanisms to induce Ms to produce high levels of active TGF-β, whereas nonpathogenic strains that produce low-grade infection induce relatively low levels of active TGF-β (356). TGF-β made by infected Ms suppresses NO production (244) and can inﬂuence T cell differentiation by inhibiting the production of TNF-α and IFN-γ. Leishmania species can also activate TGF-β using parasite-derived cathepsins to increase local concentrations of active TGF-β to modulate local iNOS and arginase levels (357). Systemic administration of TGF-β increases the infectivity of relatively avirulent strains of Leishmania, and in vivo neutralization of TGF-β is sufﬁcient to inhibit the development of fatal Leishmaniasis in susceptible BALB/c mice (356). When BALB/c mice expressing CD4DNR were challenged with L. major, they arrested infection as effectively as genetically resistant C57BL/6 mice, suggesting susceptibility to L. major infection requires TGF-β signaling in T cells (89). In another study, blockade of TGF-β signaling in NK cells caused the generation of large numbers of NK cells capable of producing considerable IFN-γ, which led to Th1 polarization and protection from L. major infection in BALB/c mice (22). Collectively, these in vivo studies show that TGF-β-mediated inhibition of Th1 differentiation is involved in Leishmaniasis in susceptible mice.

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Toxoplasma gondii. Toxoplasma is another obligate intracellular parasite that infects Ms. As in the case of Leishmania, infection of mouse Ms with T. gondi results in the release of TGF-β, which was associated with the downregulation of TNF-α and its receptors (358, 359). Elevated TGF-β levels were found in spleen cells from SCID mice following exposure to killed T. gondi or after oral exposure to live T. gondi (200). In the same model, TGFβ could inhibit the protective effects of IL-12, and anti-TGF-β antibody partially protected mice from lethal infection (200). From these studies, there appear to be close parallels in how TGF-β regulates infection by a number of protozoan pathogens.

Malaria. In murine malaria infections, TNF-α, IFN-γ, and NO are associated with parasite clearance. TNF-α and IFN-γ act synergistically to activate Ms to phagocytose malaria-infected red blood cells and to release NO. Overproduction of IFN-γ, TNF-α, or LT-α predisposes the host to severe pathology. IL-10 and TGF-β play a key role in limiting malaria pathology (360). When infected mice were treated with neutralizing antibody to TGF-β, the virulence of lethal Plasmodium berghei strains was exacerbated, and a normally resolving chabaudi infection became lethal (361). However, when mice infected with P. berghei were treated with recombinant TGF-β, IL-10 was upregulated, TNF-α was downregulated, parasite replication was slowed, and survival time was extended (361). Mortality is increased in IL-10−/− mice when they are infected with the normally nonlethal P. chabaudi (362), and mortality is further exacerbated by neutralization of TGF-β (363). Moreover, metalloproteinases and a thrombospondin-like molecule from malaria activate TGF-β (364). These studies suggest that TGF-β is associated with protection rather than disease in murine models of malaria.

Bacteria Mycobacteria. Mycobacteria are obligate intracellular pathogens of Ms that cause tuberculosis, leprosy, and opportunistic infections due to immunosuppression. Similar to most protozoan infections, mycobacteria induce M production of active TGF-β and suppress their antibacterial activity to aid their pathogenesis. Both puriﬁed protein derivative and lipoarabinomannan, a cell wall component of tuberculosis, induce TGF-β from human peripheral blood mononuclear cell (PBMC)-derived Ms (365, 366). Listeria. Listeria monocytogenes is a facultative intracellular bacterium and a strong inducer of Th1 response (367). Cytokines such as IFN-γ, TNF-α, and IL-6 play an important role in host resistance to Listeria, and TGF-β plays a protective role in Listeria infection (368). Mice injected with antiTGF-β antibody became susceptible to listeriosis, whereas the administration of human platelet-derived TGF-β1 enhanced their resistance, even though endogenous levels of IFN-γ, TNF-α, and IL-6 were reduced in these mice (368). The mechanism by which TGF-β confers resistance to lethal doses of L. monocytogenes in mice is not yet clear and requires more investigation.

Yeast Candida albicans is an opportunistic fungal pathogen that can cause morbidity and mortality in immunocompromised hosts. An effective Th1 response and production of IFN-γ is required for clearance of C. albicans in mouse models of infection. As in Listeria infection, studies demonstrate that TGF-β plays a beneﬁcial role in acquired resistance against C. albicans infection in mice. Administration of recombinant TGF-β to mice susceptible to C. albicans had a protective effect and delayed the progression of disease (369). In contrast, local TGF-β suppresses

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host defense in immunocompromised hosts (370). Taken together, these data suggest that TGF-β may play a protective or suppressive role depending on the host’s immune status.

Virus

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There are reports of increased TGF-β during many viral infections, such as inﬂuenza, HIV, hepatitis B virus (HBV), hepatitis C virus (HCV), and others. The neuraminidase glycoprotein of inﬂuenza A and B viruses have been shown to directly activate latent TGF-β (371). In the case of HCV, investigators observed enhanced TGF-β production and lack of TGF-β responses (372) and showed that HCV core protein directly upregulates TGFβ1 transcript (373). HCV proteins can also perturb TGF-β signaling via their interac-

tions with Smad molecules (374). Similarly, HIV Tat can induce TGF-β1 in antigenstimulated PBMCs, monocytes, T cells, and astrocytes, and this induction may be in part responsible for the immunosuppressive effects of HIV-1 Tat (375). TGF-β1 also enhances M responsiveness to SDF-1α stimulation and susceptibility to HIV-1 by selectively increasing expression of CXCR4 and enhancing the entry of HIV-1 into human monocyte-derived Ms (376).

SUMMARY AND FUTURE PERSPECTIVES The ﬁrst study of TGF-β on immune cell function was carried out two decades ago (13). TGF-β is now known for its activities on virtually all leukocyte lineages (Figure 5).

Figure 5 Pleiotropic effects of TGF-β on leukocytes. All leukocytes produce and respond to TGF-β. The yin-yang symbol illustrates the fact that TGF-β exerts both stimulatory and inhibitory effects on immune cells. Selected immunological processes regulated by TGF-β are depicted (MC, mast cell; EO, eosinophil; MO/M, monocyte/macrophage). 124

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The dominant role of TGF-β in the immune system is to induce tolerance, as well as to contain and resolve inﬂammation (Figure 1). TGF-β also positively regulates immune responses by recruiting leukocytes at the initial phase, promoting T cell survival, and inducing IgA class switching in B cells. The activity of TGF-β is inﬂuenced by the environmental milieu. Notably, inﬂammatory cytokines such as IFN-γ and costimulatory receptors including CD28, 4-1BB, and CD40 counteract the suppressive activity of TGF-β. This ensures robust immune responses to pathogens while maintaining tolerance to self or innocuous antigens. Disturbance of this balance results in various immune disorders, and TGF-β appears to be a key regulator of these diseases. What questions remain to be addressed? First, our understanding of the cellular mechanisms of immune regulation by TGF-β remains incomplete. Cell type-speciﬁc inactivation of TGF-β signaling has revealed the activities of TGF-β in T, B, and NK cells.

Similar studies can be extended to other leukocyte lineages. All leukocytes produce at least one form of TGF-β, the function and regulation of which is mostly unknown. Generation of TGF-β conditional knockout mouse models will help address this issue. Second, the molecular mechanisms of TGFβ regulation of immune cell functions need further investigation. A recent study in epithelial cells has revealed versatile interactions between signaling molecules of the TGF-β pathway and those of the other signaling pathways (377). To understand how immune cells read and respond to the combinatorial inputs from TGF-β and environmental cues remains a big challenge. Third, translation of our knowledge on TGF-β to cure immune disorders is largely unexplored. Owing to its pleiotropic activities, future attempts to harness the TGF-β system for therapeutic purposes need to focus on speciﬁc targeting strategies. We are conﬁdent that these exciting areas will enthrall investigators for at least another two decades.

ACKNOWLEDGMENTS Because of space restrictions, we were able to cite only a fraction of the relevant literature. We apologize to any colleagues whose contributions may not be appropriately acknowledged in this review or those we could cite only in secondary references. We are grateful to Elizabeth E. Eynon, Sean T. Kim, and Martin Kriegel for reading the manuscript and for incisive and ¨ helpful comments and to Goran K. Hansson for helpful suggestions. M.O. Li is an American Cancer Society fellow, Y.Y. Wan and S. Sanjabi are Cancer Research Institute fellows, and A.K.L. Robertson is a Wenner-Gren Fellow. R.A. Flavell is an investigator of the Howard Hughes Medical Institute. Work from this laboratory has been supported by grants from ADA (7-03-RA-23) and NIDDK (DK53015, DK51665).

LITERATURE CITED 1. Kappler JW, Roehm N, Marrack P. 1987. T cell tolerance by clonal elimination in the thymus. Cell 49:273–80 2. Goodnow CC, Cyster JG, Hartley SB, Bell SE, Cooke MP, et al. 1995. Self-tolerance checkpoints in B lymphocyte development. Adv. Immunol. 59:279–368 3. Medzhitov R, Janeway CA Jr. 1997. Innate immunity: the virtues of a nonclonal system of recognition. Cell 91:295–98 4. Guerder S, Picarella DE, Linsley PS, Flavell RA. 1994. Costimulator B7-1 confers antigen-presenting-cell function to parenchymal tissue and in conjunction with tumor www.annualreviews.org • Regulation of Immune Responses by TGF-β

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5.

6.

7.

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8. 9. 10. 11. 12. First study of TGF-β on immune cells.

First demonstration of the requirement of TGF-β1 for immune tolerance.

13.

14.

15.

16.

17.

Demonstrated TGF-β regulation of T cell activity in vivo.

Showed TGF-β regulation of B cell activity in vivo.

18. 19.

20.

21. Study of TGF-β regulation of NK cell activity in vivo.

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22.

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necrosis factor α leads to autoimmunity in transgenic mice. Proc. Natl. Acad. Sci. USA 91:5138–42 Li M, Carpio DF, Zheng Y, Bruzzo P, Singh V, et al. 2001. An essential role of the NF-κB/Toll-like receptor pathway in induction of inﬂammatory and tissue-repair gene expression by necrotic cells. J. Immunol. 166:7128–35 Leadbetter EA, Rifkin IR, Hohlbaum AM, Beaudette BC, Shlomchik MJ, MarshakRothstein A. 2002. Chromatin-IgG complexes activate B cells by dual engagement of IgM and Toll-like receptors. Nature 416:603–7 Sakaguchi S. 2004. Naturally arising CD4+ regulatory T cells for immunologic selftolerance and negative control of immune responses. Annu. Rev. Immunol. 22:531–62 Moore KW, de Waal Malefyt R, Coffman RL, O’Garra A. 2001. Interleukin-10 and the interleukin-10 receptor. Annu. Rev. Immunol. 19:683–765 Letterio JJ, Roberts AB. 1998. Regulation of immune responses by TGF-β. Annu. Rev. Immunol. 16:137–61 Wahl SM. 1994. Transforming growth factor β: the good, the bad, and the ugly. J. Exp. Med. 180:1587–90 Blobe GC, Schiemann WP, Lodish HF. 2000. Role of transforming growth factor β in human disease. N. Engl. J. Med. 342:1350–58 Newfeld SJ, Wisotzkey RG, Kumar S. 1999. Molecular evolution of a developmental pathway: phylogenetic analyses of transforming growth factor-β family ligands, receptors and Smad signal transducers. Genetics 152:783–95 Kehrl JH, Wakefield LM, Roberts AB, Jakowlew S, Alvarez-Mon M, et al. 1986. Production of transforming growth factor β by human T lymphocytes and its potential role in the regulation of T cell growth. J. Exp. Med. 163:1037–50 Shull MM, Ormsby I, Kier AB, Pawlowski S, Diebold RJ, et al. 1992. Targeted disruption of the mouse transforming growth factor-β1 gene results in multifocal inflammatory disease. Nature 359:693–99 Kulkarni AB, Huh CG, Becker D, Geiser A, Lyght M, et al. 1993. Transforming growth factor β1 null mutation in mice causes excessive inﬂammatory response and early death. Proc. Natl. Acad. Sci. USA 90:770–74 Yang X, Letterio JJ, Lechleider RJ, Chen L, Hayman R, et al. 1999. Targeted disruption of SMAD3 results in impaired mucosal immunity and diminished T cell responsiveness to TGF-β. EMBO J. 18:1280–91 Datto MB, Frederick JP, Pan L, Borton AJ, Zhuang Y, et al. 1999. Targeted disruption of Smad3 reveals an essential role in transforming growth factor β-mediated signal transduction. Mol. Cell. Biol. 19:2495–504 Gorelik L, Flavell RA. 2000. Abrogation of TGFβ signaling in T cells leads to spontaneous T cell differentiation and autoimmune disease. Immunity 12:171–81 Lucas PJ, Kim SJ, Melby SJ, Gress RE. 2000. Disruption of T cell homeostasis in mice expressing a T cell-speciﬁc dominant negative transforming growth factor β II receptor. J. Exp. Med. 191:1187–96 Nakao A, Miike S, Hatano M, Okumura K, Tokuhisa T, et al. 2000. Blockade of transforming growth factor β/Smad signaling in T cells by overexpression of Smad7 enhances antigen-induced airway inﬂammation and airway reactivity. J. Exp. Med. 192:151–58 Cazac BB, Roes J. 2000. TGF-β receptor controls B cell responsiveness and induction of IgA in vivo. Immunity 13:443–51 Laouar Y, Sutterwala FS, Gorelik L, Flavell RA. 2005. Transforming growth factorβ controls T helper type 1 cell development through regulation of natural killer cell interferon-γ. Nat. Immunol. 6:600–7

Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

23. Chang H, Brown CW, Matzuk MM. 2002. Genetic analysis of the mammalian transforming growth factor-β superfamily. Endocr. Rev. 23:787–823 24. Govinden R, Bhoola KD. 2003. Genealogy, expression, and cellular function of transforming growth factor-β. Pharmacol. Ther. 98:257–65 25. Dubois CM, Laprise MH, Blanchette F, Gentry LE, Leduc R. 1995. Processing of transforming growth factor β1 precursor by human furin convertase. J. Biol. Chem. 270:10618– 24 26. Annes JP, Munger JS, Rifkin DB. 2003. Making sense of latent TGF β activation. J. Cell Sci. 116:217–24 27. Nunes I, Shapiro RL, Rifkin DB. 1995. Characterization of latent TGF-β activation by murine peritoneal macrophages. J. Immunol. 155:1450–59 28. Crawford SE, Stellmach V, Murphy-Ullrich JE, Ribeiro SM, Lawler J, et al. 1998. Thrombospondin-1 is a major activator of TGF-β1 in vivo. Cell 93:1159–70 29. Yehualaeshet T, O’Connor R, Green-Johnson J, Mai S, Silverstein R, et al. 1999. Activation of rat alveolar macrophage-derived latent transforming growth factor β-1 by plasmin requires interaction with thrombospondin-1 and its cell surface receptor, CD36. Am. J. Pathol. 155:841–51 30. Munger JS, Huang X, Kawakatsu H, Grifﬁths MJ, Dalton SL, et al. 1999. The integrin αvβ6 binds and activates latent TGFβ1: a mechanism for regulating pulmonary inﬂammation and ﬁbrosis. Cell 96:319–28 31. Annes JP, Chen Y, Munger JS, Rifkin DB. 2004. Integrin αVβ6-mediated activation of latent TGF-β requires the latent TGF-β binding protein-1. J. Cell Biol. 165:723–34 32. Van Obberghen-Schilling E, Roche NS, Flanders KC, Sporn MB, Roberts AB. 1988. Transforming growth factor β1 positively regulates its own expression in normal and transformed cells. J. Biol. Chem. 263:7741–46 33. Kim SJ, Jeang KT, Glick AB, Sporn MB, Roberts AB. 1989. Promoter sequences of the human transforming growth factor-β1 gene responsive to transforming growth factor-β1 autoinduction. J. Biol. Chem. 264:7041–45 34. Kim SJ, Angel P, Lafyatis R, Hattori K, Kim KY, et al. 1990. Autoinduction of transforming growth factor β1 is mediated by the AP-1 complex. Mol. Cell. Biol. 10:1492–97 35. Ventura JJ, Kennedy NJ, Flavell RA, Davis RJ. 2004. JNK regulates autocrine expression of TGF-β1. Mol. Cell 15:269–78 36. Kim SJ, Park K, Koeller D, Kim KY, Wakeﬁeld LM, et al. 1992. Post-transcriptional regulation of the human transforming growth factor-β1 gene. J. Biol. Chem. 267:13702– 7 37. Allison RS, Mumy ML, Wakeﬁeld LM. 1998. Translational control elements in the major human transforming growth factor-β1 mRNA. Growth Factors 16:89–100 38. Roberts AB, Sporn MB. 1992. Differential expression of the TGF-β isoforms in embryogenesis suggests speciﬁc roles in developing and adult tissues. Mol. Reprod. Dev. 32:91–98 39. Brown CB, Boyer AS, Runyan RB, Barnett JV. 1999. Requirement of type III TGF-β receptor for endocardial cell transformation in the heart. Science 283:2080–82 40. Massague J. 1998. TGF-β signal transduction. Annu. Rev. Biochem. 67:753–91 41. ten Dijke P, Hill CS. 2004. New insights into TGF-β-Smad signalling. Trends Biochem. Sci. 29:265–73 42. Huse M, Muir TW, Xu L, Chen YG, Kuriyan J, Massague J. 2001. The TGF β receptor activation process: an inhibitor- to substrate-binding switch. Mol. Cell 8:671–82 43. Inman GJ, Nicolas FJ, Hill CS. 2002. Nucleocytoplasmic shuttling of Smads 2, 3, and 4 permits sensing of TGF-β receptor activity. Mol. Cell 10:283–94 www.annualreviews.org • Regulation of Immune Responses by TGF-β

127

ARI

23 February 2006

15:9

44. Johnson K, Kirkpatrick H, Comer A, Hoffmann FM, Laughon A. 1999. Interaction of Smad complexes with tripartite DNA-binding sites. J. Biol. Chem. 274:20709–16 45. Shi Y, Wang YF, Jayaraman L, Yang H, Massague J, Pavletich NP. 1998. Crystal structure of a Smad MH1 domain bound to DNA: insights on DNA binding in TGF-β signaling. Cell 94:585–94 46. Zawel L, Dai JL, Buckhaults P, Zhou S, Kinzler KW, et al. 1998. Human Smad3 and Smad4 are sequence-speciﬁc transcription activators. Mol. Cell 1:611–67 47. Liu F, Pouponnot C, Massague J. 1997. Dual role of the Smad4/DPC4 tumor suppressor in TGFβ-inducible transcriptional complexes. Genes Dev. 11:3157–67 48. Nakao A, Afrakhte M, Moren A, Nakayama T, Christian JL, et al. 1997. Identiﬁcation of Smad7, a TGFβ-inducible antagonist of TGF-β signalling. Nature 389:631–35 49. Kavsak P, Rasmussen RK, Causing CG, Bonni S, Zhu H, et al. 2000. Smad7 binds to Smurf2 to form an E3 ubiquitin ligase that targets the TGF β receptor for degradation. Mol. Cell 6:1365–75 50. Ebisawa T, Fukuchi M, Murakami G, Chiba T, Tanaka K, et al. 2001. Smurf1 interacts with transforming growth factor-β type I receptor through Smad7 and induces receptor degradation. J. Biol. Chem. 276:12477–80 51. Attisano L, Wrana JL. 2000. Smads as transcriptional co-modulators. Curr. Opin. Cell Biol. 12:235–43 52. Massague J, Wotton D. 2000. Transcriptional control by the TGF-β/Smad signaling system. EMBO J. 19:1745–54 53. ten Dijke P, Miyazono K, Heldin CH. 2000. Signaling inputs converge on nuclear effectors in TGF-β signaling. Trends Biochem. Sci. 25:64–70 54. Derynck R, Zhang YE. 2003. Smad-dependent and Smad-independent pathways in TGF-β family signalling. Nature 425:577–84 55. Shi Y, Massague J. 2003. Mechanisms of TGF-β signaling from cell membrane to the nucleus. Cell 113:685–700 56. Engel ME, McDonnell MA, Law BK, Moses HL. 1999. Interdependent SMAD and JNK signaling in transforming growth factor-β-mediated transcription. J. Biol. Chem. 274:37413–20 57. Yu L, Hebert MC, Zhang YE. 2002. TGF-β receptor-activated p38 MAP kinase mediates Smad-independent TGF-β responses. EMBO J. 21:3749–59 58. Kretzschmar M, Doody J, Timokhina I, Massague J. 1999. A mechanism of repression of TGFβ/Smad signaling by oncogenic Ras. Genes Dev. 13:804–16 59. Funaba M, Zimmerman CM, Mathews LS. 2002. Modulation of Smad2-mediated signaling by extracellular signal-regulated kinase. J. Biol. Chem. 277:41361–68 60. Blanchette F, Rivard N, Rudd P, Grondin F, Attisano L, Dubois CM. 2001. Cross-talk between the p42/p44 MAP kinase and Smad pathways in transforming growth factor β1-induced furin gene transactivation. J. Biol. Chem. 276:33986–94 61. Choy L, Derynck R. 1998. The type II transforming growth factor (TGF)-β receptorinteracting protein TRIP-1 acts as a modulator of the TGF-β response. J. Biol. Chem. 273:31455–62 62. McGonigle S, Beall MJ, Pearce EJ. 2002. Eukaryotic initiation factor 2α subunit associates with TGFβ receptors and 14–3–3ε and acts as a modulator of the TGFβ response. Biochemistry 41:579–87 63. Griswold-Prenner I, Kamibayashi C, Maruoka EM, Mumby MC, Derynck R. 1998. Physical and functional interactions between type I transforming growth factor β receptors and Bα, a WD-40 repeat subunit of phosphatase 2A. Mol. Cell. Biol. 18:6595–604

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

128

Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

64. Gorelik L, Flavell RA. 2002. Transforming growth factor-β in T-cell biology. Nat. Rev. Immunol 2:46–53 65. Cerwenka A, Swain SL. 1999. TGF-β1: immunosuppressant and viability factor for T lymphocytes. Microbes Infect. 1:1291–96 66. Brabletz T, Pfeuffer I, Schorr E, Siebelt F, Wirth T, Serﬂing E. 1993. Transforming growth factor β and cyclosporin A inhibit the inducible activity of the interleukin-2 gene in T cells through a noncanonical octamer-binding site. Mol. Cell. Biol. 13:1155–62 67. McKarns SC, Schwartz RH, Kaminski NE. 2004. Smad3 is essential for TGF-β1 to suppress IL-2 production and TCR-induced proliferation, but not IL-2-induced proliferation. J. Immunol. 172:4275–84 68. Tzachanis D, Freeman GJ, Hirano N, van Puijenbroek AA, Delfs MW, et al. 2001. Tob is a negative regulator of activation that is expressed in anergic and quiescent T cells. Nat. Immunol. 2:1174–82 69. Hannon GJ, Beach D. 1994. p15INK4B is a potential effector of TGF-β-induced cell cycle arrest. Nature 371:257–61 70. Datto MB, Li Y, Panus JF, Howe DJ, Xiong Y, Wang XF. 1995. Transforming growth factor β induces the cyclin-dependent kinase inhibitor p21 through a p53-independent mechanism. Proc. Natl. Acad. Sci. USA 92:5545–49 71. Polyak K, Kato JY, Solomon MJ, Sherr CJ, Massague J, et al. 1994. p27Kip1, a cyclin-Cdk inhibitor, links transforming growth factor-β and contact inhibition to cell cycle arrest. Genes Dev. 8:9–22 72. Coffey RJ Jr, Bascom CC, Sipes NJ, Graves-Deal R, Weissman BE, et al. 1988. Selective inhibition of growth-related gene expression in murine keratinocytes by transforming growth factor β. Mol. Cell. Biol. 8:3088–93 73. Wolfraim LA, Walz TM, James Z, Fernandez T, Letterio JJ. 2004. p21Cip1 and p27Kip1 act in synergy to alter the sensitivity of naive T cells to TGF-β-mediated G1 arrest through modulation of IL-2 responsiveness. J. Immunol. 173:3093–102 74. Ruegemer JJ, Ho SN, Augustine JA, Schlager JW, Bell MP, et al. 1990. Regulatory effects of transforming growth factor-β on IL-2- and IL-4-dependent T cell-cycle progression. J. Immunol. 144:1767–76 75. Genestier L, Kasibhatla S, Brunner T, Green DR. 1999. Transforming growth factor β1 inhibits Fas ligand expression and subsequent activation-induced cell death in T cells via downregulation of c-Myc. J. Exp. Med. 189:231–39 76. Nelson BH, Martyak TP, Thompson LJ, Moon JJ, Wang T. 2003. Uncoupling of promitogenic and antiapoptotic functions of IL-2 by Smad-dependent TGF-β signaling. J. Immunol. 170:5563–70 77. McKarns SC, Schwartz RH. 2005. Distinct effects of TGF-β1 on CD4+ and CD8+ T cell survival, division, and IL-2 production: a role for T cell intrinsic Smad3. J. Immunol. 174:2071–83 78. Cottrez F, Groux H. 2001. Regulation of TGF-β response during T cell activation is modulated by IL-10. J. Immunol. 167:773–78 79. Sung JL, Lin JT, Gorham JD. 2003. CD28 co-stimulation regulates the effect of transforming growth factor-β1 on the proliferation of naive CD4+ T cells. Int. Immunol. 3:233–45 80. Bommireddy R, Ormsby I, Yin M, Boivin GP, Babcock GF, Doetschman T. 2003. TGFβ1 inhibits Ca2+ -calcineurin-mediated activation in thymocytes. J. Immunol. 170:3645–52 81. Lucas PJ, McNeil N, Hilgenfeld E, Choudhury B, Kim SJ, et al. 2004. Transforming growth factor-β pathway serves as a primary tumor suppressor in CD8+ T cell tumorigenesis. Cancer Res. 64:6524–29 www.annualreviews.org • Regulation of Immune Responses by TGF-β

129

ARI

23 February 2006

15:9

82. Wolfraim LA, Fernandez TM, Mamura M, Fuller WL, Kumar R, et al. 2004. Loss of Smad3 in acute T-cell lymphoblastic leukemia. N. Engl. J. Med. 351:552–59 83. Murphy KM, Reiner SL. 2002. The lineage decisions of helper T cells. Nat. Rev. Immunol. 2:933–44 84. Sad S, Mosmann TR. 1994. Single IL-2-secreting precursor CD4 T cell can develop into either Th1 or Th2 cytokine secretion phenotype. J. Immunol. 153:3514–22 85. Gorelik L, Fields PE, Flavell RA. 2000. Cutting edge: TGF-β inhibits Th type 2 development through inhibition of GATA-3 expression. J. Immunol. 165:4773–77 86. Heath VL, Murphy EE, Crain C, Tomlinson MG, O’Garra A. 2000. TGF-β1 downregulates Th2 development and results in decreased IL-4-induced STAT6 activation and GATA-3 expression. Eur. J. Immunol. 30:2639–49 87. Chen CH, Seguin-Devaux C, Burke NA, Oriss TB, Watkins SC, et al. 2003. Transforming growth factor β blocks Tec kinase phosphorylation, Ca2+ inﬂux, and NFATc translocation causing inhibition of T cell differentiation. J. Exp. Med. 197:1689–99 88. Gorham JD, Guler ML, Fenoglio D, Gubler U, Murphy KM. 1998. Low dose TGF-β attenuates IL-12 responsiveness in murine Th cells. J. Immunol. 161:1664–70 89. Gorelik L, Constant S, Flavell RA. 2002. Mechanism of transforming growth factor βinduced inhibition of T helper type 1 differentiation. J. Exp. Med. 195:1499–505 90. Afkarian M, Sedy JR, Yang J, Jacobson NG, Cereb N, et al. 2002. T-bet is a STAT1induced regulator of IL-12R expression in naive CD4+ T cells. Nat. Immunol. 3:549–57 91. Szabo SJ, Kim ST, Costa GL, Zhang X, Fathman CG, Glimcher LH. 2000. A novel transcription factor, T-bet, directs Th1 lineage commitment. Cell 100:655–69 92. Lin JT, Martin SL, Xia L, Gorham JD. 2005. TGF-β1 uses distinct mechanisms to inhibit IFN-γ expression in CD4+ T cells at priming and at recall: differential involvement of Stat4 and T-bet. J. Immunol. 174:5950–58 93. Ludviksson BR, Seegers D, Resnick AS, Strober W. 2000. The effect of TGF-β1 on immune responses of naive versus memory CD4+ Th1/Th2 T cells. Eur. J. Immunol. 30:2101–11 94. Ulloa L, Doody J, Massague J. 1999. Inhibition of transforming growth factor-β/SMAD signalling by the interferon-γ/STAT pathway. Nature 397:710–13 95. Kitani A, Fuss I, Nakamura K, Kumaki F, Usui T, Strober W. 2003. Transforming growth factor (TGF)-β1-producing regulatory T cells induce Smad-mediated interleukin 10 secretion that facilitates coordinated immunoregulatory activity and amelioration of TGFβ1-mediated ﬁbrosis. J. Exp. Med. 198:1179–88 96. Fuss IJ, Boirivant M, Lacy B, Strober W. 2002. The interrelated roles of TGF-β and IL-10 in the regulation of experimental colitis. J. Immunol. 168:900–8 97. Schmitt E, Germann T, Goedert S, Hoehn P, Huels C, et al. 1994. IL-9 production of naive CD4+ T cells depends on IL-2, is synergistically enhanced by a combination of TGF-β and IL-4, and is inhibited by IFN-γ. J. Immunol. 153:3989–96 98. Rudner LA, Lin JT, Park IK, Cates JM, Dyer DA, et al. 2003. Necroinﬂammatory liver disease in BALB/c background, TGF-β1-deﬁcient mice requires CD4+ T cells. J. Immunol. 170:4785–92 99. Letterio JJ, Geiser AG, Kulkarni AB, Dang H, Kong L, et al. 1996. Autoimmunity associated with TGF-β1-deﬁciency in mice is dependent on MHC class II antigen expression. J. Clin. Invest. 98:2109–19 100. Seder RA, Ahmed R. 2003. Similarities and differences in CD4+ and CD8+ effector and memory T cell generation. Nat. Immunol. 4:835–42

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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Li et al.

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ANRV270-IY24-04

ARI

23 February 2006

15:9

101. Ranges GE, Figari IS, Espevik T, Palladino MA Jr. 1987. Inhibition of cytotoxic T cell development by transforming growth factor β and reversal by recombinant tumor necrosis factor α. J. Exp. Med. 166:991–98 102. Smyth MJ, Strobl SL, Young HA, Ortaldo JR, Ochoa AC. 1991. Regulation of lymphokine-activated killer activity and pore-forming protein gene expression in human peripheral blood CD8+ T lymphocytes. Inhibition by transforming growth factor-β. J. Immunol. 146:3289–97 103. Bonig H, Banning U, Hannen M, Kim YM, Verheyen J, et al. 1999. Transforming growth factor-β1 suppresses interleukin-15-mediated interferon-γ production in human T lymphocytes. Scand. J. Immunol. 50:612–18 104. Ahmadzadeh M, Rosenberg SA. 2005. TGF-β1 attenuates the acquisition and expression of effector function by tumor antigen-speciﬁc human memory CD8 T cells. J. Immunol. 174:5215–23 105. Glimcher LH, Townsend MJ, Sullivan BM, Lord GM. 2004. Recent developments in the transcriptional regulation of cytolytic effector cells. Nat. Rev. Immunol. 4:900–11 106. Croft M. 2003. Co-stimulatory members of the TNFR family: keys to effective T-cell immunity? Nat. Rev. Immunol. 3:609–20 107. Kim YJ, Stringﬁeld TM, Chen Y, Broxmeyer HE. 2005. Modulation of cord blood CD8+ T-cell effector differentiation by TGF-β1 and 4-1BB costimulation. Blood 105:274–81 108. Kobayashi S, Yoshida K, Ward JM, Letterio JJ, Longenecker G, et al. 1999. β2 microglobulin-deﬁcient background ameliorates lethal phenotype of the TGF-β1 null mouse. J. Immunol. 163:4013–19 109. Gorelik L, Flavell RA. 2001. Immune-mediated eradication of tumors through the blockade of transforming growth factor-β signaling in T cells. Nat. Med. 7:1118–22 110. Gershon RK. 1975. A disquisition on suppressor T cells. Transplant Rev. 26:170–85 111. Fontenot JD, Rudensky AY. 2005. A well adapted regulatory contrivance: regulatory T cell development and the forkhead family transcription factor Foxp3. Nat. Immunol. 6:331–37 112. Zelenay S, Lopes-Carvalho T, Caramalho I, Moraes-Fontes MF, Rebelo M, Demengeot J. 2005. Foxp3+ CD25− CD4 T cells constitute a reservoir of committed regulatory cells that regain CD25 expression upon homeostatic expansion. Proc. Natl. Acad. Sci. USA 102:4091–96 113. Liang S, Alard P, Zhao Y, Parnell S, Clark SL, Kosiewicz MM. 2005. Conversion of CD4+ CD25− cells into CD4+ CD25+ regulatory T cells in vivo requires B7 costimulation, but not the thymus. J. Exp. Med. 201:127–37 114. Chen Y, Kuchroo VK, Inobe J, Hafler DA, Weiner HL. 1994. Regulatory T cell clones induced by oral tolerance: suppression of autoimmune encephalomyelitis. Science 265:1237–40 115. Horwitz DA, Zheng SG, Gray JD. 2003. The role of the combination of IL-2 and TGF-β or IL-10 in the generation and function of CD4+ CD25+ and CD8+ regulatory T cell subsets. J. Leukoc. Biol. 74:471–78 116. Chen W, Jin W, Hardegen N, Lei KJ, Li L, et al. 2003. Conversion of peripheral CD4+ CD25− naive T cells to CD4+ CD25+ regulatory T cells by TGF-β induction of transcription factor Foxp3. J. Exp. Med. 198:1875–86 117. Fantini MC, Becker C, Monteleone G, Pallone F, Galle PR, Neurath MF. 2004. Cutting edge: TGF-β induces a regulatory phenotype in CD4+ CD25− T cells through Foxp3 induction and down-regulation of Smad7. J. Immunol. 172:5149–53 118. Zheng SG, Wang JH, Gray JD, Soucier H, Horwitz DA. 2004. Natural and induced CD4+ CD25+ cells educate CD4+ CD25− cells to develop suppressive activity: the role of IL-2, TGF-β, and IL-10. J. Immunol. 172:5213–21 www.annualreviews.org • Regulation of Immune Responses by TGF-β

Study of TGF-β regulation of oral tolerance.

First demonstration of TGF-β induction of FoxP3 and regulatory T cell generation.

131

ARI

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119. Wan YY, Flavell RA. 2005. Identifying Foxp3-expressing suppressor T cells with a bicistronic reporter. Proc. Natl. Acad. Sci. USA 102:5126–31 120. Peng Y, Laouar Y, Li MO, Green EA, Flavell RA. 2004. TGF-β regulates in vivo expansion of Foxp3-expressing CD4+ CD25+ regulatory T cells responsible for protection against diabetes. Proc. Natl. Acad. Sci. USA 101:4572–77 121. Huber S, Schramm C, Lehr HA, Mann A, Schmitt S, et al. 2004. Cutting edge: TGF-β signaling is required for the in vivo expansion and immunosuppressive capacity of regulatory CD4+ CD25+ T cells. J. Immunol. 173:6526–31 122. Marie JC, Letterio JJ, Gavin M, Rudensky AY. 2005. TGF-β1 maintains suppressor function and Foxp3 expression in CD4+ CD25+ regulatory T cells. J. Exp. Med. 201:1061– 67 123. Mamura M, Lee W, Sullivan TJ, Felici A, Sowers AL, et al. 2004. CD28 disruption exacerbates inﬂammation in Tgf-β1−/− mice: in vivo suppression by CD4+ CD25+ regulatory T cells independent of autocrine TGF-β1. Blood 103:4594–601 124. Fahlen L, Read S, Gorelik L, Hurst SD, Coffman RL, et al. 2005. T cells that cannot respond to TGF-β escape control by CD4+ CD25+ regulatory T cells. J. Exp. Med. 201:737–46 125. Fontenot JD, Rasmussen JP, Williams LM, Dooley JL, Farr AG, Rudensky AY. 2005. Regulatory T cell lineage speciﬁcation by the forkhead transcription factor Foxp3. Immunity 22:329–41 126. Green DR, Droin N, Pinkoski M. 2003. Activation-induced cell death in T cells. Immunol. Rev. 193:70–81 127. Marrack P, Kappler J. 2004. Control of T cell viability. Annu. Rev. Immunol. 22:765–87 128. Gunnlaugsdottir B, Maggadottir SM, Ludviksson BR. 2005. Anti-CD28-induced costimulation and TCR avidity regulates the differential effect of TGF-β1 on CD4+ and CD8+ naive human T-cells. Int. Immunol. 17:35–44 129. Swain SL, Huston G, Tonkonogy S, Weinberg A. 1991. Transforming growth factor-β and IL-4 cause helper T cell precursors to develop into distinct effector helper cells that differ in lymphokine secretion pattern and cell surface phenotype. J. Immunol. 147:2991–3000 130. Weinberg AD, Whitham R, Swain SL, Morrison WJ, Wyrick G, et al. 1992. Transforming growth factor-β enhances the in vivo effector function and memory phenotype of antigen-speciﬁc T helper cells in experimental autoimmune encephalomyelitis. J. Immunol. 148:2109–17 131. de Jong R, van Lier RA, Ruscetti FW, Schmitt C, Debre P, Mossalayi MD. 1994. Differential effect of transforming growth factor-β1 on the activation of human naive and memory CD4+ T lymphocytes. Int. Immunol. 6:631–38 132. Zhang X, Giangreco L, Broome HE, Dargan CM, Swain SL. 1995. Control of CD4 effector fate: transforming growth factor β1 and interleukin 2 synergize to prevent apoptosis and promote effector expansion. J. Exp. Med. 182:699–709 133. Swain SL, Croft M, Dubey C, Haynes L, Rogers P, et al. 1996. From naive to memory T cells. Immunol. Rev. 150:143–67 134. Rich S, Van Nood N, Lee HM. 1996. Role of alpha 5 beta 1 integrin in TGF-beta 1costimulated CD8+ T cell growth and apoptosis. J. Immunol. 157:2916–23 135. Cerwenka A, Kovar H, Majdic O, Holter W. 1996. Fas- and activation-induced apoptosis are reduced in human T cells preactivated in the presence of TGF-β1. J. Immunol. 156:459–64 136. Chen W, Jin W, Tian H, Sicurello P, Frank M, et al. 2001. Requirement for transforming growth factor β1 in controlling T cell apoptosis. J. Exp. Med. 194:439–53

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23 February 2006

15:9

137. Bommireddy R, Saxena V, Ormsby I, Yin M, Boivin GP, et al. 2003. TGF-β1 regulates lymphocyte homeostasis by preventing activation and subsequent apoptosis of peripheral lymphocytes. J. Immunol. 170:4612–22 138. Refaeli Y, Van Parijs L, Alexander SI, Abbas AK. 2002. Interferon γ is required for activation-induced death of T lymphocytes. J. Exp. Med. 196:999–1005 139. Weiner HL. 2001. Induction and mechanism of action of transforming growth factor-βsecreting Th3 regulatory cells. Immunol. Rev. 182:207–14 140. Nakamura K, Kitani A, Strober W. 2001. Cell contact-dependent immunosuppression by CD4+ CD25+ regulatory T cells is mediated by cell surface-bound transforming growth factor β. J. Exp. Med. 194:629–44 141. Oida T, Zhang X, Goto M, Hachimura S, Totsuka M, et al. 2003. CD4+ CD25− T cells that express latency-associated peptide on the surface suppress CD4+ CD45RBhigh -induced colitis by a TGF-β-dependent mechanism. J. Immunol. 170:2516–22 142. Green EA, Gorelik L, McGregor CM, Tran EH, Flavell RA. 2003. CD4+ CD25+ T regulatory cells control anti-islet CD8+ T cells through TGF-β-TGF-β receptor interactions in type 1 diabetes. Proc. Natl. Acad. Sci. USA 100:10878–83 143. Piccirillo CA, Letterio JJ, Thornton AM, McHugh RS, Mamura M, et al. 2002. CD4+ CD25+ regulatory T cells can mediate suppressor function in the absence of transforming growth factor β1 production and responsiveness. J. Exp. Med. 196:237–46 144. Powrie F, Carlino J, Leach MW, Mauze S, Coffman RL. 1996. A critical role for transforming growth factor-β but not interleukin 4 in the suppression of T helper type 1mediated colitis by CD45RBlow CD4+ T cells. J. Exp. Med. 183:2669–74 145. Nakamura K, Kitani A, Fuss I, Pedersen A, Harada N, et al. 2004. TGF-β1 plays an important role in the mechanism of CD4+ CD25+ regulatory T cell activity in both humans and mice. J. Immunol. 172:834–42 146. Chen W, Jin W, Wahl SM. 1998. Engagement of cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) induces transforming growth factor β (TGF-β) production by murine CD4+ T cells. J. Exp. Med. 188:1849–57 147. Gomes NA, Gattass CR, Barreto-De-Souza V, Wilson ME, DosReis GA. 2000. TGF-β mediates CTLA-4 suppression of cellular immunity in murine kalaazar. J. Immunol. 164:2001–8 148. Kitani A, Chua K, Nakamura K, Strober W. 2000. Activated self-MHC-reactive T cells have the cytokine phenotype of Th3/T regulatory cell 1 T cells. J. Immunol. 165:691–702 149. Sullivan TJ, Letterio JJ, van Elsas A, Mamura M, van Amelsfort J, et al. 2001. Lack of a role for transforming growth factor-β in cytotoxic T lymphocyte antigen-4-mediated inhibition of T cell activation. Proc. Natl. Acad. Sci. USA 98:2587–92 150. Esplugues E, Sancho D, Vega-Ramos J, Martinez C, Syrbe U, et al. 2003. Enhanced antitumor immunity in mice deﬁcient in CD69. J. Exp. Med. 197:1093–106 151. Sancho D, Gomez M, Viedma F, Esplugues E, Gordon-Alonso M, et al. 2003. CD69 downregulates autoimmune reactivity through active transforming growth factor-β production in collagen-induced arthritis. J. Clin. Invest. 112:872–82 152. Seder RA, Marth T, Sieve MC, Strober W, Letterio JJ, et al. 1998. Factors involved in the differentiation of TGF-β-producing cells from naive CD4+ T cells: IL-4 and IFN-γ have opposing effects, while TGF-β positively regulates its own production. J. Immunol. 160:5719–28 153. Neurath MF, Weigmann B, Finotto S, Glickman J, Nieuwenhuis E, et al. 2002. The transcription factor T-bet regulates mucosal T cell activation in experimental colitis and Crohn’s disease. J. Exp. Med. 195:1129–43 www.annualreviews.org • Regulation of Immune Responses by TGF-β

133

ARI

23 February 2006

15:9

154. Chen W, Frank ME, Jin W, Wahl SM. 2001. TGF-β released by apoptotic T cells contributes to an immunosuppressive milieu. Immunity 14:715–25 155. Stavnezer J. 1995. Regulation of antibody production and class switching by TGF-β. J. Immunol. 155:1647–51 156. Lebman DA, Edmiston JS. 1999. The role of TGF-β in growth, differentiation, and maturation of B lymphocytes. Microbes Infect. 1:1297–304 157. Kee BL, Rivera RR, Murre C. 2001. Id3 inhibits B lymphocyte progenitor growth and survival in response to TGF-β. Nat. Immunol. 2:242–47 158. Kehrl JH, Roberts AB, Wakeﬁeld LM, Jakowlew S, Sporn MB, Fauci AS. 1986. Transforming growth factor β is an important immunomodulatory protein for human B lymphocytes. J. Immunol. 137:3855–60 159. Petit-Koskas E, Genot E, Lawrence D, Kolb JP. 1988. Inhibition of the proliferative response of human B lymphocytes to B cell growth factor by transforming growth factorβ. Eur. J. Immunol. 18:111–16 160. Smeland EB, Blomhoff HK, Holte H, Ruud E, Beiske K, et al. 1987. Transforming growth factor type β (TGF β) inhibits G1 to S transition, but not activation of human B lymphocytes. Exp. Cell Res. 171:213–22 161. Bouchard C, Fridman WH, Sautes C. 1997. Effect of TGF-β1 on cell cycle regulatory proteins in LPS-stimulated normal mouse B lymphocytes. J. Immunol. 159:4155–64 162. Kamesaki H, Nishizawa K, Michaud GY, Cossman J, Kiyono T. 1998. TGF-β1 induces the cyclin-dependent kinase inhibitor p27Kip1 mRNA and protein in murine B cells. J. Immunol. 160:770–77 163. Roes J, Choi BK, Cazac BB. 2003. Redirection of B cell responsiveness by transforming growth factor β receptor. Proc. Natl. Acad. Sci. USA 100:7241–46 164. Patil S, Wildey GM, Brown TL, Choy L, Derynck R, Howe PH. 2000. Smad7 is induced by CD40 and protects WEHI 231 B-lymphocytes from transforming growth factor-βinduced growth inhibition and apoptosis. J. Biol. Chem. 275:38363–70 165. Briskin M, Kuwabara MD, Sigman DS, Wall R. 1988. Induction of κ transcription by interferon-γ without activation of NF-κB. Science 242:1036–37 166. Kehrl JH, Thevenin C, Rieckmann P, Fauci AS. 1991. Transforming growth factor-β suppresses human B lymphocyte Ig production by inhibiting synthesis and the switch from the membrane form to the secreted form of Ig mRNA. J. Immunol. 146:4016–23 167. Lebman DA, Lee FD, Coffman RL. 1990. Mechanism for transforming growth factor β and IL-2 enhancement of IgA expression in lipopolysaccharide-stimulated B cell cultures. J. Immunol. 144:952–59 168. Kim PH, Kagnoff MF. 1990. Transforming growth factor beta 1 increases IgA isotype switching at the clonal level. J. Immunol. 145:3773–78 169. Defrance T, Vanbervliet B, Briere F, Durand I, Rousset F, Banchereau J. 1992. Interleukin 10 and transforming growth factor β cooperate to induce anti-CD40-activated naive human B cells to secrete immunoglobulin A. J. Exp. Med. 175:671–82 170. van Vlasselaer P, Punnonen J, de Vries JE. 1992. Transforming growth factor-β directs IgA switching in human B cells. J. Immunol. 148:2062–67 171. McIntyre TM, Klinman DR, Rothman P, Lugo M, Dasch JR, et al. 1993. Transforming growth factor β1 selectivity stimulates immunoglobulin G2b secretion by lipopolysaccharide-activated murine B cells. J. Exp. Med. 177:1031–37 172. Fagarasan S, Honjo T. 2003. Intestinal IgA synthesis: regulation of front-line body defences. Nat. Rev. Immunol. 3:63–72 173. McIntyre TM, Kehry MR, Snapper CM. 1995. Novel in vitro model for high-rate IgA class switching. J. Immunol. 154:3156–61

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

134

Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

174. Shockett P, Stavnezer J. 1991. Effect of cytokines on switching to IgA and α germline transcripts in the B lymphoma I.29 μ: transforming growth factor-β activates transcription of the unrearranged Cα gene. J. Immunol. 147:4374–83 175. Lin YC, Stavnezer J. 1992. Regulation of transcription of the germ-line Ig α constant region gene by an ATF element and by novel transforming growth factor-β1-responsive elements. J. Immunol. 149:2914–25 176. Shi MJ, Stavnezer J. 1998. CBF α3 (AML2) is induced by TGF-β1 to bind and activate the mouse germline Ig α promoter. J. Immunol. 161:6751–60 177. Hanai J, Chen LF, Kanno T, Ohtani-Fujita N, Kim WY, et al. 1999. Interaction and functional cooperation of PEBP2/CBF with Smads. Synergistic induction of the immunoglobulin germline Cα promoter. J. Biol. Chem. 274:31577–82 178. Pardali E, Xie XQ, Tsapogas P, Itoh S, Arvanitidis K, et al. 2000. Smad and AML proteins synergistically confer transforming growth factor β1 responsiveness to human germ-line IgA genes. J. Biol. Chem. 275:3552–60 179. Park SR, Lee JH, Kim PH. 2001. Smad3 and Smad4 mediate transforming growth factorβ1-induced IgA expression in murine B lymphocytes. Eur. J. Immunol. 31:1706–15 180. Park SR, Lee EK, Kim BC, Kim PH. 2003. p300 cooperates with Smad3/4 and Runx3 in TGFβ1-induced IgA isotype expression. Eur. J. Immunol. 33:3386–92 181. Borsutzky S, Cazac BB, Roes J, Guzman CA. 2004. TGF-β receptor signaling is critical for mucosal IgA responses. J. Immunol. 173:3305–9 182. van Ginkel FW, Wahl SM, Kearney JF, Kweon MN, Fujihashi K, et al. 1999. Partial IgAdeﬁciency with increased Th2-type cytokines in TGF-β1 knockout mice. J. Immunol. 163:1951–57 183. Letterio JJ, Geiser AG, Kulkarni AB, Roche NS, Sporn MB, Roberts AB. 1994. Maternal rescue of transforming growth factor-β1 null mice. Science 264:1936–38 184. Holder MJ, Knox K, Gordon J. 1992. Factors modifying survival pathways of germinal center B cells. Glucocorticoids and transforming growth factor-β, but not cyclosporin A or anti-CD19, block surface immunoglobulin-mediated rescue from apoptosis. Eur. J. Immunol. 22:2725–28 185. Lomo J, Blomhoff HK, Beiske K, Stokke T, Smeland EB. 1995. TGF-β1 and cyclic AMP promote apoptosis in resting human B lymphocytes. J. Immunol. 154:1634–43 186. Arsura M, Wu M, Sonenshein GE. 1996. TGF β1 inhibits NF-κB/Rel activity inducing apoptosis of B cells: transcriptional activation of IκBα. Immunity 5:31–40 187. Wildey GM, Patil S, Howe PH. 2003. Smad3 potentiates transforming growth factor β (TGFβ)-induced apoptosis and expression of the BH3-only protein Bim in WEHI 231 B lymphocytes. J. Biol. Chem. 278:18069–77 188. Hayakawa K, Asano M, Shinton SA, Gui M, Allman D, et al. 1999. Positive selection of natural autoreactive B cells. Science 285:113–16 189. Lam KP, Rajewsky K. 1999. B cell antigen receptor speciﬁcity and surface density together determine B-1 versus B-2 cell development. J. Exp. Med. 190:471–77 190. Snapper CM, Waegell W, Beernink H, Dasch JR. 1993. Transforming growth factor-β1 is required for secretion of IgG of all subclasses by LPS-activated murine B cells in vitro. J. Immunol. 151:4625–36 191. Zan H, Cerutti A, Dramitinos P, Schaffer A, Casali P. 1998. CD40 engagement triggers switching to IgA1 and IgA2 in human B cells through induction of endogenous TGF-β: evidence for TGF-β but not IL-10-dependent direct Sμ → Sα and sequential Sμ → Sγ, Sγ → Sα DNA recombination. J. Immunol. 161:5217–25 www.annualreviews.org • Regulation of Immune Responses by TGF-β

135

ARI

23 February 2006

15:9

192. Caver TE, O’Sullivan FX, Gold LI, Gresham HD. 1996. Intracellular demonstration of active TGFβ1 in B cells and plasma cells of autoimmune mice. IgG-bound TGFβ1 suppresses neutrophil function and host defense against Staphylococcus aureus infection. J. Clin. Invest. 98:2496–506 193. Biron CA, Nguyen KB, Pien GC, Cousens LP, Salazar-Mather TP. 1999. Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu. Rev. Immunol. 17:189–220 194. Hamerman JA, Ogasawara K, Lanier LL. 2005. NK cells in innate immunity. Curr. Opin. Immunol. 17:29–35 195. Martin-Fontecha A, Thomsen LL, Brett S, Gerard C, Lipp M, et al. 2004. Induced recruitment of NK cells to lymph nodes provides IFN-γ for TH 1 priming. Nat. Immunol. 5:1260–65 196. Perussia B, Chan SH, D’Andrea A, Tsuji K, Santoli D, et al. 1992. Natural killer (NK) cell stimulatory factor or IL-12 has differential effects on the proliferation of TCR-alpha beta+, TCR-gamma delta+ T lymphocytes, and NK cells. J. Immunol. 149:3495–502 197. Tripp CS, Wolf SF, Unanue ER. 1993. Interleukin 12 and tumor necrosis factor α are costimulators of interferon γ production by natural killer cells in severe combined immunodeﬁciency mice with listeriosis, and interleukin 10 is a physiologic antagonist. Proc. Natl. Acad. Sci. USA 90:3725–29 198. Rook AH, Kehrl JH, Wakeﬁeld LM, Roberts AB, Sporn MB, et al. 1986. Effects of transforming growth factor β on the functions of natural killer cells: depressed cytolytic activity and blunting of interferon responsiveness. J. Immunol. 136:3916–20 199. Bellone G, Aste-Amezaga M, Trinchieri G, Rodeck U. 1995. Regulation of NK cell functions by TGF-β 1. J. Immunol. 155:1066–73 200. Hunter CA, Bermudez L, Beernink H, Waegell W, Remington JS. 1995. Transforming growth factor-β inhibits interleukin-12-induced production of interferon-γ by natural killer cells: a role for transforming growth factor-β in the regulation of T cell-independent resistance to Toxoplasma gondii. Eur. J. Immunol. 25:994–1000 201. Su HC, Ishikawa R, Biron CA. 1993. Transforming growth factor-β expression and natural killer cell responses during virus infection of normal, nude, and SCID mice. J. Immunol. 151:4874–90 202. Ortaldo JR, Mason AT, O’Shea JJ, Smyth MJ, Falk LA, et al. 1991. Mechanistic studies of transforming growth factor-β inhibition of IL-2-dependent activation of CD3-large granular lymphocyte functions. Regulation of IL-2R β (p75) signal transduction. J. Immunol. 146:3791–98 203. Malygin AM, Meri S, Timonen T. 1993. Regulation of natural killer cell activity by transforming growth factor-β and prostaglandin E2. Scand. J. Immunol. 37:71–76 204. Moretta A, Biassoni R, Bottino C, Mingari MC, Moretta L. 2000. Natural cytotoxicity receptors that trigger human NK-cell-mediated cytolysis. Immunol. Today 21:228–34 205. Bauer S, Groh V, Wu J, Steinle A, Phillips JH, et al. 1999. Activation of NK cells and T cells by NKG2D, a receptor for stress-inducible MICA. Science 285:727–29 206. Castriconi R, Cantoni C, Della Chiesa M, Vitale M, Marcenaro E, et al. 2003. Transforming growth factor β1 inhibits expression of NKp30 and NKG2D receptors: consequences for the NK-mediated killing of dendritic cells. Proc. Natl. Acad. Sci. USA 100:4120– 25 207. Lee JC, Lee KM, Kim DW, Heo DS. 2004. Elevated TGF-β1 secretion and downmodulation of NKG2D underlies impaired NK cytotoxicity in cancer patients. J. Immunol. 172:7335–40

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

136

Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

208. Gray JD, Hirokawa M, Ohtsuka K, Horwitz DA. 1998. Generation of an inhibitory circuit involving CD8+ T cells, IL-2, and NK cell-derived TGF-β: contrasting effects of antiCD2 and anti-CD3. J. Immunol. 160:2248–54 209. Steinman RM, Hawiger D, Nussenzweig MC. 2003. Tolerogenic dendritic cells. Annu. Rev. Immunol. 21:685–711 210. Strobl H, Knapp W. 1999. TGF-β1 regulation of dendritic cells. Microbes Infect. 1:1283– 90 211. Strobl H, Riedl E, Scheinecker C, Bello-Fernandez C, Pickl WF, et al. 1996. TGF-β1 promotes in vitro development of dendritic cells from CD34+ hemopoietic progenitors. J. Immunol. 157:1499–507 212. Geissmann F, Prost C, Monnet JP, Dy M, Brousse N, Hermine O. 1998. Transforming growth factor β1, in the presence of granulocyte/macrophage colony-stimulating factor and interleukin 4, induces differentiation of human peripheral blood monocytes into dendritic Langerhans cells. J. Exp. Med. 187:961–66 213. Jaksits S, Kriehuber E, Charbonnier AS, Rappersberger K, Stingl G, Maurer D. 1999. CD34+ cell-derived CD14+ precursor cells develop into Langerhans cells in a TGF-β1dependent manner. J. Immunol. 163:4869–77 214. Zhang Y, Zhang YY, Ogata M, Chen P, Harada A, et al. 1999. Transforming growth factorβ1 polarizes murine hematopoietic progenitor cells to generate Langerhans cell-like dendritic cells through a monocyte/macrophage differentiation pathway. Blood 93:1208–20 215. Riedl E, Strobl H, Majdic O, Knapp W. 1997. TGF-β1 promotes in vitro generation of dendritic cells by protecting progenitor cells from apoptosis. J. Immunol. 158:1591–97 216. Borkowski TA, Letterio JJ, Farr AG, Udey MC. 1996. A role for endogenous transforming growth factor β1 in Langerhans cell biology: the skin of transforming growth factor β1 null mice is devoid of epidermal Langerhans cells. J. Exp. Med. 184:2417–22 217. Borkowski TA, Letterio JJ, Mackall CL, Saitoh A, Wang XJ, et al. 1997. A role for TGFβ1 in Langerhans cell biology. Further characterization of the epidermal Langerhans cell defect in TGFβ1 null mice. J. Clin. Invest. 100:575–81 218. Thomas RM, Belsito DV, Huang C, Chen LZ, Ormsby I, et al. 2001. Appearance of Langerhans cells in the epidermis of Tgfb1−/− SCID mice: paracrine and autocrine effects of transforming growth factor-β1 and -β2(1). J. Invest. Dermatol. 117:1574–80 219. Hacker C, Kirsch RD, Ju XS, Hieronymus T, Gust TC, et al. 2003. Transcriptional proﬁling identiﬁes Id2 function in dendritic cell development. Nat. Immunol. 4:380–86 220. Strobl H, Bello-Fernandez C, Riedl E, Pickl WF, Majdic O, et al. 1997. ﬂt3 ligand in cooperation with transforming growth factor-β1 potentiates in vitro development of Langerhans-type dendritic cells and allows single-cell dendritic cell cluster formation under serum-free conditions. Blood 90:1425–34 221. Geissmann F, Revy P, Regnault A, Lepelletier Y, Dy M, et al. 1999. TGF-β1 prevents the noncognate maturation of human dendritic Langerhans cells. J. Immunol. 162:4567–75 222. Ronger-Savle S, Valladeau J, Claudy A, Schmitt D, Peguet-Navarro J, et al. 2005. TGFβ inhibits CD1d expression on dendritic cells. J. Invest. Dermatol. 124:116–18 223. Yamaguchi Y, Tsumura H, Miwa M, Inaba K. 1997. Contrasting effects of TGF-β1 and TNF-α on the development of dendritic cells from progenitors in mouse bone marrow. Stem Cells 15:144–53 224. Dong Y, Tang L, Letterio JJ, Benveniste EN. 2001. The Smad3 protein is involved in TGF-β inhibition of class II transactivator and class II MHC expression. J. Immunol. 167:311–19 www.annualreviews.org • Regulation of Immune Responses by TGF-β

An in vivo demonstration of TGF-β1 in Langerhans cell development.

137

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

First demonstration of TGF-β induction of chemotaxis.

138

15:9

225. Tada Y, Asahina A, Fujita H, Mitsui H, Torii H, et al. 2004. Differential effects of LPS and TGF-β on the production of IL-6 and IL-12 by Langerhans cells, splenic dendritic cells, and macrophages. Cytokine 25:155–61 226. Gruschwitz MS, Hornstein OP. 1992. Expression of transforming growth factor type β on human epidermal dendritic cells. J. Invest. Dermatol. 99:114–16 227. de Saint-Vis B, Fugier-Vivier I, Massacrier C, Gaillard C, Vanbervliet B, et al. 1998. The cytokine proﬁle expressed by human dendritic cells is dependent on cell subtype and mode of activation. J. Immunol. 160:1666–76 228. Morelli AE, Zahorchak AF, Larregina AT, Colvin BL, Logar AJ, et al. 2001. Cytokine production by mouse myeloid dendritic cells in relation to differentiation and terminal maturation induced by lipopolysaccharide or CD40 ligation. Blood 98:1512–23 229. Steinman RM, Hawiger D, Liu K, Bonifaz L, Bonnyay D, et al. 2003. Dendritic cell function in vivo during the steady state: a role in peripheral tolerance. Ann. NY Acad. Sci. 987:15–25 230. Gregory CD, Devitt A. 2004. The macrophage and the apoptotic cell: an innate immune interaction viewed simplistically?Immunology 113:1–14 231. Ashcroft GS. 1999. Bidirectional regulation of macrophage function by TGF-β. Microbes Infect. 1:1275–82 232. Wahl SM, Hunt DA, Wakefield LM, McCartney-Francis N, Wahl LM, et al. 1987. Transforming growth factor type β induces monocyte chemotaxis and growth factor production. Proc. Natl. Acad. Sci. USA 84:5788–92 233. Wiseman DM, Polverini PJ, Kamp DW, Leibovich SJ. 1988. Transforming growth factorβ (TGF β) is chemotactic for human monocytes and induces their expression of angiogenic activity. Biochem. Biophys. Res. Commun. 157:793–800 234. Allen JB, Manthey CL, Hand AR, Ohura K, Ellingsworth L, Wahl SM. 1990. Rapid onset synovial inﬂammation and hyperplasia induced by transforming growth factor β. J. Exp. Med. 171:231–47 235. Wahl SM, Allen JB, Weeks BS, Wong HL, Klotman PE. 1993. Transforming growth factor β enhances integrin expression and type IV collagenase secretion in human monocytes. Proc. Natl. Acad. Sci. USA 90:4577–81 236. Bauvois B, Rouillard D, Sanceau J, Wietzerbin J. 1992. IFN-γ and transforming growth factor-β1 differently regulate ﬁbronectin and laminin receptors of human differentiating monocytic cells. J. Immunol. 148:3912–19 237. Turner M, Chantry D, Feldmann M. 1990. Transforming growth factor β induces the production of interleukin 6 by human peripheral blood mononuclear cells. Cytokine 2:211– 16 238. Riddick CA, Serio KJ, Hodulik CR, Ring WL, Regan MS, Bigby TD. 1999. TGF-β increases leukotriene C4 synthase expression in the monocyte-like cell line, THP-1. J. Immunol. 162:1101–7 239. Bottalico LA, Wager RE, Agellon LB, Assoian RK, Tabas I. 1991. Transforming growth factor-β1 inhibits scavenger receptor activity in THP-1 human macrophages. J. Biol. Chem. 266:22866–71 240. Han J, Hajjar DP, Tauras JM, Feng J, Gotto AM Jr, Nicholson AC. 2000. Transforming growth factor-β1 (TGF-β1) and TGF-β2 decrease expression of CD36, the type B scavenger receptor, through mitogen-activated protein kinase phosphorylation of peroxisome proliferator-activated receptor-γ. J. Biol. Chem. 275:1241–46 241. Tridandapani S, Wardrop R, Baran CP, Wang Y, Opalek JM, et al. 2003. TGF-β1 suppresses [correction of supresses] myeloid Fcγ receptor function by regulating the expression and function of the common γ-subunit. J. Immunol. 170:4572–77 Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

242. Bogdan C, Paik J, Vodovotz Y, Nathan C. 1992. Contrasting mechanisms for suppression of macrophage cytokine release by transforming growth factor-β and interleukin-10. J. Biol. Chem. 267:23301–8 243. Werner F, Jain MK, Feinberg MW, Sibinga NE, Pellacani A, et al. 2000. Transforming growth factor-β1 inhibition of macrophage activation is mediated via Smad3. J. Biol. Chem. 275:36653–58 244. Vodovotz Y, Bogdan C, Paik J, Xie QW, Nathan C. 1993. Mechanisms of suppression of macrophage nitric oxide release by transforming growth factor β. J. Exp. Med. 178:605–13 245. Tsunawaki S, Sporn M, Ding A, Nathan C. 1988. Deactivation of macrophages by transforming growth factor-β. Nature 334:260–62 246. Janeway CA Jr, Medzhitov R. 2002. Innate immune recognition. Annu. Rev. Immunol. 20:197–216 247. Imai K, Takeshita A, Hanazawa S. 2000. Transforming growth factor-β inhibits lipopolysaccharide-stimulated expression of inﬂammatory cytokines in mouse macrophages through downregulation of activation protein 1 and CD14 receptor expression. Infect. Immun. 68:2418–23 248. Naiki Y, Michelsen KS, Zhang W, Chen S, Doherty TM, Arditi M. 2005. Transforming growth factor-β differentially inhibits MyD88-dependent, but not TRAM- and TRIFdependent, lipopolysaccharide-induced TLR4 signaling. J. Biol. Chem. 280:5491–95 249. Nandan D, Reiner NE. 1997. TGF-β attenuates the class II transactivator and reveals an accessory pathway of IFN-γ action. J. Immunol. 158:1095–101 250. Takeuchi M, Alard P, Streilein JW. 1998. TGF-β promotes immune deviation by altering accessory signals of antigen-presenting cells. J. Immunol. 160:1589–97 251. Du C, Sriram S. 1998. Mechanism of inhibition of LPS-induced IL-12p40 production by IL-10 and TGF-β in ANA-1 cells. J. Leukoc. Biol. 64:92–97 252. Fadok VA, Bratton DL, Konowal A, Freed PW, Westcott JY, Henson PM. 1998. Macrophages that have ingested apoptotic cells in vitro inhibit proinﬂammatory cytokine production through autocrine/paracrine mechanisms involving TGF-β, PGE2, and PAF. J. Clin. Invest. 101:890–98 253. McDonald PP, Fadok VA, Bratton D, Henson PM. 1999. Transcriptional and translational regulation of inﬂammatory mediator production by endogenous TGF-β in macrophages that have ingested apoptotic cells. J. Immunol. 163:6164–72 254. Fadok VA, Voelker DR, Campbell PA, Cohen JJ, Bratton DL, Henson PM. 1992. Exposure of phosphatidylserine on the surface of apoptotic lymphocytes triggers speciﬁc recognition and removal by macrophages. J. Immunol. 148:2207–16 255. Fadok VA, Bratton DL, Rose DM, Pearson A, Ezekewitz RA, Henson PM. 2000. A receptor for phosphatidylserine-speciﬁc clearance of apoptotic cells. Nature 405:85–90 256. Huynh ML, Fadok VA, Henson PM. 2002. Phosphatidylserine-dependent ingestion of apoptotic cells promotes TGF-β1 secretion and the resolution of inﬂammation. J. Clin. Invest. 109:41–50 257. Otsuka M, Tsuchiya S, Aramaki Y. 2004. Involvement of ERK, a MAP kinase, in the production of TGF-β by macrophages treated with liposomes composed of phosphatidylserine. Biochem. Biophys. Res. Commun. 324:1400–5 258. Pilette C, Ouadrhiri Y, Van Snick J, Renauld JC, Staquet P, et al. 2002. IL-9 inhibits oxidative burst and TNF-α release in lipopolysaccharide-stimulated human monocytes through TGF-β. J. Immunol. 168:4103–11 259. Bastos KR, Alvarez JM, Marinho CR, Rizzo LV, Lima MR. 2002. Macrophages from IL-12p40-deﬁcient mice have a bias toward the M2 activation proﬁle. J. Leukoc. Biol. 71:271–78 www.annualreviews.org • Regulation of Immune Responses by TGF-β

Identified TGF-β as an inhibitor of macrophage activation.

139

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23 February 2006

15:9

260. Gruber BL, Marchese MJ, Kew RR. 1994. Transforming growth factor-β1 mediates mast cell chemotaxis. J. Immunol. 152:5860–67 261. Olsson N, Piek E, ten Dijke P, Nilsson G. 2000. Human mast cell migration in response to members of the transforming growth factor-β family. J. Leukoc. Biol. 67:350–56 262. Rosbottom A, Scudamore CL, von der Mark H, Thornton EM, Wright SH, Miller HR. 2002. TGF-β1 regulates adhesion of mucosal mast cell homologues to laminin-1 through expression of integrin α7 J. Immunol. 169:5689–95 263. Gomez G, Ramirez CD, Rivera J, Patel M, Norozian F, et al. 2005. TGF-β1 inhibits mast cell FcεRI expression. J. Immunol. 174:5987–93 264. Mesples B, Fontaine RH, Lelievre V, Launay JM, Gressens P. 2005. Neuronal TGFβ1 mediates IL-9/mast cell interaction and exacerbates excitotoxicity in newborn mice. Neurobiol. Dis. 18:193–205 265. Bissonnette EY, Enciso JA, Befus AD. 1997. TGF-β1 inhibits the release of histamine and tumor necrosis factor-α from mast cells through an autocrine pathway. Am. J. Respir. Cell Mol. Biol. 16:275–82 266. Gebhardt T, Lorentz A, Detmer F, Trautwein C, Bektas H, et al. 2005. Growth, phenotype, and function of human intestinal mast cells are tightly regulated by transforming growth factor β1. Gut 54:928–34 267. Broide DH, Wasserman SI, Alvaro-Gracia J, Zvaiﬂer NJ, Firestein GS. 1989. Transforming growth factor-β1 selectively inhibits IL-3-dependent mast cell proliferation without affecting mast cell function or differentiation. J. Immunol. 143:1591–97 268. Miller HR, Wright SH, Knight PA, Thornton EM. 1999. A novel function for transforming growth factor-β1: upregulation of the expression and the IgE-independent extracellular release of a mucosal mast cell granule-speciﬁc β-chymase, mouse mast cell protease-1. Blood 93:3473–86 269. Pennington DW, Lopez AR, Thomas PS, Peck C, Gold WM. 1992. Dog mastocytoma cells produce transforming growth factor β1. J. Clin. Invest. 90:35–41 270. Gordon JR, Galli SJ. 1994. Promotion of mouse ﬁbroblast collagen gene expression by mast cells stimulated via the FcεRI. Role for mast cell-derived transforming growth factor β and tumor necrosis factor α. J. Exp. Med. 180:2027–37 271. Lindstedt KA, Wang Y, Shiota N, Saarinen J, Hyytiainen M, et al. 2001. Activation of paracrine TGF-β1 signaling upon stimulation and degranulation of rat serosal mast cells: a novel function for chymase. FASEB J. 15:1377–88 272. Kobayashi SD, Voyich JM, DeLeo FR. 2003. Regulation of the neutrophil-mediated inﬂammatory response to infection. Microbes Infect. 5:1337–44 273. Klion AD, Nutman TB. 2004. The role of eosinophils in host defense against helminth parasites. J. Allergy Clin. Immunol. 113:30–37 274. Munitz A, Levi-Schaffer F. 2004. Eosinophils: ‘new’ roles for ‘old’ cells. Allergy 59:268–75 275. Reibman J, Meixler S, Lee TC, Gold LI, Cronstein BN, et al. 1991. Transforming growth factor β1, a potent chemoattractant for human neutrophils, bypasses classic signaltransduction pathways. Proc. Natl. Acad. Sci. USA 88:6805–9 276. Brandes ME, Mai UE, Ohura K, Wahl SM. 1991. Type I transforming growth factor-β receptors on neutrophils mediate chemotaxis to transforming growth factor-β. J. Immunol. 147:1600–6 277. Chen CC, Manning AM. 1996. TGF-β1, IL-10 and IL-4 differentially modulate the cytokine-induced expression of IL-6 and IL-8 in human endothelial cells. Cytokine 8:58– 65

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

140

Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

278. Park SK, Yang WS, Lee SK, Ahn H, Park JS, et al. 2000. TGF-β1 down-regulates inﬂammatory cytokine-induced VCAM-1 expression in cultured human glomerular endothelial cells. Nephrol. Dial. Transplant. 15:596–604 279. Smith WB, Noack L, Khew-Goodall Y, Isenmann S, Vadas MA, Gamble JR. 1996. Transforming growth factor-β1 inhibits the production of IL-8 and the transmigration of neutrophils through activated endothelium. J. Immunol. 157:360–68 280. Luttmann W, Franz P, Matthys H, Virchow JC Jr. 1998. Effects of TGF-β on eosinophil chemotaxis. Scand. J. Immunol. 47:127–30 281. Lagraoui M, Gagnon L. 1997. Enhancement of human neutrophil survival and activation by TGF-β1. Cell. Mol. Biol. 43:313–18 282. Balazovich KJ, Fernandez R, Hinkovska-Galcheva V, Suchard SJ, Boxer LA. 1996. Transforming growth factor-β1 stimulates degranulation and oxidant release by adherent human neutrophils. J. Leukoc. Biol. 60:772–77 283. Myrtek D, Knoll M, Matthiesen T, Krause S, Lohrmann J, et al. 2004. Expression of interleukin-13 receptor alpha 1-subunit on peripheral blood eosinophils is regulated by cytokines. Immunology 112:597–604 284. Alam R, Forsythe P, Stafford S, Fukuda Y. 1994. Transforming growth factor β abrogates the effects of hematopoietins on eosinophils and induces their apoptosis. J. Exp. Med. 179:1041–45 285. Fava RA, Olsen NJ, Postlethwaite AE, Broadley KN, Davidson JM, et al. 1991. Transforming growth factor β1 (TGF-β1) induced neutrophil recruitment to synovial tissues: implications for TGF-β-driven synovial inﬂammation and hyperplasia. J. Exp. Med. 173:1121–32 286. Kobayashi T, Iijima K, Kita H. 2003. Marked airway eosinophilia prevents development of airway hyper-responsiveness during an allergic response in IL-5 transgenic mice. J. Immunol. 170:5756–63 287. Tanaka H, Komai M, Nagao K, Ishizaki M, Kajiwara D, et al. 2004. Role of interleukin-5 and eosinophils in allergen-induced airway remodeling in mice. Am. J. Respir. Cell Mol. Biol. 31:62–68 288. Cho JY, Miller M, Baek KJ, Han JW, Nayar J, et al. 2004. Inhibition of airway remodeling in IL-5-deﬁcient mice. J. Clin. Invest. 113:551–60 289. Prud’homme GJ, Piccirillo CA. 2000. The inhibitory effects of transforming growth factor-β-1 (TGF-β1) in autoimmune diseases. J. Autoimmun. 14:23–42 290. Dang H, Geiser AG, Letterio JJ, Nakabayashi T, Kong L, et al. 1995. SLE-like autoantibodies and Sjogren’s syndrome-like lymphoproliferation in TGF-β knockout mice. J. Immunol. 155:3205–12 291. Yaswen L, Kulkarni AB, Fredrickson T, Mittleman B, Schiffman R, et al. 1996. Autoimmune manifestations in the transforming growth factor-β1 knockout mouse. Blood 87:1439–45 292. Ohtsuka K, Gray JD, Stimmler MM, Toro B, Horwitz DA. 1998. Decreased production of TGF-β by lymphocytes from patients with systemic lupus erythematosus. J. Immunol. 160:2539–45 293. Ohtsuka K, Gray JD, Stimmler MM, Horwitz DA. 1999. The relationship between defects in lymphocyte production of transforming growth factor-β1 in systemic lupus erythematosus and disease activity or severity. Lupus 8:90–94 294. Lowrance JH, O’Sullivan FX, Caver TE, Waegell W, Gresham HD. 1994. Spontaneous elaboration of transforming growth factor β suppresses host defense against bacterial infection in autoimmune MRL/lpr mice. J. Exp. Med. 180:1693–703 www.annualreviews.org • Regulation of Immune Responses by TGF-β

141

ARI

23 February 2006

15:9

295. Raz E, Dudler J, Lotz M, Baird SM, Berry CC, et al. 1995. Modulation of disease activity in murine systemic lupus erythematosus by cytokine gene delivery. Lupus 4:286–92 296. Holmdahl R, Bockermann R, Backlund J, Yamada H. 2002. The molecular pathogenesis of collagen-induced arthritis in mice—a model for rheumatoid arthritis. Ageing Res. Rev. 1:135–47 297. Thorbecke GJ, Shah R, Leu CH, Kuruvilla AP, Hardison AM, Palladino MA. 1992. Involvement of endogenous tumor necrosis factor α and transforming growth factor β during induction of collagen type II arthritis in mice. Proc. Natl. Acad. Sci. USA 89:7375– 79 298. Wahl SM, Allen JB, Costa GL, Wong HL, Dasch JR. 1993. Reversal of acute and chronic synovial inﬂammation by anti-transforming growth factor β. J. Exp. Med. 177:225–30 299. Schramm C, Kriegsmann J, Protschka M, Huber S, Hansen T, et al. 2004. Susceptibility to collagen-induced arthritis is modulated by TGFβ responsiveness of T cells. Arthritis Res. Ther. 6:R114–19 300. King C, Davies J, Mueller R, Lee MS, Krahl T, et al. 1998. TGF-β1 alters APC preference, polarizing islet antigen responses toward a Th2 phenotype. Immunity 8:601–13 301. Grewal IS, Grewal KD, Wong FS, Wang H, Picarella DE, et al. 2002. Expression of transgene encoded TGF-β in islets prevents autoimmune diabetes in NOD mice by a local mechanism. J. Autoimmun. 19:9–22 302. Moritani M, Yoshimoto K, Wong SF, Tanaka C, Yamaoka T, et al. 1998. Abrogation of autoimmune diabetes in nonobese diabetic mice and protection against effector lymphocytes by transgenic paracrine TGF-β1. J. Clin. Invest. 102:499–506 303. Belghith M, Bluestone JA, Barriot S, Megret J, Bach JF, Chatenoud L. 2003. TGF-βdependent mechanisms mediate restoration of self-tolerance induced by antibodies to CD3 in overt autoimmune diabetes. Nat. Med. 9:1202–8 304. Herold KC, Hagopian W, Auger JA, Poumian-Ruiz E, Taylor L, et al. 2002. Anti-CD3 monoclonal antibody in new-onset type 1 diabetes mellitus. N. Engl. J. Med. 346:1692–98 305. Johns LD, Flanders KC, Ranges GE, Sriram S. 1991. Successful treatment of experimental allergic encephalomyelitis with transforming growth factor-β1. J. Immunol. 147:1792–96 306. Racke MK, Dhib-Jalbut S, Cannella B, Albert PS, Raine CS, McFarlin DE. 1991. Prevention and treatment of chronic relapsing experimental allergic encephalomyelitis by transforming growth factor-β1. J. Immunol. 146:3012–17 307. Kuruvilla AP, Shah R, Hochwald GM, Liggitt HD, Palladino MA, Thorbecke GJ. 1991. Protective effect of transforming growth factor β1 on experimental autoimmune diseases in mice. Proc. Natl. Acad. Sci. USA 88:2918–21 308. Racke MK, Cannella B, Albert P, Sporn M, Raine CS, McFarlin DE. 1992. Evidence of endogenous regulatory function of transforming growth factor-β1 in experimental allergic encephalomyelitis. Int. Immunol. 4:615–20 309. Issazadeh S, Mustafa M, Ljungdahl A, Hojeberg B, Dagerlind A, et al. 1995. Interferon γ, interleukin 4 and transforming growth factor β in experimental autoimmune encephalomyelitis in Lewis rats: dynamics of cellular mRNA expression in the central nervous system and lymphoid cells. J. Neurosci. Res. 40:579–90 310. Johns LD, Sriram S. 1993. Experimental allergic encephalomyelitis: neutralizing antibody to TGF β1 enhances the clinical severity of the disease. J. Neuroimmunol. 47:1–7 311. Bai XF, Shi FD, Xiao BG, Li HL, van der Meide PH, Link H. 1997. Nasal administration of myelin basic protein prevents relapsing experimental autoimmune encephalomyelitis in DA rats by activating regulatory cells expressing IL-4 and TGF-β mRNA. J. Neuroimmunol. 80:65–75

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

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Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

312. Nowell PC. 1976. The clonal evolution of tumor cell populations. Science 194:23–28 313. Foulds L. 1954. The experimental study of tumor progression: a review. Cancer Res. 14:327–39 314. Kinzler KW, Vogelstein B. 1996. Lessons from hereditary colorectal cancer. Cell 87:159– 70 315. Weinberg RA. 1996. How cancer arises. Sci. Am. 275:62–70 316. Pardoll D. 2003. Does the immune system see tumors as foreign or self?Annu. Rev. Immunol. 21:807–39 317. Dunn GP, Old LJ, Schreiber RD. 2004. The three Es of cancer immunoediting. Annu. Rev. Immunol. 22:329–60 318. Chang HL, Gillett N, Figari I, Lopez AR, Palladino MA, Derynck R. 1993. Increased transforming growth factor β expression inhibits cell proliferation in vitro, yet increases tumorigenicity and tumor growth of Meth A sarcoma cells. Cancer Res. 53:4391–98 319. Stander M, Naumann U, Dumitrescu L, Heneka M, Loschmann P, et al. 1998. Decorin gene transfer-mediated suppression of TGF-β synthesis abrogates experimental malignant glioma growth in vivo. Gene Ther. 5:1187–94 320. Fakhrai H, Dorigo O, Shawler DL, Lin H, Mercola D, et al. 1996. Eradication of established intracranial rat gliomas by transforming growth factor β antisense gene therapy. Proc. Natl. Acad. Sci. USA 93:2909–14 321. Friese MA, Wischhusen J, Wick W, Weiler M, Eisele G, et al. 2004. RNA interference targeting transforming growth factor-β enhances NKG2D-mediated antiglioma immune response, inhibits glioma cell migration and invasiveness, and abrogates tumorigenicity in vivo. Cancer Res. 64:7596–603 322. Torre-Amione G, Beauchamp RD, Koeppen H, Park BH, et al. 1990. A highly immunogenic tumor transfected with a murine transforming growth factor type β1 cDNA escapes immune surveillance. Proc. Natl. Acad. Sci. USA 87:1486–90 323. Nishikawa H, Kato T, Tawara I, Takemitsu T, Saito K, et al. 2005. Accelerated chemically induced tumor development mediated by CD4+ CD25+ regulatory T cells in wild-type hosts. Proc. Natl. Acad. Sci. USA 102:9253–57 324. Shimizu J, Yamazaki S, Sakaguchi S. 1999. Induction of tumor immunity by removing CD25+ CD4+ T cells: a common basis between tumor immunity and autoimmunity. J. Immunol. 163:5211–18 325. Onizuka S, Tawara I, Shimizu J, Sakaguchi S, Fujita T, Nakayama E. 1999. Tumor rejection by in vivo administration of anti-CD25 (interleukin-2 receptor α) monoclonal antibody. Cancer Res. 59:3128–33 326. Sutmuller RP, van Duivenvoorde LM, van Elsas A, Schumacher TN, Wildenberg ME, et al. 2001. Synergism of cytotoxic T lymphocyte-associated antigen 4 blockade and depletion of CD25+ regulatory T cells in antitumor therapy reveals alternative pathways for suppression of autoreactive cytotoxic T lymphocyte responses. J. Exp. Med. 194:823–32 327. Chen ML, Pittet MJ, Gorelik L, Flavell RA, Weissleder R, et al. 2005. Regulatory T cells suppress tumor-speciﬁc CD8 T cell cytotoxicity through TGF-β signals in vivo. Proc. Natl. Acad. Sci. USA 102:419–24 328. Ormandy LA, Hillemann T, Wedemeyer H, Manns MP, Greten TF, Korangy F. 2005. Increased populations of regulatory T cells in peripheral blood of patients with hepatocellular carcinoma. Cancer Res. 65:2457–64 329. Beyer M, Kochanek M, Darabi K, Popov A, Jensen M, et al. 2005. Reduced frequencies and suppressive function of CD4+ CD25high regulatory T cells in patients with chronic lymphocytic leukemia after therapy with ﬂudarabine. Blood 106:2018–25 www.annualreviews.org • Regulation of Immune Responses by TGF-β

143

ARI

23 February 2006

15:9

330. Hansson GK. 2005. Inﬂammation, atherosclerosis, and coronary artery disease. N. Engl. J. Med. 352:1685–95 331. Binder CJ, Chang MK, Shaw PX, Miller YI, Hartvigsen K, et al. 2002. Innate and acquired immunity in atherogenesis. Nat. Med. 8:1218–26 332. Libby P. 2002. Inﬂammation in atherosclerosis. Nature 420:868–74 333. Mallat Z, Gojova A, Marchiol-Fournigault C, Esposito B, Kamate C, et al. 2001. Inhibition of transforming growth factor-β signaling accelerates atherosclerosis and induces an unstable plaque phenotype in mice. Circ. Res. 89:930–34 334. Lutgens E, Gijbels M, Smook M, Heeringa P, Gotwals P, et al. 2002. Transforming growth factor-β mediates balance between inﬂammation and ﬁbrosis during plaque progression. Arterioscler. Thromb. Vasc. Biol. 22:975–82 335. Robertson AK, Rudling M, Zhou X, Gorelik L, Flavell RA, Hansson GK. 2003. Disruption of TGF-β signaling in T cells accelerates atherosclerosis. J. Clin. Invest. 112:1342–50 336. Gojova A, Brun V, Esposito B, Cottrez F, Gourdy P, et al. 2003. Speciﬁc abrogation of transforming growth factor-β signaling in T cells alters atherosclerotic lesion size and composition in mice. Blood 102:4052–58 337. Argmann CA, Van Den Diepstraten CH, Sawyez CG, Edwards JY, Hegele RA, et al. 2001. Transforming growth factor-β1 inhibits macrophage cholesteryl ester accumulation induced by native and oxidized VLDL remnants. Arterioscler. Thromb. Vasc. Biol. 21:2011– 18 338. Amento EP, Ehsani N, Palmer H, Libby P. 1991. Cytokines and growth factors positively and negatively regulate interstitial collagen gene expression in human vascular smooth muscle cells. Arterioscler. Thromb. 11:1223–30 339. Fabunmi RP, Baker AH, Murray EJ, Booth RF, Newby AC. 1996. Divergent regulation by growth factors and cytokines of 95 kDa and 72 kDa gelatinases and tissue inhibitors or metalloproteinases-1, -2, and -3 in rabbit aortic smooth muscle cells. Biochem. J. 315(Pt. 1):335–42 340. Cipollone F, Fazia M, Mincione G, Iezzi A, Pini B, et al. 2004. Increased expression of transforming growth factor-β1 as a stabilizing factor in human atherosclerotic plaques. Stroke 35:2253–57 341. Grainger DJ, Kemp PR, Metcalfe JC, Liu AC, Lawn RM, et al. 1995. The serum concentration of active transforming growth factor-β is severely depressed in advanced atherosclerosis. Nat. Med. 1:74–79 342. Hansen G, McIntire JJ, Yeung VP, Berry G, Thorbecke GJ, et al. 2000. CD4+ T helper cells engineered to produce latent TGF-β1 reverse allergen-induced airway hyperreactivity and inﬂammation. J. Clin. Invest. 105:61–70 343. Scherf W, Burdach S, Hansen G. 2005. Reduced expression of transforming growth factor β1 exacerbates pathology in an experimental asthma model. Eur. J. Immunol. 35:198–206 344. Redington AE, Madden J, Frew AJ, Djukanovic R, Roche WR, et al. 1997. Transforming growth factor-β1 in asthma. Measurement in bronchoalveolar lavage ﬂuid. Am. J. Respir. Crit. Care Med. 156:642–47 345. Branton MH, Kopp JB. 1999. TGF-β and ﬁbrosis. Microbes Infect. 1:1349–65 346. Bouma G, Strober W. 2003. The immunological and genetic basis of inﬂammatory bowel disease. Nat. Rev. Immunol. 3:521–33 347. Monteleone G, Pallone F, MacDonald TT. 2004. Smad7 in TGF-β-mediated negative regulation of gut inﬂammation. Trends Immunol. 25:513–17 348. Monteleone G, Kumberova A, Croft NM, McKenzie C, Steer HW, MacDonald TT. 2001. Blocking Smad7 restores TGF-β1 signaling in chronic inﬂammatory bowel disease. J. Clin. Invest. 108:601–9

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

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Li et al.

Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-04

ARI

23 February 2006

15:9

349. Ming M, Ewen ME, Pereira ME. 1995. Trypanosome invasion of mammalian cells requires activation of the TGF β signaling pathway. Cell 82:287–96 350. Silva JS, Twardzik DR, Reed SG. 1991. Regulation of Trypanosoma cruzi infections in vitro and in vivo by transforming growth factor β (TGF-β). J. Exp. Med. 174:539–45 351. Hansen DS, Villacres-Eriksson M, Akerblom L, Hellman U, Segura E, et al. 1998. An immunoafﬁnity-puriﬁed Trypanosoma cruzi antigen suppresses cellular proliferation through a TGF-β-mediated mechanism. Scand. J. Immunol. 47:509–16 352. Olsson T, Bakhiet M, Hojeberg B, Ljungdahl A, Edlund C, et al. 1993. CD8 is critically involved in lymphocyte activation by a T. brucei brucei-released molecule. Cell 72:715–27 353. Sacks D, Noben-Trauth N. 2002. The immunology of susceptibility and resistance to Leishmania major in mice. Nat. Rev. Immunol. 2:845–58 354. Scott P. 1991. IFN-γ modulates the early development of Th1 and Th2 responses in a murine model of cutaneous leishmaniasis. J. Immunol. 147:3149–55 355. Belosevic M, Finbloom DS, Van Der Meide PH, Slayter MV, Nacy CA. 1989. Administration of monoclonal anti-IFN-γ antibodies in vivo abrogates natural resistance of C3H/HeN mice to infection with Leishmania major. J. Immunol. 143:266–74 356. Barral A, Barral-Netto M, Yong EC, Brownell CE, Twardzik DR, Reed SG. 1993. Transforming growth factor β as a virulence mechanism for Leishmania braziliensis. Proc. Natl. Acad. Sci. USA 90:3442–46 357. Gantt KR, Schultz-Cherry S, Rodriguez N, Jeronimo SM, Nascimento ET, et al. 2003. Activation of TGF-β by Leishmania chagasi: importance for parasite survival in macrophages. J. Immunol. 170:2613–20 358. Langermans JA, Nibbering PH, Van Vuren-Van Der Hulst ME, Van Furth R. 2001. Transforming growth factor-β suppresses interferon-γ-induced toxoplasmastatic activity in murine macrophages by inhibition of tumour necrosis factor-α production. Parasite Immunol. 23:169–75 359. Bermudez LE, Covaro G, Remington J. 1993. Infection of murine macrophages with Toxoplasma gondii is associated with release of transforming growth factor β and downregulation of expression of tumor necrosis factor receptors. Infect. Immun. 61:4126–30 360. Omer FM, Kurtzhals JA, Riley EM. 2000. Maintaining the immunological balance in parasitic infections: a role for TGF-β? Parasitol. Today 16:18–23 361. Omer FM, Riley EM. 1998. Transforming growth factor β production is inversely correlated with severity of murine malaria infection. J. Exp. Med. 188:39–48 362. Li C, Corraliza I, Langhorne J. 1999. A defect in interleukin-10 leads to enhanced malarial disease in Plasmodium chabaudi chabaudi infection in mice. Infect. Immun. 67:4435–42 363. Li C, Sanni LA, Omer F, Riley E, Langhorne J. 2003. Pathology of Plasmodium chabaudi chabaudi infection and mortality in interleukin-10-deﬁcient mice are ameliorated by antitumor necrosis factor α and exacerbated by anti-transforming growth factor β antibodies. Infect. Immun. 71:4850–56 364. Omer FM, de Souza JB, Corran PH, Sultan AA, Riley EM. 2003. Activation of transforming growth factor β by malaria parasite-derived metalloproteinases and a thrombospondin-like molecule. J. Exp. Med. 198:1817–27 365. Toossi Z, Young TG, Averill LE, Hamilton BD, Shiratsuchi H, Ellner JJ. 1995. Induction of transforming growth factor β1 by puriﬁed protein derivative of Mycobacterium tuberculosis. Infect. Immun. 63:224–28 366. Dahl KE, Shiratsuchi H, Hamilton BD, Ellner JJ, Toossi Z. 1996. Selective induction of transforming growth factor β in human monocytes by lipoarabinomannan of Mycobacterium tuberculosis. Infect. Immun. 64:399–405 www.annualreviews.org • Regulation of Immune Responses by TGF-β

145

ARI

23 February 2006

15:9

367. Pamer EG. 2004. Immune responses to Listeria monocytogenes. Nat. Rev. Immunol. 4:812– 23 368. Nakane A, Asano M, Sasaki S, Nishikawa S, Miura T, et al. 1996. Transforming growth factor β is protective in host resistance against Listeria monocytogenes infection in mice. Infect. Immun. 64:3901–4 369. Spaccapelo R, Romani L, Tonnetti L, Cenci E, Mencacci A, et al. 1995. TGF-β is important in determining the in vivo patterns of susceptibility or resistance in mice infected with Candida albicans. J. Immunol. 155:1349–60 370. Letterio JJ, Lehrnbecher T, Pollack G, Walsh TJ, Chanock SJ. 2001. Invasive candidiasis stimulates hepatocyte and monocyte production of active transforming growth factor β. Infect. Immun. 69:5115–20 371. Schultz-Cherry S, Hinshaw VS. 1996. Inﬂuenza virus neuraminidase activates latent transforming growth factor β. J. Virol. 70:8624–29 372. Cecere A, Marotta F, Vangieri B, Tancredi L, Gattoni A. 2004. Progressive liver injury in chronic hepatitis C infection is related to altered cellular immune response and to different citokine proﬁle. Panminerva Med. 46:171–87 373. Taniguchi H, Kato N, Otsuka M, Goto T, Yoshida H, et al. 2004. Hepatitis C virus core protein upregulates transforming growth factor-β1 transcription. J. Med. Virol. 72:52–59 374. Cheng PL, Chang MH, Chao CH, Lee YH. 2004. Hepatitis C viral proteins interact with Smad3 and differentially regulate TGF-β/Smad3-mediated transcriptional activation. Oncogene 23:7821–38 375. Reinhold D, Wrenger S, Kahne T, Ansorge S. 1999. HIV-1 Tat: immunosuppression via TGF-β1 induction. Immunol. Today 20:384–85 376. Chen S, Tuttle DL, Oshier JT, Knot HJ, Streit WJ, et al. 2005. Transforming growth factor-β1 increases CXCR4 expression, stromal-derived factor-1α-stimulated signalling and human immunodeﬁciency virus-1 entry in human monocyte-derived macrophages. Immunology 114:565–74 377. Barrios-Rodiles M, Brown KR, Ozdamar B, Bose R, Liu Z, et al. 2005. High-throughput mapping of a dynamic signaling network in mammalian cells. Science 307:S1621–25

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Contents

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:99-146. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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The Eosinophil Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Marc E. Rothenberg and Simon P. Hogan Division of Allergy and Immunology, Department of Pediatrics, Cincinnati Children’s Hospital Medical Center, University of Cincinnati College of Medicine, Cincinnati, Ohio 45229; email: [email protected], [email protected]

Annu. Rev. Immunol. 2006. 24:147–74 First published online as a Review in Advance on November 4, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090720 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0147$20.00

Key Words allergy, asthma, cellular trafﬁcking, chemokines, mucosal immunity

Abstract Eosinophils have been considered end-stage cells involved in host protection against parasites. However, numerous lines of evidence have now changed this perspective by showing that eosinophils are pleiotropic multifunctional leukocytes involved in initiation and propagation of diverse inﬂammatory responses, as well as modulators of innate and adaptive immunity. In this review, we summarize the biology of eosinophils, focusing on the growing properties of eosinophil-derived products, including the constituents of their granules as well as the mechanisms by which they release their pleiotropic mediators. We examine new views on the role of eosinophils in homeostatic function, including developmental biology and innate and adaptive immunity (as well as interaction with mast cells and T cells). The molecular steps involved in eosinophil development and trafﬁcking are described, with special attention to the important role of the transcription factor GATA-1, the eosinophil-selective cytokine IL-5, and the eotaxin subfamily of chemokines. We also review the role of eosinophils in disease processes, including infections, asthma, and gastrointestinal disorders, and new data concerning genetically engineered eosinophil-deﬁcient mice. Finally, strategies for targeted therapeutic intervention in eosinophil-mediated mucosal diseases are conceptualized.

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INTRODUCTION

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Eotaxin family of chemokines: the group of related eosinophil-selective chemoattractant proteins eotaxin-1, eotaxin-2, and eotaxin-3 Major basic protein (MBP): a major protein in eosinophil granules EPO: eosinophil peroxidase

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Eosinophils are multifunctional leukocytes implicated in the pathogenesis of numerous inﬂammatory processes, including parasitic helminth infections and allergic diseases (1– 3). In response to diverse stimuli, eosinophils are recruited from the circulation into inﬂammatory foci, where they modulate immune responses through an array of mechanisms. Triggering of eosinophils by engagement of receptors for cytokines, immunoglobulins, and complement can lead to the secretion of an array of proinﬂammatory cytokines [IL-2, IL-4, IL-5, IL-10, IL-12, IL-13, IL-16, IL18, and TGF (transforming growth factor)α/β], chemokines (RANTES and eotaxin1), and lipid mediators [platelet-activating factor and leukotriene C4 (LTC4)] (4) (Figure 1). These molecules have proinﬂammatory effects, including upregulation of adhesion systems, modulation of cellular trafﬁcking, and activation and regulation of vascular permeability, mucus secretion, and smooth muscle constriction. Eosinophils can initiate antigen-speciﬁc immune responses by acting as antigen-presenting cells (APCs). Furthermore, eosinophils can serve as major effector cells inducing tissue damage and dysfunction by releasing toxic granule proteins and lipid mediators (5). In this review, we summarize the biology of eosinophils, focusing on the growing properties of eosinophil-derived products, including the constituents of their granules as well as the mechanisms by which they release their pleiotropic mediators. We examine new views on the role of eosinophils in homeostatic function, including developmental biology and innate and adaptive immunity (including interaction with mast cells and T cells). The molecular steps involved in eosinophil development and trafﬁcking are described, with special attention to the important role of the transcription factor GATA1 and the eosinophil-selective cytokine IL-5 and the eotaxin subfamily of chemokines. Furthermore, we review the role of eosinophils

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in disease processes, including infections, asthma, and gastrointestinal disorders. We also review new data concerning genetically engineered eosinophil-deﬁcient mice. Finally, strategies for targeted therapeutic intervention in eosinophil-mediated diseases are conceptualized.

EOSINOPHIL GRANULE PROTEINS Eosinophils secrete an array of cytotoxic granule cationic proteins [major basic protein (MBP), eosinophil cationic protein (ECP), eosinophil peroxidase (EPO), and eosinophilderived neurotoxin (EDN)] that are capable of inducing tissue damage and dysfunction (5). Eosinophil granules contain a crystalloid core composed of MBP-1 (and MBP2) and a matrix composed of ECP, EDN, and EPO (5). MBP, EPO, and ECP are toxic to a variety of tissues, including heart, brain, and bronchial epithelium (6–9). ECP and EDN are ribonucleases and have been shown to possess antiviral activity, and ECP causes voltage-insensitive, ion-selective toxic pores in the membranes of target cells, possibly facilitating the entry of other cytotoxic molecules (10–13). ECP also has a number of additional noncytotoxic activities, including suppression of T cell proliferative responses and immunoglobulin synthesis by B cells, induction of mast cell degranulation, and stimulation of airway mucus secretion and glycosaminoglycan production by human ﬁbroblasts (14). MBP directly alters smooth muscle contraction responses by dysregulating vagal muscarinic M2 and M3 receptor function and by inducing mast cell and basophil degranulation (15–17). MBP has recently been implicated in regulating peripheral nerve plasticity (18). EPO, which constitutes ∼25% of the total protein mass of speciﬁc granules, catalyzes the oxidation of pseudohalides [thyiocyanate (SCN− )], halides [chloride (Cl− ), bromide (Br− ), iodide (I− )], and nitric oxide (nitrite) to form highly reactive oxygen species

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Figure 1 Schematic diagram of an eosinophil and its multifunctional effects. Eosinophils are bilobed granulocytes with eosinophilic staining secondary granules. The secondary granules contain four primary cationic proteins, designated eosinophil peroxidase (EPO), major basic protein (MBP), eosinophil cationic protein (ECP), and eosinophil-derived neurotoxin (EDN). All four proteins are cytotoxic molecules; in addition, ECP and EDN are ribonucleases. Eosinophils respond to diverse stimuli, including nonspeciﬁc tissue injury, infections, allografts, allergens, and tumors. In addition to releasing their preformed cationic proteins, eosinophils can also release a variety of cytokines, chemokines, lipid mediators, and neuromodulators. Eosinophils directly communicate with T cells and mast cells in a bidirectional manner. Eosinophils activate T cells by serving as APCs, and eosinophil-derived MBP is a mast cell secretagogue. Eosinophils can also regulate T cell polarization through synthesis of indoleamine 2,3-dioxygenase (IDO), an enzyme involved in oxidative metabolism of tryptophan, catalyzing the conversion of tryptophan to kynurenines (KYN), a regulator of Th1/Th2 balance.

(hypohalous acids) and reactive nitrogen metabolites (perioxynitrate). These molecules oxidize nucleophilic targets on proteins, promoting oxidative stress and subsequent cell death by apoptosis and necrosis (19–21). Eosinophils predominantly secrete their granule protein by regulated exocytosis and degranulation (22). In a process of piecemeal degranulation, eosinophils selectively release components of their speciﬁc granules (23). For example, activation of human eosinophils by IFN-γ promotes the mobilization of granule-derived RANTES to the cell periphery without inducing cationic protein release (24, 25). Regulated exocytosis occurs by the formation of a docking complex com-

posed of soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptors (SNAREs) located on the vesicle (v-SNAREs) and the target membrane (t-SNAREs). SNAREs are classiﬁed into two categories based on the presence of a conserved amino acid (arginine [R] or glutamine [Q]). Human eosinophils express the Q-SNAREs SNAP-23 and syntaxin-4, which are predominantly localized to the plasma membrane (26), and the R-SNARE VAMP (vesicle-associated membrane protein)-2, which is localized to cytoplasmic secretory vesicles. It is postulated that receptor-coupled activation of eosinophils leads to rapid mobilization of cytoplasmic vesicles to the plasma membrane, leading to www.annualreviews.org • The Eosinophil

SNARE: soluble N-ethylmaleimidesensitive factor attachment protein (SNAP) receptor

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the formation of a SNARE complex (VAMP2/SNAP-23/syntaxin-4) and subsequent mediator release (22).

EOSINOPHILS AND HOMEOSTATIC FUNCTION

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Early clinical investigations have demonstrated an association between eosinophils and parasitic infections, leading investigators to hypothesize that eosinophils were classical end-stage effector cells involved in host defense (27). However, in recent years, eosinophils have been shown to be involved in numerous biological processes, including postpubertal mammary gland development (28), estrus cycling (29, 30), organ transplantation (31), viral infection (13), allergic inﬂammatory responses, and neoplasia (32).

Eosinophils and Reproduction Eosinophils are a prevalent cell population in the female reproductive tract, with numbers reaching maximum levels at estrus. Eosinophils are predominantly localized to the endometrial stroma subadjacent to the luminal and glandular epithelium and at the endometrial-myometrial junction (28). Eosinophil recruitment into the uterus is regulated by IL-5; however, while uterine eosinophil numbers are depleted in IL5-deﬁcient mice, a residual population of eosinophils is still present, and their localization in the subepithelial stroma is comparable to wild-type mice, suggesting that IL-5independent mechanisms regulate the tissuespeciﬁc recruitment of eosinophils into the uterus (30). Consistent with this notion, in response to ovarian steroid hormones, the expression of the eosinophil-active chemokines eotaxin-1, RANTES, and MIP-1α is upregulated, paralleling eosinophil inﬁltration into the uterus (29, 33, 34). Indeed, eotaxin-1deﬁcient mice not only have a deﬁciency of uterine eosinophils, but also have a two-week delay in the onset of estrus, along with a delay in the ﬁrst age of parturition, suggesting a role 150

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for eosinophils in preparing the mature uterus for pregnancy (35). Furthermore, eosinophils inﬁltrate the endometrium following copulation (36), and investigators have postulated that this cell may have a role in blastocyst implantation and protection against infection; however, this has yet to be proven (37, 38). Interestingly, eosinophil MBP is ectopically expressed by the uterus during pregnancy, but this is not directly related to eosinophils (39). Eosinophils have also been implicated in postnatal mammary gland development (40). Eosinophils reside in the postnatal developing mammary gland and are predominantly localized around the head of the terminal end buds. The expression level of eotaxin-1 mRNA is low between zero and four weeks of age; however, it is signiﬁcantly increased in the mammary gland at ﬁve weeks of age. Notably, increased expression of eotaxin-1 at this time coincides with eosinophil inﬁltration into the head of the terminal end bud (40). Depletion of eosinophils from the postnatal mammary gland by deletion of the eotaxin-1 gene results in reduction in terminal end bud formation and reduced branching complexity of the ductal tree (40). It is likely that eosinophils regulate mammary gland ductal outgrowth through local secretion of eosinophil-derived TGF-β (40).

Thymic Eosinophils Eosinophils migrate into the thymus during the neonatal period, localizing to the corticomedullary region and reaching maximum levels by two weeks of age. Interestingly, their absolute levels are approximately equivalent to that of thymic dendritic cells (41). In mice, a second inﬂux of eosinophils is observed at 16 weeks of age, corresponding to the commencement of thymic involution. Eosinophils localize to the medullary region. Thymic eosinophils express high levels of MHC class II molecules and moderate levels of MHC class I and the costimulatory molecules CD86 (B7.2) and CD30L (CD153) (Figure 2). Furthermore, thymic eosinophils

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Figure 2 Eosinophil surface markers. This schematic diagram lists surface molecules expressed by human eosinophils. Molecules have been listed generally based on convincing evidence for their expression as assessed by ﬂow cytometry or inferred by cellular responsiveness to speciﬁc stimuli. Cluster designation (CD) for particular molecules is indicated based on the most recent classiﬁcation (www.ncbi.nlm.nih. gov/prow/).

are CD11b/CD11c double positive and appear to be activated as they lose expression of GL-1 and CD62L and upregulate CD25 and CD69 surface expression. Analysis of thymic eosinophil cytokine production reveals that eosinophils express mRNA for the proinﬂammatory cytokines TNF-α, TGF-β, IL1α, and IL-6 and the Th2-cytokines IL-4 and IL-13 (41). Notably, the recruitment of eosinophils into the thymus is regulated by

eotaxin-1, which is constitutively expressed in the thymus (42). It has been postulated that eosinophils are associated with MHC class I–restricted thymocyte deletion. Consistent with this notion, the biphasic recruitment of eosinophils and their anatomical localization within discrete compartments of the thymus coincide with negative selection of double-positive thymocytes (41). Employing an experimental model www.annualreviews.org • The Eosinophil

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of acute negative selection, researchers have demonstrated increased thymic eosinophil levels in MHC class I–restricted male (HY) antigen T cell receptor (TCR) transgenic mice following cognate peptide injection. In addition, eosinophils are associated with clusters of apoptotic bodies, suggesting eosinophil-mediated MHC class I–restricted thymocyte deletion. Thymic eosinophils have the capacity to promote thymocyte apoptosis as they express costimulatory molecules that are involved in clonal deletions, such as CD30 ligand (CD153) and CD66 (41). Additionally, eosinophils may induce thymocyte apoptosis through free radicals, as thymic eosinophils express high levels of NADPH oxidase activity; notably, developing thymocytes have increased sensitivity to free radicals owing to the downregulation of Cu2+ /Zn2+ superoxide dismutase.

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EOSINOPHILS AND IMMUNE REGULATION In recent years, investigators have shown that eosinophils can perform numerous immune functions, including antigen presentation (43, 44) and exacerbation of inﬂammatory responses through their capacity to release a range of largely preformed cytokines and lipid mediators (2, 5).

Antigen Presentation Recent clinical and experimental investigations have shown that eosinophils can function as APCs (Figure 1). Eosinophils can process and present a variety of microbial, viral, and parasitic antigens. (45). In addition, granulocyte-macrophage colony stimulating factor (GM-CSF)-treated eosinophils promote T cell proliferation in response to staphylococcal superantigen (Staphylococcus enterotoxins A, B, and E) stimulation (46). Furthermore, eosinophils incubated with human rhinovirus-16 promote rhinovirus-16speciﬁc T cell proliferation and IFN-γ secretion (47). Eosinophils can also effectively 152

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present soluble antigens to CD4+ T cells, thereby promoting T cell proliferation and polarization. Adoptive transfer of antigenpulsed eosinophils results in eosinophildependent T cell proliferation (44). Furthermore, addition of antigen to eosinophil and T cell cocultures promotes heightened T cell proliferative responses (43). The capacity of eosinophils to present antigen has been debated in some publications. It is interesting to note that the failure of eosinophils to present antigen may be related to the methods used for isolating eosinophils. For example, lysis of erythrocytes with ammonium chloride, an inhibitor of lysosome acidiﬁcation (needed for antigen presentation), negatively correlates with eosinophil antigen presentation activity (43, 48). Eosinophils secrete an array of cytokines (IL-2, IL-4, IL-6, IL-10, IL-12) capable of promoting T cell proliferation, activation, and Th1/Th2 polarization (4, 43, 44, 49) (Figure 1). Recent attention has been drawn to the ability of murine eosinophils to produce IL-4. Employing mice with enhanced green ﬂuorescent protein (GFP) in the IL-4 gene locus (4get mice), investigators have demonstrated that eosinophils are a primary source of GFP following parasitic infection or anti-IgD treatment (a strong Th2 stimulator). Notably, although the IL4 gene locus is transcriptionally active in eosinophils, the amount of IL-4 protein production appears to be lower than in T cells and basophils (50–52). Furthermore, murine eosinophils promote IL-4, IL-5, and IL-13 secretion by CD4+ T cells (44). Eosinophils can also regulate T cell polarization through their synthesis of indoleamine 2,3-dioxygenase (IDO), an enzyme involved in oxidative metabolism of tryptophan, converting tryptophan to kynurenines (KYN). KYN regulates Th1 and Th2 imbalance by promoting Th1 cell apoptosis (53). The eosinophilmediated T cell proliferative and cytokine secretion responses are dependent on costimulation. Indeed, blockade of CD80, CD86, and CTLA-4 by neutralizing antibodies inhibits

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eosinophil-elicited T cell proliferation and cytokine secretion (45). Fluorescent labeling studies revealed that eosinophils instilled into the trachea of mice trafﬁc into the draining peritracheal lymph nodes and localize to the T cell–rich paracortical regions (B cell zones) within 24 h (43). Employing models of allergic airway disease and gastrointestinal allergy, investigators have demonstrated that inhalation of antigen promotes eosinophil homing to the draining endotracheal lymph nodes and Peyer’s patches (44, 54–56). Interestingly, a recent investigation suggests that eosinophils can only promote proliferation of effector T cells but not naive T cells (48). Moreover, eosinophils pulsed with OVA peptide and cocultured with OVAspeciﬁc TCR transgenic T cells (D011.10 T cells) induced effector T cell proliferation; however, when cocultured with naive CD4+ T cells, no T cell proliferation was observed. It is tempting to speculate that eosinophils trafﬁc to draining lymph nodes to recruit activated effector T cells and promote proliferation of effector T cells.

Mast Cell Regulation A substantial body of literature has emerged demonstrating that eosinophils have the capacity to regulate mast cell function (Figure 1). Notably, human umbilical cord blood–derived mast cells can be activated by MBP to release histamine, PGD-2, GM-CSF, TNF-α, and IL-8 (57). The activation of mast cells by MBP elicits not only exocytosis, but also eicosanoid generation and cytokine production, both of which are prominent responses following FcRI-dependent activation of mast cells (57). Incubation of rat peritoneal mast cells with native MBP, EPO, and ECP (but not EDN) results in concentration-dependent histamine release (15). Several studies have shown that MBP induces mast cell activation via a pathway similar to that observed with other polybasic compounds such as substance P, com-

pound 48/80, and bradykinin (16). Freshly isolated human lung mast cells are resistant to IgE-independent activation; however, highly puriﬁed lung mast cells cocultured with human lung ﬁbroblasts are sensitive to IgEindependent activation by MBP (57). Interestingly, activation of eosinophils with the mast cell protease chymase promotes production of eosinophil-derived stem cell factor, a critical mast cell growth factor. Eosinophils also produce nerve growth factor (NGF) (58), a cytokine not only involved in survival and functional maintenance of sympathetic neurons but also in immune regulation. For example, NGF promotes mast cell survival and activation (59, 60). NGF is preformed in eosinophils and acts in an autocrine fashion by activating release of EPO (58). EPO activates rat peritoneal muscles to release histamine, suggesting a role for eosinophil-derived NGF in mast cell–eosinophil interactions. Thus, eosinophils and mast cells communicate in a bidirectional fashion.

EOSINOPHIL DEVELOPMENT Eosinophils are produced in the bone marrow from pluripotential stem cells, which ﬁrst differentiate into a hybrid precursor with shared properties of basophils and eosinophils and then into a separate eosinophil lineage (61). Eosinophil lineage speciﬁcation is dictated by the interplay of at least three classes of transcription factors, including GATA-1 (a zinc family ﬁnger member), PU.1 (an ETS family member), and C/EBP members (CCAAT/enhancer-binding protein family) (62–64) (Figure 3). Although these transcription factors are expressed in a variety of hematopoietic lineages, their mechanism of action in eosinophils is unique. In particular, graded expression of PU.1 speciﬁes distinct cell lineage fates, with low levels specifying lymphocytic and high levels myeloid differentiation (65–67). Although GATA-1 and PU.1 antagonize each other’s function in most cell types, they have synergistic activity in regulating eosinophil lineage speciﬁcation (and www.annualreviews.org • The Eosinophil

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Figure 3 Schematic representation of eosinophil trafﬁcking. Eosinophils develop in the bone marrow, where they differentiate from hematopoietic progenitor cells into mature eosinophils under the control of critical transcription factors, especially GATA-1. The eosinophilopoietins IL-3, IL-5, and GM-CSF regulate eosinophil expansion, especially in conditions of hypereosinophilia. Eosinophil migration out of the bone marrow into the circulation is primarily regulated by IL-5. Circulating eosinophils subsequently interact with the endothelium by processes involving rolling, adhesion, and diapedesis. Depending on the target organ, eosinophils cross the endothelium into tissues by a regulated process involving the coordinated interaction between networks involving the chemokine eotaxin-1, eosinophil adhesion molecules (α4 β1 , α4 β7 , αm β2 , αL β2 ), and adhesion receptors on the endothelium (MAdCAM-1, VCAM-1, and ICAM-1). Under homeostatic conditions, eosinophils trafﬁc into the thymus, mammary gland, uterus, and most prominently into the gastrointestinal tract.

eosinophil granule protein transcription) (67). The speciﬁcity of these factors for eosinophils is conserved across species, as C/EBP factors and GATA-1 drive differentiation of chicken progenitor cells into eosinophils (62). Of these 154

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transcription factors, GATA-1 is clearly the most important for eosinophil lineage speciﬁcation, as revealed by the loss of the eosinophil lineage in mice harboring a targeted deletion of the high-afﬁnity GATA-binding site in

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Figure 4

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Double GATA site. Sequence alignment of the palindromic double GATA high-afﬁnity binding site identiﬁed in a hypersensitivity region of the murine GATA-1 regulatory locus with the dual GATA sites found in the promoters of three human eosinophil lineage-selective genes, including the human IL-5Rα gene promoter 1, human MBP promoter 2, and the human CCR3 regulatory exon-1. This ﬁgure was kindly provided by Drs. J. Du and S. Ackerman, University of Illinois, Chicago.

the GATA-1 promoter (68) and by eosinophil differentiation experiments in vitro (69). In particular, the speciﬁc activity of GATA-1 in eosinophils but not other GATA-1+ lineages (mast cells, megakaryocytes, and erythroid cells) appears to be mediated by a high-afﬁnity palindromic (or double) GATA site (67). This double GATA site is present in the downstream GATA-1 promoter and also in the regulatory regions of eosinophil-speciﬁc genes, including the eotaxin receptor CC chemokine receptor-3 (CCR3), MBP, and the IL-5 receptor alpha (IL-5Rα) gene (Figure 4), and it accounts for eosinophil-speciﬁc gene expression (67, 68, 70). For example, the tandem double GATA site in the human MBP-P2 promoter is required for both promoter activity in human eosinophil cell lines and for the synergistic transactivation by GATA-1 and PU.1 (67). Three cytokines, IL-3, IL-5, and GMCSF, are particularly important in regulating eosinophil development (71–74) (Figure 3). These eosinophilopoietins likely provide permissive proliferative and differentiation signals following the instructive signals speciﬁed by the transcription factors GATA-1, PU.1, and C/EBPs. These cytokines are encoded by closely linked genes on chromosome 5q31. They bind to receptors that share a common beta chain and have unique alpha chains (75). Of these three cytokines, IL-5 is the most speciﬁc to the eosinophil lineage and is responsible for selective differentiation of eosinophils (76). IL-5 also stimulates the release of eosinophils from the bone

marrow into the peripheral circulation (77). The critical role of IL-5 in the production of eosinophils is best demonstrated by genetic manipulation of mice. Overproduction of IL-5 in transgenic mice results in profound eosinophilia (78–81), and deletion of the IL-5 gene causes a marked reduction of eosinophils in the blood and lungs after allergen challenge (82, 83). The overproduction of one or a combination of these three cytokines occurs in humans with eosinophilia, and diseases with selective eosinophilia are often accompanied by overproduction of IL-5 (84). The critical role of IL-5 in regulating eosinophils in humans has been demonstrated by several clinical trials with humanized anti-IL-5 antibody; this currently unapproved drug dramatically lowers eosinophil levels in the blood and to a lesser extent in the inﬂamed lung (85–87).

Double GATA site: a high-afﬁnity palindromic (or double) GATA site located in the regulatory region of eosinophil-speciﬁc genes CCR3: CC chemokine receptor-3, the major eosinophil chemokine receptor that binds the eotaxin family of chemokines Eosinophilopoietins: IL-3, IL-5, and GM-CSF

EOSINOPHIL TRAFFICKING Under baseline conditions, most eosinophils trafﬁc into the gastrointestinal tract where they normally reside within the lamina propria of all segments except the esophagus (88) (Figure 3). The gastrointestinal eosinophil is the predominant population of eosinophils. Under baseline conditions, eosinophil levels in the gastrointestinal tract occur independently of lymphocytes and enteric ﬂora, indicating unique regulation compared with other leukocytes (88). Indeed, the recruitment of gastrointestinal eosinophils is regulated by the constitutive expression of eotaxin-1, www.annualreviews.org • The Eosinophil

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as demonstrated by the marked decrease of this population of eosinophils in eotaxin-1deﬁcient mice. The importance of eotaxin-1 in regulating the baseline level of eosinophils is reinforced by the observation that mice with the targeted deletion of CCR3 (but not eotaxin-2-deﬁcient mice) also have a deﬁciency in gastrointestinal eosinophils (89, 90). In addition to trafﬁcking into the gastrointestinal tract, under homeostatic conditions, eosinophils home into the thymus, mammary gland, and uterus, also under the regulation of eotaxin-1 (40, 91) (Figure 3). Of note, trafﬁcking into the uterus is regulated by estrogen, as eosinophil and eotaxin-1 levels cycle along with estrus (29). The trafﬁcking of eosinophils into inﬂammatory sites involves a number of cytokines (in particular, Th2 and endothelial cell products IL-4, IL-5, and IL-13) (92–94), adhesion molecules (e.g., β1-, β2-, and β7integrins) (95), chemokines (e.g., RANTES and the eotaxins) (96), and other recently identiﬁed molecules (e.g., acidic mammalian chitinase) (97). Tissue eosinophils likely can survive for at least two weeks based on in vitro observations (92). Of the cytokines implicated in modulating leukocyte recruitment, only IL-5 and the eotaxins selectively regulate eosinophil trafﬁcking (98). IL-5 regulates growth, differentiation, activation, and survival of eosinophils and provides an essential signal for the expansion and mobilization of eosinophils from the bone marrow into the lung following allergen exposure (77). However, antigen-induced tissue eosinophilia can occur independently of IL-5, as demonstrated by residual tissue eosinophils in trials using anti-IL-5 in patients with asthma (86) and using IL-5-deﬁcient mice (82, 99). Recent studies have demonstrated an important role for the eotaxin subfamily of chemokines in eosinophil recruitment to the lung (96). Eotaxin was initially discovered using a biological assay in guinea pigs designed to identify the molecules responsible for allergeninduced eosinophil accumulation in the lungs (98, 100, 101). Subsequently, using genomic

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analyses, two additional chemokines were identiﬁed in the human genome that encode for CC chemokines with eosinophilselective chemoattractant activity and have thus been designated eotaxin-2 and eotaxin3 (96). Eotaxin-2 and eotaxin-3 are only distantly related to eotaxin-1 because they are only ∼30% identical in sequence and are located in a different chromosomal position (102, 103). The speciﬁc activity of all eotaxins is mediated by the selective expression of the seven-transmembrane spanning, G protein–coupled receptor CCR3, primarily expressed on eosinophils (104–106). Notably, the eotaxin chemokines cooperate with IL-5 in the induction of tissue eosinophilia. IL-5 increases the pool of eotaxin-responsive cells and primes eosinophils to respond to CCR3 ligands (96). Furthermore, when given exogenously, eotaxins cooperate with IL-5 to induce substantial production of IL-13 in the lung (96). The ﬁnding that IL-4 and IL-13 are potent inducers of the eotaxin chemokines by a STAT6-dependent pathway provides an integrated mechanism to explain the eosinophilia associated with Th2 responses (96). Recent studies have identiﬁed that eosinophil recruitment to the lung is dependent on STAT6 and a bone marrow–derived lung tissue resident non-T or non-B cell (51); in particular, eotaxin-2 production by airway macrophages likely accounts for this (90, 107). Of further interest, recently CCR3 has been shown also to deliver a powerful negative signal in eosinophils, depending on the ligand engaged. For example, pretreatment with the chemokine Mig inhibits eosinophil responses by a CCR3- and Rac2-dependent mechanism (108). Using eotaxin-1 and eotaxin-2 single- and double-gene-deﬁcient mice or neutralizing antibodies, investigators have shown that both chemokines have nonoverlapping roles in regulating the temporal and regional distribution of eosinophils in an allergic inﬂammatory site (90, 109, 110). In a standard experimental asthma model induced by systemic sensitization with OVA/alum followed

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by respiratory OVA challenge, only a modest reduction in lung eosinophils was found in CCR3-deﬁcient mice (89). However, when the same CCR3-deﬁcient mouse line was subjected to experimental asthma induction by epicutaneous OVA sensitization, there was a marked deﬁciency of lung and bronchoalveolar lavage eosinophils (111). It was proposed that these apparently conﬂicting results may be related to the sensitization protocol (111), but the reason for this apparent discrepancy remains unclear. Notably, another CCR3-deﬁcient mouse strain has recently been shown to have a profound reduction in eosinophil recruitment to the lung in the standard OVA/alum systemic sensitization model (107). Substantial preclinical evidence now supports a role for the eotaxin chemokines in human allergic disease (96). Experimental induction of cutaneous and pulmonary latephase responses in humans has revealed that the eotaxin chemokines are produced by tissue resident cells (e.g., respiratory epithelial cells and skin ﬁbroblasts) and allergeninduced inﬁltrative cells (e.g., macrophages and eosinophils). Following allergen challenge in the human lung, eotaxin-1 is induced early (6 h) and correlates with early eosinophil recruitment; in contrast, eotaxin-2 correlates with eosinophil accumulation at 24 h (96). In another study, eotaxin-1 and eotaxin-2 mRNA was increased in patients with asthma compared with normal controls; however, there was no further increase following allergen challenge (96). In contrast, eotaxin-3 mRNA was dramatically enhanced 24 h after allergen challenge (96). The chemoattractant activity of the bronchoalveolar lavage ﬂuid from patients with asthma is inhibited by antibodies against RANTES, MCP (monocyte chemoattractant protein)-3, MCP-4, and eotaxin-1 (96). Further support for an important role of eotaxin-1 in human asthma is derived from analysis of a single nucleotide polymorphism (SNP) in the eotaxin-1 gene. A naturally occurring mutation encoding for a change in the last amino acid in the signal peptide

(alanine→threonine) results in less effective cellular secretion of eotaxin-1 in vitro and in vivo (112). Notably, this SNP is associated with reduced levels of circulating eotaxin-1 and eosinophils and improved lung function (e.g., FEV1) (112). Furthermore, a SNP in the eotaxin-3 gene is associated with atopy in a Korean population (113). Recently, the activity of eotaxin-1 and eotaxin-2 in humans has been investigated by injection of these chemokines into the skin of humans; both eotaxin-1 and eotaxin-2 induce an immediate wheal and ﬂare response associated with mast cell degranulation and subsequent inﬁltrations by eosinophils, basophils, and neutrophils (114). The inﬁltration by neutrophils is likely to be mediated indirectly by the mast cell degranulation. These results provide substantial evidence that the biological activities attributed to eotaxins in animals are conserved in humans. Eosinophils express numerous adhesion molecules, and most attention has focused on their highly expressed integrins, including α4 β7 , the CD18 family of molecules (β2integrins), and the very late antigen (VLA)4 molecules (β1-integrins) (95) (Figure 1). The CD18 family of molecules includes lymphocyte function antigen (LFA)-1 and Mac-1 that interact with endothelial cells via intercellular adhesion molecule (ICAM)-1. VLA4 interacts with endothelium via vascular cell adhesion molecule (VCAM)-1, as well as ﬁbronectin. The α4 β7 integrin interacts with the mucosal addressin cell adhesion molecule (MAdCAM)-1 expressed by vascular endothelium in the intestinal tract. These integrins have variable roles in eosinophil trafﬁcking during inﬂammation, but the role of speciﬁc adhesion molecules in the baseline homing of eosinophils into the gastrointestinal tract has yet to be elucidated. For example, in β7 genetargeted mice, there is a delay and reduced magnitude in the development of intestinal eosinophilia following Trichinella spiralis infection (115) and when the eotaxin-1 intestine transgene is expressed, but no changes in the baseline level of small intestine eosinophils www.annualreviews.org • The Eosinophil

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(81). Analysis of anti-β1-treated mice or VLA-4-deﬁcient mice has shown the critical participation of this family of molecules in regulating eosinophil homing to the allergic lung (116–118). Indeed, eotaxin-1-stimulated eosinophils have increased expression and avidity of VLA-4 (119). It has also become clear that engagement of eosinophil adhesion molecules with their ligands not only induces a proadhesive pathway, but also activates expression of a series of proinﬂammatory genes within eosinophils, including GMCSF, that then propagate eosinophil survival by a paracrine pathway. Numerous other pathways for regulating eosinophil accumulation and trafﬁcking are operational in various inﬂammatory models. However, recently several lines of evidence have focused attention on the importance of arachidonic acid metabolites, especially leukotriene B4 (LTB4), the cysteinyl leukotrienes (LTC4, LTCD4, and LTE4), and prostaglandin (PG) D2. Notably, cysteinyl leukotriene type 1 receptor antagonists (now approved for asthma therapy) reduce blood and lung eosinophilia. Mice with the targeted deletion of the LTB4 receptor also have markedly reduced allergen-induced lung eosinophilia (120). Furthermore, eosinophils express high levels of a high-afﬁnity PGD2 type 2 receptor. Interestingly, this receptor is also expressed by basophils and Th2 cells [and is now designated chemoattractant receptor Th2 cells (CRTH2)] and appears to co-mediate Th2 cell and eosinophil/basophil recruitment (121). Eosinophils have recently also been shown to express high levels of the histamine receptor 4 (H4) that mediates eosinophil chemoattraction and activation in vitro (122).

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defend the host against parasitic helminths. This is based on several lines of evidence, including (a) the ability of eosinophils to mediate antibody- (or complement-) dependent cellular toxicity against helminths in vitro (27), (b) the observation that eosinophil levels increase during helminthic infections and that eosinophils aggregate and degranulate in the local vicinity of damaged parasites in vivo, and (c) the results in experimental parasite infected mice that have been depleted of eosinophils by IL-5 neutralization and/or gene targeting (123). Murine studies are particularly problematic because mice are not the natural hosts of many of the experimental parasites; nevertheless, in some primary infection models, a role for IL-5 in protective immunity has been suggested following infection with Strongyloides venezuelensis, Strongyloides ratti, Nippostrongyloides brasiliensis, and Heligmosomoides polygyrus (123, 124). These in vivo studies need to be interpreted with caution because IL-5 neutralization may have effects on other IL-5 receptor bearing cells (including murine B cells, human basophils, and possibly human respiratory smooth muscle cells) (76, 125–127). Other approaches, including analysis of CCR3- and eotaxin-1deﬁcient mice, have recently demonstrated a role for eosinophils in the encystment of larvae in Trichinella spiralis and in controlling the Brugia malayi microﬁlariae, respectively (128, 129). Perhaps analysis of the recently generated eosinophil-deﬁcient mice following experimental parasitic infection will provide further compelling evidence that eosinophils participate in host defense against parasites. Thus, although the debate continues, it seems likely that eosinophils participate in the protective immunity against selected helminths. Evidence is emerging that eosinophils may also have a protective role in other infections, especially against RNA viruses such as respiratory syncytial virus (RSV) and the related natural rodent pathogen, pneumonia virus of mice (PVM), in vivo (13, 130). Notably, eosinophil granule proteins include abundant

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ribonucleases [such as human ECP and EDN, and at least 11 eosinophil-associated ribonuclease (EAR) orthologs in mice] that degrade single-stranded RNA containing viruses (13). In fact, ECP and EDN are the most divergent coding sequences in the entire human genome (compared with other primates) (13). Despite their divergence, they have conserved ribonuclease activity across species, strongly implicating evolutionary pressure to preserve this critical enzymatic activity.

Asthma Elevated levels of eosinophil granule proteins (e.g., MBP) have been found in bronchoalveolar lavage ﬂuid from patients with asthma, and importantly these concentrations are sufﬁcient to induce cytotoxicity of a variety of host tissue, including respiratory epithelial cells in vitro (3). Direct degranulation of mast cells and basophils, triggered by MBP, is thought also to be involved in disease pathogenesis (3). In addition to being cytotoxic, MBP directly increases smooth muscle reactivity by causing dysfunction of vagal muscarinic M2 receptors, which is thought to contribute to the development of airway hyperreactivity (AHR), a cardinal feature of asthma (131). Additionally, eosinophils generate large amounts of the cysteinyl leukotrienes (132). Of note, eosinophil granule proteins contain all the biochemical machinery necessary to synthesize cysteinyl leukotrienes (132). These mediators lead to increased vascular permeability and mucus secretion and are potent smooth muscle constrictors. Indeed, inhibitors of cysteinyl leukotrienes are effective therapeutic agents for the treatment of allergic airway disease. Multiple studies employing experimental models of asthma (primarily in mice, guinea pigs, and monkeys) have demonstrated that neutralization of IL-5 can block various aspects of asthma (82, 133). Although extensive investigations have implicated the eosinophil as a central effector cell in asthma and an important clinical target for the resolution

of this disease, the role of this granulocyte in the development and exacerbation of asthma pathogenesis has been controversial. This controversy stems in part from distinctions between human asthma and experimental murine models of asthma. For example, in contrast to human asthma, mice with eosinophil lung disease triggered by allergens or helminthic infection have variable levels of eosinophil degranulation (50, 134). In experimental models, inhibition of the actions of IL-5 consistently suppresses pulmonary eosinophilia in response to antigen inhalation; however, this effect does not always correlate with a reduction of AHR (135). This dichotomy is highlighted by ﬁndings in allergic IL-5-deﬁcient mice of the C57BL/6 strain (82) that do not develop antigen-induced AHR, whereas IL-5-deﬁcient BALB/c mice develop enhanced reactivity independent of this factor (136). Although eosinophil trafﬁcking to the allergic lung is profoundly attenuated in IL-5-deﬁcient mice or in those treated with anti-IL-5 antibodies in comparison to wild-type responses (137–139), a marked residual tissue eosinophilia can persist in these mice after allergen inhalation (82, 140, 141). Furthermore, the degree of residual tissue eosinophilia is under genetic regulation, as lung eosinophilia is 10- to 100-fold greater in the BALB/c strain, where AHR persists, compared with the C57BL/6 strain, where AHR is abolished in the absence of IL-5 (82, 99, 138). Studies with transgenic mice overexpressing IL-5 (in T cells, lung epithelial cells, or enterocytes) have demonstrated that overexpression of IL-5 is sufﬁcient for the development of eosinophilia (78–81, 142); however, elevated levels of eosinophils are not universally associated with the development of asthma-like changes in the lung. Indeed, clinical studies in patients have shown that AHR correlates with mast cell localization near pulmonary nerves, whereas pulmonary eosinophilia relates more strongly with chronic cough (143). However, depletion of murine eosinophils (by administration www.annualreviews.org • The Eosinophil

Airway hyperresponsiveness or hyperreactivity (AHR): increased constriction of the airways to various stimuli such as methacholine

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Airway remodeling: microscopic changes (e.g., goblet cell metaplasia, collagen deposition, smooth muscle hyperplasia) in the lungs associated with functional alterations in lung function PHIL mice: genetically engineered eosinophil-deﬁcient mice produced by insertion of the diphtheria toxin A chain into the EPO gene locus dbl-GATA-1 mice: genetically engineered eosinophil-deﬁcient mice produced by deleting the high-afﬁnity double GATA site in the GATA-1 promoter

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of complement-ﬁxing antibodies against CCR3) has demonstrated an important role for eosinophils in the development of asthmaassociated AHR (144); a role for other CCR3+ cells was not ruled out, but there was no evidence for CCR3 expression by non-eosinophils (144). Accordingly, a humanized antibody against IL-5 has recently been tested for asthma (85). In the early studies with this reagent, patients with mild to moderate asthma were shown to have a drop in their circulating and sputum eosinophil levels (85); however, no clinical beneﬁt (e.g., improvement in FEV1) was demonstrated. This result prompted some investigators to conclude that eosinophils were not effector cells in human asthma (85); however, the anti-IL5 study was not properly designed to address the efﬁcacy of this drug (145). In support of these preclinical studies, a very recent study has demonstrated that anti-IL-5 in humans blocks lung eosinophil recruitment by only 55% (146), providing evidence that accessory molecules (in addition to IL-5) regulate lung eosinophilia. Thus, anti-IL-5 treatment does not completely resolve tissue eosinophilia in the allergic lung, and therefore this cell may still contribute to disease pathogenesis even in

Table 1 Effect of eosinophil depletion on experimental asthma parametersa Mouse line

PHIL

Δdbl-GATA

Asthma parameter BALF eosinophils Lung tissue eosinophils BALF mononuclear cells AHR Mucus production Collagen deposition Th2 antibody production Th2 cytokines

+ + NE + + ND ND +

+ + NE NE NE + NE NE

a If the genetic manipulation of the mouse resulted in protection from or reduction in severity of the asthma parameter, the parameter is labeled with a “+”. If there was no change in the asthma parameter between the genetically modiﬁed mouse and wild-type control mice, the parameter is labeled with “NE” for no effect. If the asthma parameter was not measured, the parameter is labeled with “ND” for not determined.

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the presence of IL-5 neutralization. With the discovery of the eotaxins, and the ﬁnding that IL-5 cooperates with eotaxins in regulating eosinophil tissue recruitment, it became critical to determine if the lack of efﬁcacy of antiIL-5 in humans was related to the inability of this drug to block eosinophil tissue recruitment or to the noneffector role of eosinophils. One possibility is that local chemokine systems (eotaxins) can operate independently of IL-5 to recruit eosinophils into the allergic lung. Studies with eotaxin-1 gene-targeted mice, IL-5 gene-targeted mice, and eotaxin1/IL-5 double-gene-targeted mice have revealed an independent and synergistic role for both of these molecules in regulating the tissue level of eosinophils in the asthmatic lung and in the induction of AHR (139). Although early studies with anti-IL-5 in human asthma have continued to ﬁnd no improvement in airﬂow measurements (FEV1), pathological markers of chronic airway remodeling (e.g., deposition of tenascin, procollagen III, and lumican) are improved by anti-IL-5 (146). Decreased levels of TGF-β in the bronchoalveolar lavage ﬂuid following anti-IL-5 treatment have been found, suggesting that eosinophilderived TGF-β regulates lung remodeling. In support for a role of eosinophils in the pathogenesis of human asthma, a very recent study has demonstrated improved clinical outcome when asthma treatment decisions are based on monitoring sputum eosinophil counts rather than conventional guidelines from the British Thoracic Society (147). Recently, two different lines of eosinophildeﬁcient mice were developed (see Table 1 and Eosinophil-Deﬁcient Mice). Lee et al. (148) targeted the depletion of eosinophils by using an eosinophil-speciﬁc promoter to drive expression of a cytocidal protein, diphtheria toxin A chain. The eosinophil-deﬁcient character of these mice (called PHIL mice) was assessed by examination of peripheral blood and by immunohistochemistry of tissues with abundant resident populations (e.g., bone marrow, uterus, small intestine, and thymus) using antibodies speciﬁc for eosinophil

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granule proteins. In comparison, Yu et al. (68) developed mice harboring a deletion of a high-afﬁnity GATA-binding site in the GATA-1 promoter (dbl-GATA) which led to the speciﬁc ablation of the eosinophil lineage. RT-PCR analysis of gene expression in the bone marrow of the dbl-GATA mice revealed no expression of EPO, but expression of MBP was only partially reduced and CCR3 expression remained unchanged. Nevertheless, eosinophil deﬁciency in these mice was veriﬁed by morphological observation of cells from the blood, bone marrow, and spleen. Using both lines of eosinophildeﬁcient mice, eosinophils were shown to have an integral role in experimental allergic asthma. However, their speciﬁc contribution toward allergen-induced AHR and mucus cell metaplasia was different (Table 1). Perhaps dbl-GATA mice have residual eosinophils or unappreciated hematological abnormalities, or alternatively, diptheria toxin treatment of PHIL mice may induce toxic effects on noneosinophils; these and other explanations for the distinct results will hopefully be uncovered soon. It should be noted that dbl-GATA mice had impaired development of lung remodeling in a chronic model of asthma, consistent with the results of anti-IL-5 in patients with asthma. Taken together, compelling evidence now exists that eosinophils are prominent effector cells in eliciting multiple parameters of experimental asthma.

Gastrointestinal Disorders The accumulation of eosinophils in the gastrointestinal tract is a common feature of numerous disorders, such as drug reactions, helminth infections, hypereosinophilic syndromes, eosinophilic gastroenteritis, allergic colitis, inﬂammatory bowel disease, and gastroesophageal reﬂux disease (150). A subset of these diseases, referred to as primary eosinophil-associated gastrointestinal disorders (EGID), includes eosinophilic esophagitis (EE), eosinophilic gastritis, and

Eosinophil-Deficient Mice Two different lines of eosinophil-deﬁcient mice have recently been developed. One group targeted the depletion of eosinophils using an eosinophil-speciﬁc promoter (the EPO gene) to drive expression of a cytocidal protein diphtheria toxin A (13). These mice (called PHIL) are protected from the development of AHR in a model of experimental asthma. Another group developed mice harboring a deletion of the high-afﬁnity GATA-binding site in the GATA-1 promoter (dbl-GATA); this led to the speciﬁc ablation of the eosinophil lineage even when these mice were crossed with IL-5 transgenic mice (48). The dbl-GATA mice are protected from features of airway remodeling but not AHR in an experimental model of asthma. It is anticipated that these newly generated eosinophil-deﬁcient mouse lines will transform eosinophil research over the next decade, especially because the dbl-GATA mice are now commercially available from Jackson Laboratories, Inc.

eosinophilic gastroenteritis. These are hypersensitivity disorders that lie in the middle of a spectrum ranging from anaphylaxis to Celiac disease (150). EGID usually occurs independently of peripheral blood eosinophilia, indicating the signiﬁcance of gastrointestinalspeciﬁc mechanisms for regulating eosinophil levels. Indeed, in murine models of EGID, a deﬁnitive role for eosinophils and eotaxin-1 has been demonstrated. Notably, eosinophils are frequently associated near damaged enteric nerves, and indeed eotaxin-1-deﬁcient mice are protected from this feature of disease. EE is distinguished from gastroesophageal reﬂux disease by several important differences, including the relatively higher prevalence of atopy, dysphagia, male gender, familial inheritance, degree of proximal esophagitis, and intensity of esophageal pathology [e.g., epithelial hyperplasia and eosinophil density (generally >24 eosinophils/high power ﬁeld)] (150). Consistent with the high rate of atopic respiratory disease in patients with EE, experimental EE develops in mice following respiratory allergen exposure or following intratracheal IL-13 delivery (151). These www.annualreviews.org • The Eosinophil

Hypereosinophilic syndrome: a group of disorders characterized by severely elevated blood eosinophil levels and end-organ damage EGID: eosinophil-associated gastrointestinal disorders EE: eosinophilic esophagitis

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FIP1L1-PDGFRA: activated tyrosine kinase fusion gene product that occurs in hypereosinophilic syndrome owing to an 800-kb interstitial deletion in chromosome 4

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results establish an intimate immunological connection between the lung and esophagus. The epithelial hyperplasia associated with EE and the level of esophageal eosinophils is attenuated in IL-5-deﬁcient mice (152), providing strong evidence that eosinophils are effector cells in this gastrointestinal disease. Indeed, a recent preliminary evaluation of humanized anti-IL-5 in patients with EE demonstrates lowering of esophageal eosinophil levels. Supporting a connection between allergic responses in the lung and gastrointestinal tract, eotaxin-1 intestine transgenic mice not only develop intestinal eosinophilia but also AHR by an IL-13dependent mechanism (153). Thus, increased expression of eotaxin-1 in the gastrointestinal compartment can lead to increased CD4+ T cell–derived Th2 lymphocyte-cytokine production that drives aberrant immunophysiological responses in distant noninﬂamed mucosal tissue (the lung). These results provide a possible explanation for the altered lung function seen in some patients with inﬂammatory gastrointestinal disorders.

ANTI-EOSINOPHIL THERAPEUTICS Numerous drugs inhibit eosinophil production or eosinophil-derived products. They include glucocorticoids, myelosuppressive drugs, leukotriene synthesis or receptor antagonists, tyrosine kinase inhibitors, IFN-α, and humanized anti-IL-5 antibodies. The etiology of the primary disease often speciﬁes the best therapeutic strategy. For example, a subset of patients with hypereosinophilic syndrome have an 800-kb interstitial deletion on chromosome 4 (4q12) that results in the fusion of an unknown gene FIP1L1 with the platelet-derived growth factor receptor-α (PDGFRA) gene (154, 155). This fusion gene produces a constitutively active tyrosine kinase (PDGFRA) that is exquisitely sensitive to the inhibitor imatinib mesylate, which is now approved for the treatment of several malig162

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nancies (GleevecTM ). Although PDGFRA is not normally active in hematopoietic cells, the activated kinase renders cells growth factor independent, perhaps by activating STAT5 signal transduction. Thus, eosinophilic patients with FIP1L1-PDGFRA+ disease are now treated with GleevecTM as ﬁrst-line therapy (156). In addition, a variety of other activated tyrosine kinases have just been associated with hypereosinophilic syndromes, including PDGFRB, Janus kinase-2, and ﬁbroblast growth factor receptor-1. In most other individuals, glucocorticoids are the most effective agents for reducing eosinophilia (3). They suppress the transcription of a number of genes for inﬂammatory mediators, including the genes for IL-3, IL-4, IL-5, GM-CSF, and various chemokines including the eotaxins. Recently, the main action of glucocorticoids on eosinophil-active cytokines has been shown to involve mRNA destabilization, thus reducing the half-life of cytokines such as eotaxins (157). In addition, glucocorticoids inhibit the cytokinedependent survival of eosinophils (158). Systemic or topical (inhaled or intranasal) glucocorticoid treatment typically causes a rapid reduction in eosinophils, but some patients are glucocorticoid resistant and maintain eosinophilia despite high doses (159). The mechanism of glucocorticoid resistance is unclear, but a reduced level of glucocorticoid receptors and alterations in transcription factor activator protein (AP)-1 appear to be at least partially responsible (159). Glucocorticoid-resistant patients sometimes require other therapy such as myelosuppressive drugs (hydroxyurea, vincristine) or IFN-α (3). IFN-α can be especially helpful because it inhibits eosinophil degranulation and effector function (160). Notably, patients with myeloproliferative variants of hypereosinophilic syndrome can often go into remission with IFN-α therapy. Cyclophilins (e.g., cyclosporine A) have also been used because they block the transcription of numerous eosinophil-active cytokines (e.g.,

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IL-5, GM-CSF) (3). Recently, lidocaine has been shown to shorten eosinophil survival, and its effects mimic those of glucocorticoids and are noncytotoxic (161). Indeed, an early clinical trial has shown that nebulized lidocaine is safe and effective in subjects with asthma (162). Drugs that interfere with eosinophil chemotactic signals include recently approved leukotriene antagonists and inhibitors. 5lipoxygenase inhibition (e.g., zileuton) blocks the rate-limiting step in leukotriene synthesis and inhibits the generation of the eosinophil chemoattractant, LTB4, and the cysteinyl leukotrienes (163). Cysteinyl leukotriene receptor antagonists block the muscle contraction and increased vascular permeability mediated by leukocyte-derived leukotrienes (164). Some of the third generation antihistamines inhibit the vacuolization (165) and accumulation (166) of eosinophils after allergen challenge and directly inhibit eosinophils in vitro (165, 167). Cromoglycate and nedocromil inhibit the effector function of eosinophils, such as antibody-dependent cellular cytotoxicity (167). The identiﬁcation of molecules that specifically regulate eosinophil function and/or production offers new therapeutic strategies in the pipeline. Agents that interrupt eosinophil adhesion to the endothelium through the interaction of CD18/ICAM-1 (168) or VLA-4 /VCAM-1 may be useful (169, 170). Indeed, antibodies that block these pathways have recently been approved for other diseases, but their anti-eosinophil activity has yet to be determined (171). Antibodies against IL-5, now humanized by two different pharmaceutical companies, are under active clinical investigation (172, 173). Although their utility for asthma may be limited owing to redundant pathways, anti-IL-5 is particularly promising for hypereosinophilic syndromes. Numerous inhibitors of the eotaxin/CCR3 pathway, including small molecule inhibitors of CCR3 and a human anti-eotaxin-1 antibody, are being developed (96). Early results with a phase I trial of human anti-eotaxin-1 antibody in

patients with allergic rhinitis have demonstrated the ability of this apparently safe drug to lower levels of nasal eosinophils and to improve nasal patency (96). Anti-human IL-13 antibody is now in preclinical trials (174) and looks promising for lowering tissue eosinophil levels. Finally, a recently identiﬁed eosinophil surface molecule Siglec-8 may offer a therapeutic opportunity (175). Siglec-8 is a member of the sialic acid–binding lectin family and contains ITIMs (immunoreceptor tyrosinebased inhibitory motifs) that can induce efﬁcient eosinophil apoptosis when engaged by anti-Siglec-8 crosslinking antibodies. Siglec8 as well as CCR3 and CRTH2 are coexpressed by other cells involved in Th2 responses, including Th2 cells, mast cells, and basophils. Thus, agents that block these receptors may be particularly useful for allergic disorders.

PERSPECTIVE Historically, eosinophils have been considered end-stage cells involved in host protection against parasites. However, numerous lines of evidence have now changed this perspective by showing that eosinophils are pleiotropic multifunctional leukocytes involved in initiation and propagation of diverse inﬂammatory responses, as well as modulators of adaptive immunity by directly activating T cells. As normal constituents of the mucosal immune system, particularly in the gastrointestinal tract, eosinophils are likely to have a physiological function. Indeed, eosinophils have been implicated in innate immunity by being an early and possibly instrumental source of cytokines (e.g., IL-4) and have a role in developmental processes such as mammary gland development. Analysis of recently generated genetically engineered eosinophil-deﬁcient mice will soon answer critical questions concerning the true involvement of this cell type in a variety of processes. Breakthroughs in identifying key eosinophil regulatory cytokines such as IL-5 and the eotaxin subfamily of chemokines www.annualreviews.org • The Eosinophil

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have uncovered mechanisms that selectively regulate eosinophil production and localization at baseline and during inﬂammatory responses. In particular, an integrated mechanism involving Th2 cell–derived IL-5 regulating eosinophil expansion in the bone marrow and blood and Th2 cell–derived IL-13 reg-

ulating eotaxin production now explains the means by which T cells regulate eosinophils. Based on these ﬁndings, targeted therapy against key eosinophil regulators (e.g., humanized anti-IL-5 and CCR3 antagonists) will likely transform medical management of eosinophilic patients.

DISCLOSURE STATEMENT

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

M.R. has consulted on an advisory board for GlaxoSmithKline, received stock and an honorarium for consulting for Ception Therapeutics, received a speaker’s honorarium from Merck and Tanox, and has consulted and received a research grant from Cambridge Antibody Technology.

ACKNOWLEDGMENTS Dr. Rothenberg’s laboratory is indebted to the following grants and/or organizations that have partly ﬁnanced the work presented in this review: NIAID R01 AI045898, NIAID R01 AI057803, NHLBI P01 HL076383, FDA FD-R-002313, the Campaign Urging Research for Eosinophilic Disorders (CURED), and the Burroughs Wellcome Fund. The authors thank numerous instrumental colleagues who have contributed to the ideas formulated in this review, including Drs. K. Frank Austen, Fred Finkelman, Paul Foster, Ian Young, Nives Zimmermann, Anil Mishra, Steven Ackerman, James Lee, and Gerald Gleich, the dedicated laboratory workers, and Andrea Lippelman for editorial assistance. The authors are grateful to Drs. Jamie and Nancy Lee, Craig Gerard, and Alison Humbles for sharing their innovative reagents, and to the International Eosinophil Society that recently hosted a meeting where some of the information in this review was formulated.

LITERATURE CITED 1. Gleich GJ, Loegering DA. 1984. Immunobiology of eosinophils. Annu. Rev. Immunol. 2:429–59 2. Weller PF. 1994. Eosinophils: structure and functions. Curr. Opin. Immunol. 6:85–90 3. Rothenberg ME. 1998. Eosinophilia. N. Engl. J. Med. 338:1592–600 4. Kita H. 1996. The eosinophil: a cytokine-producing cell? J. Allergy Clin. Immunol. 97:889– 92 5. Gleich GJ, Adolphson CR. 1986. The eosinophilic leukocyte: structure and function. Adv. Immunol. 39:177–253 6. Tai P-C, Hayes DJ, Clark JB, Spry CJF. 1982. Toxic effects of eosinophil secretion products on isolated rat heart cells in vitro. Biochem. J. 204:75–80 7. Venge P, Dahl R, Hallgren R, Olsson I. 1980. Cationic proteins of human eosinophils and their role in the inﬂammatory reaction. In The Eosinophil in Health and Disease, ed. AAF Mahmoud, KF Austen, pp. 1131–42. New York: Grune & Stratton 8. Frigas E, Loegering DA, Gleich GJ. 1980. Cytotoxic effects of the guinea pig eosinophil major basic protein on tracheal epithelium. Lab. Invest. 42:35–43 9. Gleich GJ, Frigas E, Loegering DA, Wassom DL, Steinmuller D. 1979. The cytotoxic properties of the eosinophil major basic protein. J. Immunol. 123:2925 10. Young JD, Peterson CG, Venge P, Cohn ZA. 1986. Mechanism of membrane damage mediated by human eosinophil cationic protein. Nature 321:613–16 164

Rothenberg

·

Hogan

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-05

ARI

23 February 2006

15:12

11. Slifman NR, Loegering DA, McKean DJ, Gleich GJ. 1986. Ribonuclease activity associated with human eosinophil-derived neurotoxin and eosinophil cationic protein. J. Immunol. 137:2913–17 12. Gleich GJ, Loegering DA, Bell MP, Checkel JL, Ackerman SJ, McKean DJ. 1986. Biochemical and functional similarities between human eosinophil-derived neurotoxin and eosinophil cationic protein: homology with ribonuclease. Proc. Natl. Acad. Sci. USA 83:3146–50 13. Rosenberg HF, Domachowske JB. 2001. Eosinophils, eosinophil ribonucleases, and their role in host defense against respiratory virus pathogens. J. Leukoc. Biol. 70:691– 98 14. Venge P, Bystrom J, Carlson M, Hakansson L, Karawacjzyk M, et al. 1999. Eosinophil cationic protein (ECP): molecular and biological properties and the use of ECP as a marker of eosinophil activation in disease. Clin. Exp. Allergy 29:1172–86 15. Zheutlin LM, Ackerman SJ, Gleich GJ, Thomas LL. 1984. Stimulation of basophil and rat mast cell histamine release by eosinophil granule-derived cationic proteins. J. Immunol. 133:2180–85 16. Piliponsky AM, Pickholtz D, Gleich GJ, Levi-Schaffer F. 2001. Human eosinophils induce histamine release from antigen-activated rat peritoneal mast cells: a possible role for mast cells in late-phase allergic reactions. J. Allergy Clin. Immunol. 107:993–1000 17. Jacoby DB, Costello RM, Fryer AD. 2001. Eosinophil recruitment to the airway nerves. J. Allergy Clin. Immunol. 107:211–18 18. Morgan RK, Costello RW, Durcan N, Kingham PJ, Gleich GJ, et al. 2005. Diverse effects of eosinophil cationic granule proteins on IMR-32 nerve cell signalling and survival. Am. J. Respir. Cell Mol. Biol. 33:169–77 19. Agosti JM, Altman LC, Ayars GH, Loegering DA, Gleich GJ, Klebanoff SJ. 1987. The injurious effect of eosinophil peroxidase, hydrogen peroxide, and halides on pneumocytes in vitro. J. Allergy Clin. Immunol. 79:496–504 20. Wu W, Chen Y, Hazen SL. 1999. Eosinophil peroxidase, nitrates, protein tyrosyl residues. Implications for oxidative damage by nitrating intermediates in eosinophilic inﬂammatory disorders. J. Biol. Chem. 274:25933–44 21. MacPherson JC, Comhair SA, Erzurum SC, Klein DF, Lipscomb MF, et al. 2001. Eosinophils are a major source of nitric oxide-derived oxidants in severe asthma: Characterization of pathways available to eosinophils for generating reactive nitrogen species. J. Immunol. 166:5763–72 22. Logan MR, Odemuyiwa SO, Moqbel R. 2003. Understanding exocytosis in immune and inﬂammatory cells: the molecular basis of mediator secretion. J. Allergy Clin. Immunol. 111:923–32 23. Dvorak AM, Furitsu T, Letourneau L, Ishizaka T, Ackerman SJ. 1991. Mature eosinophils stimulated to develop in human cord blood mononuclear cell cultures supplemented with recombinant human interleukin-5. Part I. Piecemeal degranulation of speciﬁc granules and distribution of Charcot-Leyden crystal protein. Am. J. Pathol. 138:69– 82 24. Lacy P, Mahmudi-Azer S, Bablitz B, Hagen SC, Velazquez JR, et al. 1999. Rapid mobilization of intracellularly stored RANTES in response to interferon-γ in human eosinophils. Blood 94:23–32 25. Bandeira-Melo C, Gillard G, Ghiran I, Weller PF. 2000. EliCell: a gel-phase dual antibody capture and detection assay to measure cytokine release from eosinophils. J. Immunol. Methods 244:105–15 www.annualreviews.org • The Eosinophil

165

ARI

23 February 2006

15:12

26. Logan MR, Lacy P, Bablitz B, Moqbel R. 2002. Expression of eosinophil target SNAREs as potential cognate receptors for vesicle-associated membrane protein-2 in exocytosis. J. Allergy Clin. Immunol. 109:299–306 27. Butterworth AE. 1977. The eosinophil and its role in immunity to helminth infection. Curr. Top. Microbiol. Immunol. 77:127–68 28. Sferruzzi-Perri AN, Robertson SA, Dent LA. 2003. Interleukin-5 transgene expression and eosinophilia are associated with retarded mammary gland development in mice. Biol. Reprod. 69:224–33 29. Gouon-Evans V, Pollard JW. 2001. Eotaxin is required for eosinophil homing into the stroma of the pubertal and cycling uterus. Endocrinology 142:4515–21 30. Robertson SA, Mau VJ, Young IG, Matthaei KI. 2000. Uterine eosinophils and reproductive performance in interleukin 5-deﬁcient mice. J. Reprod. Fertil. 120:423–32 31. Nagral A, Ben-Ari Z, Dhillon AP, Burroughs AK. 1998. Eosinophils in acute cellular rejection in liver allografts. Liver Transpl. Surg. 4:355–62 32. Tepper RI, Coffman RL, Leder P. 1992. An eosinophil-dependent mechanism for the antitumor effect of interleukin-4. Science 257:548–51 33. Robertson SA, Allanson M, Mau VJ. 1998. Molecular regulation of uterine recruitment during early pregnancy in the mouse. Trophobl. Res. 11:101–20 34. Zhang J, Lathbury LJ, Salamonsen LA. 2000. Expression of the chemokine eotaxin and its receptor, CCR3, in human endometrium. Biol. Reprod. 62:404–11 35. Gouon-Evans V, Lin EY, Pollard JW. 2002. Requirement of macrophages and eosinophils and their cytokines/chemokines for mammary gland development. Breast Cancer Res. 4:155–64 36. McMaster MT, Newton RC, Dey SK, Andrews GK. 1992. Activation and distribution of inﬂammatory cells in the mouse uterus during the preimplantation period. J. Immunol. 148:1699–705 37. Robertson SA, Mau VJ, Hudson SA, Tremellen KP. 1997. Cytokine-leukocyte networks and the establishment of preganancy in the mouse. J. Reprod. Fertil. 107:265–77 38. De M, Choudhuri R, Wood GW. 1991. Determination of the number and distribution of macrophages, lymphocytes and granulocytes in the mouse uterus from mating through implantation. J. Leukoc. Biol. 50:252–62 39. Wagner JM, Hustin J, Bonno M, Kephart GM, Gurian KV, Gleich GJ. 1994. Pregnancyassociated major basic protein: deposition of protein and expression of mRNA at the maternal-fetal junction in early and late gestation. Placenta 15:625–40 40. Gouon-Evans V, Rothenberg ME, Pollard JW. 2000. Postnatal mammary gland development requires macrophages and eosinophils. Development 127:2269–82 41. Throsby M, Herbelin A, Pleau JM, Dardenne M. 2000. CD11c+ eosinophils in the murine thymus: developmental regulation and recruitment upon MHC class I-restricted thymocyte deletion. J. Immunol. 165:1965–75 42. Matthews AN, Friend DS, Zimmermann N, Saraﬁ MN, Luster AD, et al. 1998. Eotaxin is required for the baseline level of tissue eosinophils. Proc. Natl. Acad. Sci. USA 95:6273– 78 43. Shi HZ, Humbles A, Gerard C, Jin Z, Weller PF. 2000. Lymph node trafﬁcking and antigen presentation by endobronchial eosinophils. J. Clin. Invest. 105:945–53 44. MacKenzie JR, Mattes J, Dent LA, Foster PS. 2001. Eosinophils promote allergic disease of the lung by regulating CD4+ Th2 lymphocyte function. J. Immunol. 167:3146–55 45. Shi H. 2004. Eosinophils function as antigen-presenting cells. J. Leukoc. Biol. 76:520– 27

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-05

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Rothenberg

·

Hogan

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ANRV270-IY24-05

ARI

23 February 2006

15:12

46. Mawhorter SD, Kazura JW, Boom WH. 1994. Human eosinophils as antigen-presenting cells: relative efﬁciency for superantigen- and antigen-induced CD4+ T-cell proliferation. Immunology 81:584–91 47. Handzel ZT, Busse WW, Sedgwick JB, Vrtis R, Lee WM, et al. 1998. Eosinophils bind rhinovirus and activate virus-speciﬁc T cells. J. Immunol. 160:1279–84 48. van Rijt LS, Vos N, Hijdra D, De Vries VC, Hoogsteden HC, Lambrecht BN. 2003. Airways eosinophils accumulate in the mediastinal lymph nodes but lack antigen-presenting potential for naive T-cells. J. Immunol. 171:3372–78 49. Lacy P, Moqbel R. 2000. Eosinophil cytokines. Chem. Immunol. 76:134–55 50. Shinkai K, Mohrs M, Locksley RM. 2002. Helper T cells regulate type-2 innate immunity in vivo. Nature 420:825–29 51. Voehringer D, Shinkai K, Locksley RM. 2004. Type 2 immunity reﬂects orchestrated recruitment of cells committed to IL-4 production. Immunity 20:267–77 52. Khodoun MV, Orekhova T, Potter C, Morris S, Finkelman FD. 2004. Basophils initiate IL-4 production during a memory T-dependent response. J. Exp. Med. 200:857–70 53. Odemuyiwa SO, Ghahary A, Li Y, Puttagunta L, Lee JE, et al. 2004. Human eosinophils regulate T cell subset selection through indoleamine 2,3-dioxyenase. J. Immunol. 173:5909–13 54. Korsgren M, Erjefalt JS, Korsgren O, Sundler F, Persson CGA. 1997. Allergic eosinophilrich inﬂammation develops in lungs and airways of B cell-deﬁcient mice. J. Exp. Med. 185:885–92 55. Hogan SP, Mishra A, Brandt EB, Royalty MP, Pope SM, et al. 2001. A pathological function for eotaxin and eosinophils in eosinophilic gastrointestinal inﬂammation. Nat. Immunol. 2:353–60 56. Mishra A, Hogan SP, Brandt EB, Rothenberg ME. 2000. Peyer’s patch eosinophils: identiﬁcation, characterization, and regulation by mucosal allergen exposure, interleukin-5, and eotaxin. Blood 96:1538–44 57. Piliponsky AM, Gleich GJ, Bar I, Levi-Schaffer F. 2002. Effects of eosinophils on mast cells: a new pathway for the perpetuation of allergic inﬂammation. Mol. Immunol. 38:1369 58. Solomon A, Aloe L, Pe’er J, Frucht-Pery J, Bonini S, Levi-Schaffer F. 1998. Nerve growth factor is preformed in and activates human peripheral blood eosinophils. J. Allergy Clin. Immunol. 102:454–60 59. Bullock ED, Johnson EM Jr 1996. Nerve growth factor induces the expression of certain cytokine genes and bcl-2 in mast cells. Potential role in survival promotion. J. Biol. Chem. 271:27500–8 60. Horigome K, Bullock ED, Johnson EM Jr. 1994. Effects of nerve growth factor on rat peritoneal mast cells. Survival promotion and immediate-early gene induction. J. Biol. Chem. 269:2695–702 61. Boyce JA, Friend D, Matsumoto R, Austen KF, Owen WF. 1995. Differentiation in vitro of hybrid eosinophil/basophil granulocytes: autocrine function of an eosinophil developmental intermediate. J. Exp. Med. 182:49–57 62. McNagny K, Graf T. 2002. Making eosinophils through subtle shifts in transcription factor expression. J. Exp. Med. 195:F43–47 63. Nerlov C, Graf T. 1998. PU.1 induces myeloid lineage commitment in multipotent hematopoietic progenitors. Genes Dev. 12:2403–12 64. Nerlov C, McNagny KM, Doderlein G, Kowenz-Leutz E, Graf T. 1998. Distinct C/EBP functions are required for eosinophil lineage commitment and maturation. Genes Dev. 12:2413–23 www.annualreviews.org • The Eosinophil

167

ARI

23 February 2006

15:12

65. DeKoter RP, Singh H. 2000. Regulation of B lymphocyte and macrophage development by graded expression of PU.1. Science 288:1439–41 66. Walsh JC, DeKoter RP, Lee HJ, Smith ED, Lancki DW, et al. 2002. Cooperative and antagonistic interplay between PU.1 and GATA-2 in the speciﬁcation of myeloid cell fates. Immunity 17:665–76 67. Du J, Stankiewicz MJ, Liu Y, Xi Q, Schmitz JE, et al. 2002. Novel combinatorial interactions of GATA-1, PU.1, and C/EBP isoforms regulate transcription of the gene encoding eosinophil granule major basic protein. J. Biol. Chem. 277:43481– 94 68. Yu C, Cantor AB, Yang H, Browne C, Wells RA, et al. 2002. Targeted deletion of a high-afﬁnity GATA-binding site in the GATA-1 promoter leads to selective loss of the eosinophil lineage in vivo. J. Exp. Med. 195:1387–95 69. Hirasawa R, Shimizu R, Takahashi S, Osawa M, Takayanagi S, et al. 2002. Essential and instructive roles of GATA factors in eosinophil development. J. Exp. Med. 195:1379– 86 70. Zimmermann N, Daugherty BL, Kavanaugh JL, El-Awar FY, Moulton EA, Rothenberg ME. 2000. Analysis of the CC chemokine receptor 3 gene reveals a complex 5 exon organization, a functional role for untranslated exon 1, and a broadly active promoter with eosinophil-selective elements. Blood 96:2346–54 71. Lopez AF, Begley CG, Williamson DJ, Warren DJ, Vadas MA, Sanderson CJ. 1986. Murine eosinophil differentiation factor. An eosinophil-speciﬁc colony-stimulating factor with activity for human cells. J. Exp. Med. 163:1085–99 72. Rothenberg ME, Pomerantz JL, Owen WF, Avraham S, Soberman RJ, et al. 1988. Characterization of a human eosinophil proteoglycan, and augmentation of its biosynthesis and size by interleukin 3, interleukin 5, and granulocyte/macrophage colony stimulating factor. J. Biol. Chem. 263:13901–8 73. Lopez AF, Sanderson CJ, Gamble JR, Campbell HD, Young IG, Vadas MA. 1988. Recombinant human interleukin 5 is a selective activator of human eosinophil function. J. Exp. Med. 167:219–24 74. Takatsu K, Takaki S, Hitoshi Y. 1994. Interleukin-5 and its receptor system: implications in the immune system and inﬂammation. Adv. Immunol. 57:45–90 75. Vadas M, Lopez A, Gamble J, Khew-Goodall Y, Smith W, et al. 1994. Cytokines and allergy. J. Allergy Clin. Immunol. 94:1289–93 76. Sanderson CJ. 1992. Interleukin-5, eosinophils, and disease. Blood 79:3101–9 77. Collins PD, Marleau S, Grifﬁths-Johnson DA, Jose PJ, Williams TJ. 1995. Cooperation between interleukin-5 and the chemokine eotaxin to induce eosinophil accumulation in vivo. J. Exp. Med. 182:1169–74 78. Dent LA, Strath M, Mellor AL, Sanderson CJ. 1990. Eosinophilia in transgenic mice expressing interleukin 5. J. Exp. Med. 172:1425–31 79. Tominaga A, Takaki S, Koyama N, Katoh S, Matsumoto R, et al. 1991. Transgenic mice expressing a B cell growth and differentiation factor (interleukin 5) develop eosinophilia and autoantibody production. J. Exp. Med. 173:429–39 80. Lee J, McGarry M, Farmer S, Denzler K, Larson K, et al. 1997. Interleukin-5 expression in the lung epithelium of transgenic mice leads to pulmonary changes pathogmomonic of asthma. J. Exp. Med. 185:2143–56 81. Mishra A, Hogan SP, Brandt EB, Wagner N, Crossman MW, et al. 2002. Enterocyte expression of the eotaxin and interleukin-5 transgenes induces compartmentalized dysregulation of eosinophil trafﬁcking. J. Biol. Chem. 277:4406–12

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-05

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Rothenberg

·

Hogan

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-05

ARI

23 February 2006

15:12

82. Foster P, Hogan S, Ramsay A, Matthaei K, Young I. 1996. Interleukin-5 deﬁciency abolishes eosinophilia, airway hyperreactivity and lung damage in a mouse asthma model. J. Exp. Med. 183:195–201 83. Kopf M, Brombacher F, Hodgkin PD, Ramsay AJ, Milbourne EA, et al. 1996. IL-5deﬁcient mice have a developmental defect in CD5+ B-1 cells and lack eosinophilia but have normal antibody and cytotoxic T cell responses. Immunity 4:15–24 84. Owen WF, Rothenberg ME, Petersen J, Weller PF, Silberstein D, et al. 1989. Interleukin 5 and phenotypically altered eosinophils in the blood of patients with the idiopathic hypereosinophilic syndrome. J. Exp. Med. 170:343–48 85. Leckie MJ, ten Brinke A, Khan J, Diamant Z, O’Connor BJ, et al. 2000. Effects of an interleukin-5 blocking monoclonal antibody on eosinophils, airway hyper-responsiveness, and the late asthmatic response. Lancet 356:2144–48 86. Flood-Page P, Phipps S, Menzies-Gow A, Ong YE, Kay AB. 2003. Effect of intravenous administration of an anti-IL-5 (mepolizumab) on allergen-induced tissue eosinophilia, the late-phase allergic reaction and the expression of a marker of repair/remodeling in human atopic subjects. J. Allergy Clin. Immunol. 111:S261 87. Kips JC, O’Connor BJ, Langley SJ, Woodcock AA, Kerstjens HA, et al. 2003. Effect of SCH55700, a humanized anti-human interleukin-5 antibody, in severe persistent asthma: a pilot study. Am. J. Respir. Crit. Care Med. 167:1655–59 88. Mishra A, Hogan SP, Lee JJ, Foster PS, Rothenberg ME. 1999. Fundamental signals that regulate eosinophil homing to the gastrointestinal tract. J. Clin. Invest. 103:1719– 27 89. Humbles AA, Lu B, Friend DS, Okinaga S, Lora J, et al. 2002. The murine CCR3 receptor regulates both the role of eosinophils and mast cells in allergen-induced airway inﬂammation and hyperresponsiveness. Proc. Natl. Acad. Sci. USA 99:1479–84 90. Pope SM, Fulkerson PC, Blanchard C, Akei HS, Nikolaidis NM, et al. 2005. Identiﬁcation of a cooperative mechanism involving interleukin-13 and eotaxin-2 in experimental allergic lung inﬂammation. J. Biol. Chem. 280:13952–61 91. Rothenberg ME, Mishra A, Brandt EB, Hogan SP. 2001. Gastrointestinal eosinophils in health and disease. Adv. Immunol. 78:291–328 92. Rothenberg ME, Owen WF Jr, Silberstein DS, Soberman RJ, Austen KF, Stevens RL. 1987. Eosinophils co-cultured with endothelial cells have increased survival and functional properties. Science 237:645–47 93. Sher A, Coffman RL, Hieny S, Cheever AW. 1990. Ablation of eosinophil and IgE responses with anti-IL-5 or anti-IL-4 antibodies fails to affect immunity against Schistosoma mansoni in the mouse. J. Immunol. 145:3911–16 94. Horie S, Okubo Y, Hossain M, Sato E, Nomura H, et al. 1997. Interleukin-13 but not interleukin-4 prolongs eosinophil survival and induces eosinophil chemotaxis. Intern. Med. 36:179–85 95. Bochner BS, Schleimer RP. 1994. The role of adhesion molecules in human eosinophil and basophil recruitment. J. Allergy Clin. Immunol. 94:427–38 96. Zimmermann N, Hershey GK, Foster PS, Rothenberg ME. 2003. Chemokines in asthma: cooperative interaction between chemokines and IL-13. J. Allergy Clin. Immunol. 111:227– 42 97. Zhu Z, Zheng T, Homer RJ, Kim YK, Chen NY, et al. 2004. Acidic mammalian chitinase in asthmatic Th2 inﬂammation and IL-13 pathway activation. Science 304:1678–82 98. Rankin SM, Conroy DM, Williams TJ. 2000. Eotaxin and eosinophil recruitment: implications for human disease. Mol. Med. Today 6:20–27 www.annualreviews.org • The Eosinophil

169

ARI

23 February 2006

15:12

99. Hogan SP, Koskinen A, Foster PS. 1997. Interleukin-5 and eosinophils induce airway damage and bronchial hyperreactivity during allergic airway inﬂammation in BALB/c mice. Immunol. Cell Biol. 75:284–88 100. Jose PJ, Grifﬁths-Johnson DA, Collins PD, Walsh DT, Moqbel R, et al. 1994. Eotaxin: a potent eosinophil chemoattractant cytokine detected in a guinea pig model of allergic airways inﬂammation. J. Exp. Med. 179:881–87 101. Rothenberg ME, Luster AD, Lilly CM, Drazen JM, Leder P. 1995. Constitutive and allergen-induced expression of eotaxin mRNA in the guinea pig lung. J. Exp. Med. 181:1211–16 102. Zimmermann N, Hogan SP, Mishra A, Brandt EB, Bodette TR, et al. 2000. Murine eotaxin-2: a constitutive eosinophil chemokine induced by allergen challenge and IL-4 overexpression. J. Immunol. 165:5839–46 103. Shinkai A, Yoshisue H, Koike M, Shoji E, Nakagawa S, et al. 1999. A novel human CC chemokine, eotaxin-3, which is expressed in IL-4-stimulated vascular endothelial cells, exhibits potent activity toward eosinophils. J. Immunol. 163:1602–10 104. Murphy PM. 1994. The molecular biology of leukocyte chemoattractant receptors. Annu. Rev. Immunol. 12:593–633 105. Ponath PD, Qin S, Post TW, Wang J, Wu L, et al. 1996. Molecular cloning and characterization of a human eotaxin receptor expressed selectively on eosinophils. J. Exp. Med. 183:2437–48 106. Daugherty BL, Siciliano SJ, Demartino JA, Malkowitz L, Sirotina A, Springer MS. 1996. Cloning, expression, and characterization of the human eosinophil eotaxin receptor. J. Exp. Med. 183:2349–54 107. Pope SM, Zimmermann N, Stringer KF, Karow ML, Rothenberg ME. 2005. The eotaxin chemokines and CCR3 are fundemental regulators of allergen-induced pulmonary eosinophilia. J. Immunol. 175:5341–50 108. Fulkerson PC, Zhu H, Williams DA, Zimmermann N, Rothenberg ME. 2005. CXCL9 inhibits eosinophil responses by a CCR3- and Rac2-dependent mechanism. Blood 106:436– 43 109. Rothenberg ME, MacLean JA, Pearlman E, Luster AD, Leder P. 1997. Targeted disruption of the chemokine eotaxin partially reduces antigen-induced tissue eosinophilia. J. Exp. Med. 185:785–90 110. Gonzalo JA, Lloyd CM, Wen D, Albar JP, Wells TN, et al. 1998. The coordinated action of CC chemokines in the lung orchestrates allergic inﬂammation and airway hyperresponsiveness. J. Exp. Med. 188:157–67 111. Ma W, Bryce PJ, Humbles AA, Laouini D, Yalcindag A, et al. 2002. CCR3 is essential for skin eosinophilia and airway hyperresponsiveness in a murine model of allergic skin inﬂammation. J. Clin. Invest. 109:621–28 112. Nakamura H, Luster AD, Nakamura T, In KH, Sonna LA, et al. 2001. Variant eotaxin: its effects on the asthma phenotype. J. Allergy Clin. Immunol. 108:946–53 113. Chae SC, Park YR, Oh GJ, Lee JH, Chung HT. 2005. The suggestive association of eotaxin-2 and eotaxin-3 gene polymorphisms in Korean population with allergic rhinitis. Immunogenetics 56:760–64 114. Menzies-Gow A, Ying S, Sabroe I, Stubbs VL, Soler D, et al. 2002. Eotaxin (CCL11) and eotaxin-2 (CCL24) induce recruitment of eosinophils, basophils, neutrophils, and macrophages as well as features of early- and late-phase allergic reactions following cutaneous injection in human atopic and nonatopic volunteers. J. Immunol. 169:2712– 18

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115. Artis D, Humphreys NE, Potten CS, Wagner N, Muller W, et al. 2000. β7 integrindeﬁcient mice: delayed leukocyte recruitment and attenuated protective immunity in the small intestine during enteric helminthic infection. Eur. J. Immunol. 30:1656–64 116. Pretolani M, Rufﬁe C, Lapa e Silva JR, Joseph D, Lobb RR, Vargaftig BB. 1994. Antibody to very late activation antigen 4 prevents antigen-induced bronchial hyperreactivity and cellular inﬁltration in the guinea pig airways. J. Exp. Med. 180:795–805 117. Nakajima H, Sano H, Nishimura T, Yoshida S, Iwamoto I. 1994. Role of vascular cell adhesion molecule 1/very late activation antigen 4 and intercellular adhesion molecule 1/lymphocyte function-associated antigen 1 interactions in antigen-induced eosinophil and T cell recruitment into the tissue. J. Exp. Med. 179:1145–54 118. Gonzalo JA, Lloyd CM, Kremer L, Finger E, Martinez AC, et al. 1996. Eosinophil recruitment to the lung in a murine model of allergic inﬂammation. The role of T cells, chemokines, and adhesion receptors. J. Clin. Invest. 98:2332–45 119. Jia GQ, Gonzalo JA, Hidalgo A, Wagner D, Cybulsky M, Gutierrez-Ramos JC. 1999. Selective eosinophil transendothelial migration triggered by eotaxin via modulation of Mac-1/ICAM-1 and VLA-4/VCAM-1 interactions. Int. Immunol. 11:1–10 120. Tager AM, Dufour JH, Goodarzi K, Bercury SD, von Andrian UH, Luster AD. 2000. BLTR mediates leukotriene B4 -induced chemotaxis and adhesion and plays a dominant role in eosinophil accumulation in a murine model of peritonitis. J. Exp. Med. 192:439–46 121. Hirai H, Tanaka K, Yoshie O, Ogawa K, Kenmotsu K, et al. 2001. Prostaglandin D2 selectively induces chemotaxis in T helper type 2 cells, eosinophils, and basophils via seven-transmembrane receptor CRTH2. J. Exp. Med. 193:255–61 122. O’Reilly M, Alpert R, Jenkinson S, Gladue RP, Foo S, et al. 2002. Identiﬁcation of a histamine H4 receptor on human eosinophils—role in eosinophil chemotaxis. J. Recept. Signal Transduct. Res. 22:431–48 123. Behm CA, Ovington KS. 2000. The role of eosinophils in parasitic helminth infections: insights from genetically modiﬁed mice. Parasitol. Today 16:202–9 124. Korenaga M, Hitoshi Y, Yamaguchi N, Sato Y, Takatsu K, Tada I. 1991. The role of interleukin-5 in protective immunity to Strongyloides venezuelensis infection in mice. Immunology 72:502–7 125. Erickson LD, Foy TM, Waldschmidt TJ. 2001. Murine B1 B cells require IL-5 for optimal T cell-dependent activation. J. Immunol. 166:1531–39 126. Bischoff SC, Brunner T, De Weck AL, Dahinden CA. 1990. Interleukin 5 modiﬁes histamine release and leukotriene generation by human basophils in response to diverse agonists. J. Exp. Med. 172:1577–82 127. Hakonarson H, Maskeri N, Carter C, Chuang S, Grunstein MM. 1999. Autocrine interaction between IL-5 and IL-1β mediates altered responsiveness of atopic asthmatic sensitized airway smooth muscle. J. Clin. Invest. 104:657–67 128. Gurish MF, Humbles A, Tao H, Finkelstein S, Boyce JA, et al. 2002. CCR3 is required for tissue eosinophilia and larval cytotoxicity after infection with Trichinella spiralis. J. Immunol. 168:5730–36 129. Simons JE, Rothenberg ME, Lawrence RA. 2005. Eotaxin-1-regulated eosinophils have a critical role in innate immunity against experimental Brugia malayi infection. Eur. J. Immunol. 35:189–97 130. Adamko DJ, Yost BL, Gleich GJ, Fryer AD, Jacoby DB. 1999. Ovalbumin sensitization changes the inﬂammatory response to subsequent parainﬂuenza infection: Eosinophils mediate airway hyperresponsiveness, M2 muscarinic receptor dysfunction and antiviral effects. J. Exp. Med. 1999:1465–77 www.annualreviews.org • The Eosinophil

171

ARI

23 February 2006

15:12

131. Jacoby DB, Gleich GJ, Fryer AD. 1993. Human eosinophil major basic protein is an endogenous allosteric antagonist at the inhibitory muscarinic M2 receptor. J. Clin. Invest. 91:1314–18 132. Bandeira-Melo C, Bozza PT, Weller PF. 2002. The cellular biology of eosinophil eicosanoid formation and function. J. Allergy Clin. Immunol. 109:393–400 133. Hamelmann E, Gelfand EW. 2001. IL-5-induced airway eosinophilia—the key to asthma? Immunol. Rev. 179:182–91 134. Denzler KL, Farmer SC, Crosby JR, Borchers MT, Cieslewicz G, et al. 2000. Eosinophil major basic protein-1 does not contribute to allergen-induced airways pathologies in mouse models of asthma. J. Immunol. 165:5509–17 135. Boyce JA, Austen KF. 2005. No audible wheezing: nuggets and conundrums from mouse asthma models. J. Exp. Med. 201:1869–73 136. Hogan S, Koskinen A, Foster P. 1997. Interleukin-5 and eosinophils induce airway damage and bronchial hyperreactivity during allergic airway inﬂammation in BALB/c mice. Immunol. Cell Biol. 75:284–88 137. Foster P, Mould A, Yang M, Mackenzie J, Mattes J, et al. 2001. Elemental signals regulating eosinophil accumulation in the lung. Immunol. Rev. 179:173–81 138. Hogan SP, Koskinen A, Matthaei KI, Young IG, Foster PS. 1998. Interleukin-5-producing CD4+ T cells play a pivotal role in aeroallergen-induced eosinophilia, bronchial hyperreactivity, and lung damage in mice. Am. J. Respir. Crit. Care Med. 157:210–18 139. Mattes J, Yang M, Mahalingam S, Kuehr J, Webb DC, et al. 2002. Intrinsic defect in T cell production of interleukin (IL)-13 in the absence of both IL-5 and eotaxin precludes the development of eosinophilia and airways hyperreactivity in experimental asthma. J. Exp. Med. 195:1433–44 140. Corry D, Folkesson H, Warnock M, Erle D, Matthay M, et al. 1996. Interleukin 4, but not interleukin 5 or eosinophils, is required in a murine model of acute airway hyperreactivity. J. Exp. Med. 183:109–17 141. Hamelmann E, Oshiba A, Loader J, Larsen GL, Gleich G, et al. 1997. Antiinterleukin-5 antibody prevents airway hyperresponsiveness in a murine model of airway sensitization. Am. J. Respir. Crit. Care Med. 155:819–25 142. Lee NA, McGarry MP, Larson KA, Horton MA, Kristensen AB, Lee JJ. 1997. Expression of IL-5 in thymocytes/T cells leads to the development of a massive eosinophilia, extramedullary eosinophilopoiesis, and unique histopathologies. J. Immunol. 158:1332– 44 143. Brightling CE, Bradding P, Symon FA, Holgate ST, Wardlaw AJ, Pavord ID. 2002. Mastcell inﬁltration of airway smooth muscle in asthma. N. Engl. J. Med. 346:1699–705 144. Justice JP, Borchers MT, Crosby JR, Hines EM, Shen HH, et al. 2003. Ablation of eosinophils leads to a reduction of allergen-induced pulmonary pathology. Am. J. Physiol. Lung Cell Mol. Physiol. 284:L169–78 145. O’Byrne PM, Inman MD, Parameswaran K. 2001. The trials and tribulations of IL-5, eosinophils, and allergic asthma. J. Allergy Clin. Immunol. 108:503–8 146. Flood-Page P, Menzies-Gow A, Phipps S, Ying S, Wangoo A, et al. 2003. Anti-IL-5 treatment reduces deposition of ECM proteins in the bronchial subepithelial basement membrane of mild atopic asthmatics. J. Clin. Invest. 112:1029–36 147. Green RH, Brightling CE, McKenna S, Hargadon B, Parker D, et al. 2002. Asthma exacerbations and sputum eosinophil counts: a randomised controlled trial. Lancet 360:1715–21 148. Lee JJ, Dimina D, Macias MP, Ochkur SI, McGarry MP, et al. 2004. Deﬁning a link with asthma in mice congenitally deﬁcient in eosinophils. Science 305:1773–76

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149. Humbles AA, Lloyd CM, McMillan SJ, Friend DS, Xanthou G, et al. 2004. A critical role for eosinophils in allergic airways remodeling. Science 305:1776–79 150. Rothenberg ME. 2004. Eosinophilic gastrointestinal disorders (EGID). J. Allergy Clin. Immunol. 113:11–28 151. Mishra A, Rothenberg ME. 2003. Intratracheal IL-13 induces eosinophilic esophagitis by an IL-5, eotaxin-1, and STAT6-dependent mechanism. Gastroenterology 125:1419– 27 152. Mishra A, Hogan SP, Brandt EB, Rothenberg ME. 2001. An etiological role for aeroallergens and eosinophils in experimental esophagitis. J. Clin. Invest. 107:83–90 153. Forbes E, Smart VE, D’Aprile A, Henry P, Yang M, et al. 2004. T helper-2 immunity regulates bronchial hyperresponsiveness in eosinophil-associated gastrointestinal disease in mice. Gastroenterology 127:105–18 154. Cools J, DeAngelo DJ, Gotlib J, Stover EH, Legare RD, et al. 2003. A tyrosine kinase created by fusion of the PDGFRA and FIP1L1 genes as a therapeutic target of imatinib in idiopathic hypereosinophilic syndrome. N. Engl. J. Med. 348:1201–14 155. Cools J, Stover EH, Wlodarska I, Marynen P, Gilliland DG. 2004. The FIP1L1-PDGFRα kinase in hypereosinophilic syndrome and chronic eosinophilic leukemia. Curr. Opin. Hematol. 11:51–57 156. Gleich GJ, Leiferman KM, Pardanani A, Tefferi A, Butterﬁeld JH. 2002. Treatment of hypereosinophilic syndrome with imatinib mesilate. Lancet 359:1577–78 157. Stellato C, Matsukura S, Fal A, White J, Beck LA, et al. 1999. Differential regulation of epithelial-derived C-C chemokine expression by IL-4 and the glucocorticoid budesonide. J. Immunol. 163:5624–32 158. Schleimer RP, Bochner BS. 1994. The effect of glucocorticoids on human eosinophils. J. Allergy Clin. Immunol. 94:1202–13 159. Barnes PJ, Adcock IM. 1995. Steroid resistance in asthma. QJM 88:455–68 160. Aldebert D, Lamkhioued B, Desaint C, Gounni AS, Goldman M, et al. 1996. Eosinophils express a functional receptor for interferon α: inhibitory role of interferon α on the release of mediators. Blood 87:2354–60 161. Bankers-Fulbright JL, Kephart GM, Loegering DA, Bradford AL, Okada S, et al. 1998. Sulfonylureas inhibit cytokine-induced eosinophil survival and activation. J. Immunol. 160:5546–53 162. Hunt LW, Frigas E, Butterﬁeld JH, Kita H, Blomgren J, et al. 2004. Treatment of asthma with nebulized lidocaine: a randomized, placebo-controlled study. J. Allergy Clin. Immunol. 113:853–59 163. Kane GC, Pollice M, Kim CJ, Cohn J, Dworski RT, et al. 1996. A controlled trial of the effect of the 5-lipoxygenase inhibitor, zileuton, on lung inﬂammation produced by segmental antigen challenge in human beings. J. Allergy Clin. Immunol. 97:646– 54 164. Gaddy JN, Margolskee DJ, Bush RK, Williams VC, Busse WW. 1992. Bronchodilation with a potent and selective leukotriene D4 (LTD4) receptor antagonist (MK-571) in patients with asthma. Am. Rev. Respir. Dis. 146:358–63 165. Snyman JR, Sommers DK, Gregorowski MD, Boraine H. 1992. Effect of cetirizine, ketotifen and chlorpheniramine on the dynamics of the cutaneous hypersensitivity reaction: a comparative study. Eur. J. Clin. Pharmacol. 42:359–62 166. Redier H, Chanez P, De Vos C, Rifai N, Clauzel AM, et al. 1992. Inhibitory effect of cetirizine on the bronchial eosinophil recruitment induced by allergen inhalation challenge in allergic patients with asthma. J. Allergy Clin. Immunol. 90:215–24 www.annualreviews.org • The Eosinophil

173

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167. Rand TH, Lopez AF, Gamble JR, Vadas MA. 1988. Nedocromil sodium and cromolyn (sodium cromoglycate) selectively inhibit antibody-dependent granulocyte-mediated cytotoxicity. Int. Arch. Allergy Appl. Immunol. 87:151–58 168. Wegner CD, Gundel RH, Reilly P, Haynes N, Letts LG, Rothlein R. 1990. Intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of asthma. Science 247:456–59 169. Weg VB, Williams TJ, Lobb RR, Nourshargh S. 1993. A monoclonal antibody recognizing very late activation antigen-4 inhibits eosinophil accumulation in vivo. J. Exp. Med. 177:561–66 170. Kuijpers TW, Mul EP, Blom M, Kovach NL, Gaeta FC, et al. 1993. Freezing adhesion molecules in a state of high-avidity binding blocks eosinophil migration. J. Exp. Med. 178:279–84 171. von Andrian UH, Engelhardt B. 2003. α4 integrins as therapeutic targets in autoimmune disease. N. Engl. J. Med. 348:68–72 172. Mauser PJ, Pitman AM, Fernandez X, Foran SK, Adams GK 3rd, et al. 1995. Effects of an antibody to interleukin-5 in a monkey model of asthma. Am. J. Respir. Crit. Care Med. 152:467–72 173. Egan RW, Athwahl D, Chou CC, Emtage S, Jehn CH, et al. 1995. Inhibition of pulmonary eosinophilia and hyperreactivity by antibodies to interleukin-5. Int. Arch. Allergy Immunol. 107:321–22 174. Blanchard C, Mishra A, Saito-Akei H, Monk P, Anderson I, Rothenberg ME. 2005. Inhibition of human interleukin-13-induced respiratory and oesophageal inﬂammation by anti-human-interleukin-13 antibody (CAT-354). Clin. Exp. Allergy 35:1096–103 175. Nutku E, Aizawa H, Hudson SA, Bochner BS. 2003. Ligation of Siglec-8: a selective mechanism for induction of human eosinophil apoptosis. Blood 101:5014–20

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Annual Review of Immunology Volume 24, 2006

Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:147-174. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Human T Cell Responses Against Melanoma Thierry Boon,1,2 Pierre G. Coulie,2 Benoˆıt J. Van den Eynde,1,2 and Pierre van der Bruggen1,2 1

Ludwig Institute for Cancer Research, Brussels Branch, B-1200 Brussels, Belgium; email: [email protected], [email protected], [email protected]

2

Cellular Genetics Unit, Universit´e de Louvain, B-1200 Brussels, Belgium; email: [email protected]

Annu. Rev. Immunol. 2006. 24:175–208 First published online as a Review in Advance on December 1, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090733 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0175$20.00

Key Words vaccination, cancer, T lymphocyte

Abstract Many antigens recognized by autologous T lymphocytes have been identiﬁed on human melanoma. Melanoma patients usually mount a spontaneous T cell response against their tumor. But at some point, the responder T cells become ineffective, probably because of a local immunosuppressive process occurring at the tumor sites. Therapeutic vaccination of metastatic melanoma patients with these antigens is followed by tumor regressions only in a small minority of the patients. The T cell responses to the vaccines show correlation with the tumor regressions. The local immunosuppression may be the cause of the lack of vaccination effectiveness that is observed in most patients. In patients who do respond to the vaccine, the antivaccine T cells probably succeed in reversing focally this immunosuppression and trigger a broad activation of other antitumor T cells, which proceed to destroy the tumor.

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INTRODUCTION

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

MAGE: melanoma antigen-encoding gene

The large diversity of mouse tumor systems has predictably led to the elucidation of a wide variety of interesting mechanisms of tumor attack by the immune system, as well as of mechanisms of tumor resistance and escape (1). We suspect that for each type of human tumor, there is a subset of this wide array of processes that critically inﬂuences the tumor’s evolution and its response to immunotherapy. This crucial set of processes can only be identiﬁed by studying in depth each type of human tumor. With a few exceptions, we have restricted this review to observations made on human melanoma patients regarding antitumoral T cell responses that occur either spontaneously or following vaccination with tumor antigens. Even with this limitation, this brief review is far from exhaustive.

ANTIGENS RECOGNIZED ON MELANOMA BY AUTOLOGOUS T CELLS Genes Encoding Melanoma Antigens Human tumors bear antigens that can be recognized by autologous T lymphocytes. Some of these antigens appear to be tumor speciﬁc, whereas others are also present on normal tissues. Two main genetic mechanisms produce tumor-speciﬁc antigens: (a) Point mutations occurring in tumor cells can generate antigenic peptides either by enabling them to bind to the groove of MHC molecules or by generating new epitopes on these peptides. (b) In tumor cells, a gene is expressed that is silent in normal cells. On human melanoma, autologous T cells recognize tumor-speciﬁc antigens produced by both mechanisms: the antigens caused by mutational events and those encoded by cancer-germline genes. Somewhat surprisingly, in view of what is expected of natural tolerance, it is also possible to observe or generate autologous T cell responses against differentiation antigens common to melanoma and normal melanocytes. 176

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Antigens encoded by cancer-germline genes. A small number of gene families are expressed in male germline cells and in many tumors, but not in other normal adult tissues. Because male germline cells do not express human leukocyte antigen (HLA) molecules on their surface, they do not express antigens that can be recognized by T cells (2). As a result, the antigens encoded by these cancergermline genes are strictly tumor-speciﬁc T cell targets. These antigens are shared by many tumors. The prototype of the cancer-germline genes is the melanoma antigen-encoding (MAGE) gene family (3–5). This family is composed of 24 genes that range across three subfamilies: MAGE-A, -B, and -C, located in three different regions of the X chromosome. The expression of MAGE genes in tumors appears to be triggered by the demethylation of their promoter, apparently as a consequence of the widespread demethylation process that occurs in many tumors (6). Six additional MAGE gene families have been identiﬁed in the human genome (5, 7). These genes are expressed in normal tissues. However, because of their low similarity with the sequences of the MAGE-A, -B, and C family, not a single MAGE-A, -B, or -C antigenic peptide is encoded by one of the ubiquitously expressed MAGE genes, leaving intact the strict tumor speciﬁcity of the antigens encoded by the MAGE-A, -B, and -C genes. Other important cancer-germline gene families are the BAGE (8), GAGE (9), LAGE/NY-ESO-1 (10, 11), and SSX families (12, 13). Table 1 describes the main cancergermline genes and antigens that are discussed in this review. Investigators often state that antigens encoded by cancer-germline genes represent “self ” antigens and that tolerance must therefore be broken in the course of immunization against these antigens. In one sense, cancer-germline genes are indeed self in that they are normal genes of the human genome. However, the strict tumoral speciﬁcity of the

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Main antigenic peptides used to vaccinate melanoma patientsa

Gene

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Frequency of expression in metastatic melanoma

Presenting HLA

Peptide sequence

Position in the protein

MAGE-A1

46%

A2 A3

MAGE-A3

74%

A1 DP4

MAGE-A4

28%

MAGE-A10

47%

MAGE-A12

62%

Cw7

MAGE-C2

59%

A2

ALKDVEERV

NY-ESO-1/LAGE-2

28%

A2

SLLMWITQ(A,I,L,V)

Tyrosinase

>90%

A2

YMDGTMSQV

Melan-A/MART-1

>90%

A2

ELAGIGILTV

26–35 (A27L)

gp100

>90%

A2

IMDQVPFSV

209–217 (T210M)

50%

A2

VLPDVFIRC

(intron)

GNT-V a

KVLEYVIKV SLFRAVITK

278–286 96–104

EVDPIGHLY KKLLTQHFVQENYLEY

168–176 243–258

A2

GVYDGREHTV

230–239

A2

GLYDGMEHL

254–262

VRIGHLYIL

170–178 336–344 157–165 (C165A,I,L,V) 369–377

More information can be found at http://www.cancerimmunity.org/peptidedatabase/Tcellepitopes.htm.

antigens is hardly conveyed by the word self. The critical issue is whether there exists some measurable degree of natural immunological tolerance against these antigens. In our opinion this question has not yet been answered. One ought to be able to examine this issue with the mouse P1A gene, which codes for a mastocytoma P815 antigen and is also a cancer-germline gene (14). Normal DBA/2 mice mount a CD8 response to this antigen. Transgenic mice that express the P1A gene in all their tissues are clearly incapable of a cytotoxic T lymphocyte (CTL) response to the relevant antigens (15). But the crucial question has not yet been answered, namely, whether knockout mice will mount anti-P1A T cell responses more readily than normal mice. We expect these mice to be available in the near future. For gene P1A, as well as for MAGE genes, Kyewski and coworkers (16, 17) have shown that some sporadic gene expression occurs in medullary thymic epithelial cells, thereby opening the possibility that some form of thymic tolerance might exist. Antigens resulting from point mutations. Many antigens recognized by T cells on human tumors are encoded by a sequence con-

taining a point mutation that originated in the tumor cells (18). Many of the point mutations that have been identiﬁed because they generated a tumor antigen appear to have an oncogenic function. One mutation found in melanoma in a cyclin-dependent kinase molecule (CDK4) was also found in a familial form of melanoma (19, 20). The mutated antigens that have been identiﬁed after autologous mixed lymphocyte/tumor cell cultures (MLTC) were invariably found to result from mutations that are present on a single tumor or on very few. Thus, these mutated antigens are strictly tumor speciﬁc but not shared. This limits their use as potential vaccines. It is remarkable that MLTC have not produced T cells recognizing antigens encoded by mutated regions that are very frequent in cancer cells, such as mutated ras or p53. However, a mutated N-ras antigen was found to be recognized by tumor-inﬁltrating lymphocytes (TILs) (21). Promising candidates for shared mutated antigens are encoded by B-RAF, a member of the mitogen-activated protein (MAP) kinase cascade. The same activating mutation of B-RAF is found in more than 60% of

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TIL: tumor-inﬁltrating lymphocyte

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melanomas (22). Recently, CD4 T cells were shown to recognize a B-Raf peptide encoded by the mutated region. These CD4 T cells also recognized cells expressing the mutated B-RAF gene (23).

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Antigens encoded by genes overexpressed in tumors. The gene PRAME was identiﬁed as coding for a melanoma antigen recognized by autologous CTL. It is expressed at a high level in almost all melanomas and in many other tumors (24, 25). Survivin, an antiapoptotic protein of the inhibitor of apoptosis protein (IAP) family, was reported to be overexpressed in tumors, and melanoma cells are lysed by antisurvivin CTL (26, 27). However, no comparison of the level of survivin in tumor cells and in proliferating normal cells has been published. Telomerase has been suggested as a good target for immunotherapy, as its presence is essential for the proliferation of tumor cells. An antigenic peptide encoded by telomerase was recognized by CTL (28). However, other reports claimed that CTL of patients vaccinated with the antigenic peptide recognized cells pulsed with the peptide but did not recognize tumor cells expressing telomerase (29, 30). The antigens encoded by these genes are shared between many tumors, but they are not tumor speciﬁc, and the risk of vaccinating with these antigens must be weighed carefully.

Antigens encoded by differentiation genes. Melanoma patients have T lymphocytes capable of recognizing antigens encoded by normal melanocytic differentiation genes, such as those encoded by tyrosinase, Melan-A/Mart-1, gp100/pMel17, tyrosinase-related protein (TRP)-1, and TRP-2 (31–34). It was surprising to observe that natural tolerance against such antigens was not tighter. Vaccination with such antigens can produce vitiligo, i.e., elimination of normal melanocytes in some areas of the skin (35, 36). 178

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Processing of Tumor Antigens Antigens presented by HLA class I molecules. Most tumor antigens presented by class I molecules appear to be processed in tumor cells by the normal pathway of protein destruction by the proteasome, followed by the transport of the produced peptides into the endoplasmic reticulum by TAP (transporters associated with antigen presentation) proteins. The antigenic peptides then bind into the groove of MHC class I molecules and their N-terminal end can undergo a ﬁnal trimming by aminopeptidases. It is becoming increasingly clear that some antigenic peptides are processed much better in cells harboring immunoproteasomes, whereas others are processed better in cells with standard proteasomes (37–39). The endopeptidase activities of these proteasomes differ by their speciﬁcity, the immunoproteasome cleaving better after hydrophobic residues and the standard proteasome after acidic residues. This can affect not only the cleavage producing the proper C-terminus, but also the degree of destruction of the antigenic peptide by internal cleavage. Such proteasome preferences could have consequences for the CTL attack of tumor cells. In a noninﬂammatory environment, tumor cells are expected to express only standard proteasomes and, accordingly, only the tumor antigens produced by these proteasomes. However, upon immune attack of the tumor, IFN-γ ought to be produced, resulting in the replacement of standard proteasomes by immunoproteasomes, thereby eliminating those antigens that are produced by the former and replacing them by those produced by the latter. This could render the tumor cells resistant to some CTL. However, many tumor antigens are expressed on most tumors, and a number of them are processed by both types of proteasomes. Proteasome preferences also have implications for antitumoral vaccination. The mature dendritic cells (DCs), which present antigens to T lymphocytes in lymph nodes, express immunoproteasomes (37, 40,

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41). This means that for some antigens that are processed only by standard proteasomes, it is preferable to vaccinate with the peptide or with the recombinant virus carrying a minigene coding for the peptide rather than with the protein or a recombinant virus coding for the whole protein. A substantial minority (∼20%) of the tumor antigens are the result of a deviation from the genetic expression process that produces the standard protein. The deviation can occur at the transcription stage, as is the case for the tumor-speciﬁc antigen that results from the action of a cryptic promoter located in an intron of gene GNT-V (42). This produces a protein encoded by an intronic sequence and out-of-frame exonic sequences. Such a cryptic promoter can be located on the reverse strand of an intron and produce an antisense transcript that also produces antigenic peptides (43). A second mechanism is the translation of incompletely spliced messages, resulting in antigenic peptides spanning an exon and an intronic region (44–47). The deviation can also occur at the translational stage, with the translation of small alternative open reading frames (48–54). Posttranslational modiﬁcation such as deamidation of asparagine can also occur (55). A recent development in the area of these atypical antigens was the startling observation that peptide sequences that are noncontiguous on a protein sequence can be joined together to form an antigenic peptide (56, 57). This peptide splicing occurs in the proteasome, where one free cleavage product forms a peptide bond with another one that is still bound to the catalytic threonine (57). This enables the splicing of peptide fragments located distantly on the protein either in the direct order or in the reverse order.

Antigens presented by HLA class II molecules. Constitutive expression of MHC class II molecules is normally restricted to DCs, macrophages, and B lymphocytes. But it also occurs frequently in several types of tumor, particularly melanoma (58). Moreover,

tumors usually express class II molecules when IFN-γ is present (59). Two different pathways lead to presentation of antigenic peptides on HLA class II molecules. Through the endocytic pathway, the presenting cells can internalize external cell debris or proteins, as well as their own secreted and membrane proteins (60–62). Through the endogenous pathway, intracellular proteins reach the endosomal compartment, either after passage through the endoplasmic reticulum for proteins having appropriate signal and targeting sequences, or by direct inclusion of cytosolic elements in endosomes (autophagy) (63–65). The melanocyte protein tyrosinase follows the second pathway, being directed through its targeting sequence to melanosomes, organelles that are derived from the endosomes (66, 67). Protein MAGE-A3 does not contain a signal sequence. Accordingly, CD4 T cell clones recognizing two MAGE-A3 peptides presented by HLA-DR13 molecules could not recognize DR13 melanoma cells expressing MAGE-A3 (68). Nevertheless, three other MAGE-A3 antigenic peptides were found to be recognized by CD4 T cell clones on HLADP4, DR1, or DR11 tumor cells expressing MAGE-A3 (69–71). This presumably occurs through autophagy. An optimal anticancer vaccine may require the cooperation of CD4+ Th cells and CD8+ CTLs. Various strategies have been explored to obtain presentation of both HLA class I– and class II–restricted peptides derived from the same vaccine candidate. One interesting possibility for cytosolic proteins is the manipulation of the HLA class II presentation pathway. This manipulation can be achieved by using vectors where the protein-coding sequence is linked to a sequence encoding the targeting motif of an endosomal or lysosomal protein, such as the invariant chain, the lysosome-associated membrane protein-1 (LAMP-1), or DC-LAMP (72–78). Interestingly, targeting MAGE-A3 to the HLA class II–processing pathway induced concurrent presentation of class I– and class II–restricted

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LAMP: lysosome-associated membrane protein

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DriPs: defective ribosomal products

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peptides to CD8+ and CD4+ T cells, respectively. We observed that the HLA class I presentation by the DCs electroporated with the chimeric constructs was more efﬁcient than that by peptide-pulsed DCs. The reason for this is not clear. It could be due to the translation of an abnormal protein, inducing an increased formation of defective ribosomal products (DriPs) that are addressed to the proteasome (79).

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Optimization of Peptide Immunogenicity For immunization with antigenic peptides, an important line of research has been the substitution of some amino acids to obtain peptides with stronger binding to HLA molecules. This has been applied ﬁrst to the gp100209−217 peptide, ITDQVPFSV, which is presented by HLA-A2. The threonine in position 2 was replaced by methionine. The modiﬁed peptide was much more immunogenic in HLA-A2 transgenic mice (80, 81). Among T lymphocytes collected from patients vaccinated with the modiﬁed peptide, some recognized both the modiﬁed and the normal peptide, whereas others recognized only the modiﬁed peptide (82, 83). This approach was also applied to MelanA.A2 peptide EAAGIGILTV, where the alanine in position 2 was replaced by a leucine residue (84, 85). This increased the ability of the peptide to trigger CTL lysis and to stimulate PBL in vitro (86). T lymphocytes from melanoma patients were isolated with HLA tetramers containing the modiﬁed peptide. Some of these T lymphocytes recognized targets presenting the normal antigen, but others recognized only targets pulsed with the modiﬁed peptide (87). However, almost all T cells collected from patients vaccinated with the modiﬁed peptide recognized melanoma cell lines (88). For the NY-ESO-1 peptide SLLMWITQC, the C-terminal residue was replaced by valine, leucine, or isoleucine. Here, again, these peptides showed better 180

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binding to HLA and were more readily recognized by CTL (89, 90). In conclusion, the usefulness of vaccination with modiﬁed peptides showing increased immunogenicity has to be weighed carefully against the risk of having a signiﬁcant subset of responder T cells that fail to recognize the tumor cells.

SPONTANEOUS T CELL RESPONSES Do spontaneous antitumoral T cell responses exist? Are they beneﬁcial to some patients? To the extent that they exist, why do they fail to reject the tumors in many instances? Do they inﬂuence the state of the tumor and its sensitivity to immunotherapy by selecting for resistance?

Existence of Spontaneous T Cell Responses Evidence that strongly suggested the existence of a spontaneous antitumoral T cell response in melanoma patients predated the identiﬁcation of human tumor antigens. Rosenberg and associates (91) carried out a considerable amount of work with TILs. Single cell suspensions derived from metastases, including invaded lymph nodes that contained lymphocytes and an undeﬁned number of tumor cells, were cultured from 30–60 days in the presence of IL-2. This procedure often produced as many as 1010 –1011 T lymphocytes. Many of these lymphocyte preparations were found to exert cytolytic activity on fresh tumor cell preparations, albeit under conditions in which stringent speciﬁcity controls could not be realized (92). Adoptive transfer of these lymphocytes together with high-dose IL-2 treatment produced objective tumor responses in a higher proportion of the patients than that observed with IL-2 treatment alone (91, 93). Our early work on the identiﬁcation of antigens recognized on melanoma by autologous T cells provided unquestionable proof of the antitumoral speciﬁcity of the T cells

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found in the blood of melanoma patients (4). However, these patients had received various immunotherapy regimens, and no frequency comparison was made between normal and cancerous individuals. So, even though in retrospect we believe that, with the techniques available at that time, we would not have obtained antitumor T cell clones unless a response had occurred in these patients, our work did not clearly demonstrate the existence of such a spontaneous response. At that time, we also carried out a systematic evaluation of the antitumoral T cell response of melanoma patients, using autologous MLTC under limiting dilution conditions (94). Microcultures were considered to contain antitumor effectors when they exerted a lytic activity that was much higher on autologous tumor cells than on K562 or autologous T cell blasts. Frequencies of antitumor T cells ranging from 3 × 10−5 to 10−3 of CD8 T cells were observed in a group of six patients. The ability to conﬁrm the existence of a spontaneous antitumoral T cell response was hampered by the impossibility of providing a baseline with parallel tests run with autologous cell lines on noncancerous patients, and by the lack of knowledge of the target antigens. Parmiani and collaborators (95) compared frequencies of antitumor T cells in the blood and in metastatic sites (invaded lymph nodes and subcutaneous tumors). Frequencies ranging from 2 × 10−3 to 10−4 of CD8+ T cells were observed in the blood. In subcutaneous tumors the frequencies observed were about tenfold higher (95). They also examined melanoma metastases and found signiﬁcant differences in the frequencies of some T cell receptor (TCR)-V genes in metastases relative to the blood, suggesting that some T cells were enriched in the tumors (96). Analysis of TCR sequences indicated that some TCRs were present at high frequency. More recently, a limiting dilution analysis of tumor-speciﬁc CTLp in the blood was carried out on six melanoma patients before

vaccination with various antigens. All patients had high frequencies of antitumor CTLp ranging from 6 × 10−5 to 2 × 10−3 of the CD8 T cells (97). The antitumoral speciﬁcity of these responses now appears ﬁrmly established because in one patient the response was analyzed at a clonal level, and almost all the clones were found to be directed against antigens encoded either by the cancer-germline gene MAGE-C2 or by the melanocyte differentiation gene gp100. Letsch et al. (98) found in an ex vivo ELISPOT test that 4/5 melanoma patients had a large number of T cells that produced IFN-γ upon stimulation with an autologous cell line, with frequencies ranging from 5 × 10−3 to 10−4 of CD8 T cells.

TCR: T cell receptor CTLp: cytotoxic T lymphocyte precursor

Spontaneous responses against tumor antigens encoded by cancer-germline genes. The background level of T cell precursors against MAGE-encoded antigens in normal donors appears to be situated ∼4 × 10−7 of CD8+ T cells (see below) (99). Spontaneous (i.e., prevaccination) ampliﬁcation of anti-MAGE-3.A1 CD8 T cells was observed in a very small proportion of metastatic melanoma patients: 2/45 (100, 101). Similar observations were made for other tumorspeciﬁc antigens such as GNT-V.A2. But some other antigens of this type appear to elicit spontaneous responses more frequently. J¨ager et al. (102, 103) demonstrated that in 10/27 patients with tumors expressing NY-ESO-1, and only in those, a spontaneous CTL response against NY-ESO-1 occurred, which was strongly correlated with the occurrence of an antibody response against the NYESO-1 protein. Valmori et al. (104) showed that TILs contained anti-NY-ESO-1 CTL and found that a patient had in the blood a frequency of 4 × 10−4 of CD8+ T cells that were directed against a NY-ESO-1 peptide. Some melanoma patients have spontaneous responses against an out-of-frame antigenic peptide encoded by LAGE-1 (105). Hercend and colleagues (106–108) observed an oligoclonal ampliﬁcation of T cells

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in a spontaneously regressing melanoma lesion. Interestingly, the most frequent T cell clonotype present in the tumor was not the dominant clone in the TIL culture. Nevertheless, it proved possible to obtain a permanent T cell clone with the dominant TIL clonotype, and the clone recognized an antigen encoded by gene MAGE-A6 (106–108). A detailed analysis of the blood of one melanoma patient before vaccination indicated the presence of ﬁve ampliﬁed CTL clonotypes recognizing peptides encoded by gene MAGE-C2 (97). The frequencies of these CTL ranged from 2 × 10−6 to 4 × 10−5 of the blood CD8+ cells. These CTL were found to be highly enriched in a skin metastasis, where they reached frequencies of 0.1% (109). No response was observed in this patient against antigens encoded by other cancer-germline genes, even though these were expressed by the tumor at a similar level as MAGE-C2. A spontaneous anti-MAGE-C2 response has been found in another patient, suggesting that, like NY-ESO-1, MAGE-C2 is a preferred target for spontaneous T cell responses. These responses against MAGE-C2 antigens target several different peptides presented by several different HLA molecules. Wang et al. (110) reported that a LAGE1 antigen was the target of tumor-speciﬁc CD4+ regulatory T cell (Treg) clones derived from the TILs of a melanoma patient. Phenotypic and functional analyses demonstrated that the CD4+ Tregs expressed CD25 and GITR (glucocorticoid-induced TNF receptor family–related) molecules and exerted a suppressive activity on the proliferative response of naive CD4+ T cells to anti-CD3 antibody treatment (110).

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Treg: regulatory T cell

Spontaneous responses against mutated antigens. In the study mentioned above by Hercend et al. (108), one of the frequent T cell clonotypes present in the tumor was found to recognize a mutated antigen encoded by a myosin gene (111). A mutated tyrosine phosphatase was the target of a CD4 T cell clone isolated from the TILs of a melanoma patient 182

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(112). Sensi et al. (113) found that about 0.4% of the CD8+ T cells present in a lymph node invaded by a melanoma were speciﬁc for a unique antigen generated by a point mutation in the peroxiredoxin 5 gene. Spontaneous responses against differentiation antigens. TILs of melanoma patients were shown to contain not only CD8 T cells but also CD4 T cells recognizing melanocyte differentiation antigens (114, 115). A very interesting and exceptional situation is that of the Melan-A/Mart-126−35 antigenic peptide, which is recognized by T cells on HLA-A2. This antigenic peptide is located in the putative transmembrane region of the 118 amino acid Melan-A protein. Only in this region were Melan-A antigenic peptides identiﬁed. High frequencies (∼0.1%) of anti-Melan-A.A2 CD8 T cells are found in the blood of most melanoma patients. Still higher frequencies, of the order of several percent, are found among CD8 T cells in invaded lymph nodes or cutaneous metastases (116). But healthy individuals also harbor in their blood a frequency of anti-Melan-A.A2 of about 0.005%, a frequency that is two orders of magnitude higher than that observed for naive precursors against MAGE antigens. Romero and colleagues (116) showed that these lymphocytes, which have a naive phenotype of CCR7+ CD45RA+ , appear to have multiplied little since their exit from the thymus as judged from their content of TCR excision circles and their telomere length (117). The same group identiﬁed a large number of cross-reactive peptides in a protein data search, suggesting that presentation of these antigenic peptides in the thymus could account for the extraordinary high frequency of anti-Melan-A T cells (87). Considering the frequency in healthy individuals, can one conclude that a spontaneous Melan-A response occurs in melanoma patients? The answer appears to be yes in many patients, who show a frequency of CTL precursors that is 20-fold higher than in noncancerous individuals. Moreover, one observes, particularly in

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advanced patients, that a signiﬁcant fraction of the anti-Melan-A T cells have a memory effector phenotype of CCR7+ CD45RO+ , whereas in healthy individuals all these T cells have a naive phenotype (118). The situation is very different for the other melanocyte differentiation antigens, which appear to stimulate spontaneous CTL responses much less often. Peter Lee (119) screened 11 melanoma patients for the presence of T cells recognizing the tyrosinase369−377 peptide. Two were positive, with frequencies of 0.2 and 2% of the CD8+ cells. Frequencies around 0.2% were also observed in 2/6 patients analyzed by another group (120). We screened a dozen melanoma patients and found no spontaneous response against this tyrosinase peptide or against the gp100209−217 peptide (P.G. Coulie, unpublished data).

Benefit to the Patient Several authors have examined the correlation between intratumoral T cell inﬁltrates and clinical evolution. T cell inﬁltration in primary melanoma was reported to correlate with a better eight-year survival rate (121, 122). The inﬁltration by T lymphocytes of tumor cell areas in an invaded lymph node was also found to correspond with a better prognosis (123). However, in advanced melanoma patients (stage IV), the presence of antiMelan-A- or antityrosinase-speciﬁc T cells in the blood was not correlated with better survival (124). Similar observations were made in prostatic adenocarcinomas (125). In colorectal cancer, a study showed that invasion of CD8+ T cells inside cancer cell nests signiﬁcantly correlated with a favorable prognosis, whereas inﬁltrates at the tumor margin or in the stroma did not (126). The same conclusion was reached for esophageal carcinomas (127). In a study on epithelial ovarian cancer collected at the time of surgical debulking prior to the chemotherapy, Zhang et al. (128) reported a strong correlation between the presence of an intratu-

moral T cell inﬁltrate and both disease-free and overall survival. Interestingly, the level of RNA for cytokines, such as monokine induced by IFN-γ, macrophage-derived chemokine, and secondary lymphoid-tissue chemokine, was markedly higher in those inﬁltrated tumors that were from patients with a better progression-free survival. An interesting observation, which did not involve a truly spontaneous T cell response, was made in chronic myelogenous leukemia. IFN-α2b treatment produced about 60% overall responses. In 11 out of 12 patients who responded, CTL directed against a peptide encoded by proteinase 3 appeared following the treatment. None of the seven patients who did not respond to the treatment showed these CTL. Proteinase 3 is expressed in the azurophil granules of the normal myeloid cells and is overexpressed twoto ﬁvefold in some leukemia cells. These observations suggest that the effect of IFN-α2b may be mediated by a T cell response (129). We tend to conclude that spontaneous T cell responses correlate with a better prognosis, even though the correlation may be obscured in some circumstances by the fact that this T cell response must be initiated at one point in the evolution of the tumor and therefore must be more frequently observed in more advanced tumors. From this correlation we cannot, however, rigorously conclude that the T cell inﬁltration is the cause of a better evolution of the tumor.

Tumor Resistance and T Cell Anergy On the basis of the observations described above, one may conclude that most metastatic melanoma patients are progressing despite having mounted a quantitatively signiﬁcant T cell response against their tumor, resulting in a CTL precursor frequency of 10−4 to 10−3 of CD8 T cells in the blood. The lack of effectiveness of the antitumoral T cells does not seem to be due to their failure to reach the tumor sites. They are present there in numbers that are not very different from those observed in metastases that are regressing following

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immunotherapy, and they are considerably enriched relative to the blood (109). Considering that antitumor T cells are clearly capable of rallying to the tumor site, the spontaneous antitumoral T cell response must become ineffective at one point, either because the tumor cells have become insensitive to the effector cells or because the effector cells themselves have become unable to be stimulated by the antigen or to exert their function, a state we refer to as anergy. This anergy could result from inhibitory processes elicited by tumor cells that were selected by the immune response. A mechanism of tumor escape is the selection of tumor cell variants that no longer present the antigens that are the targets of the effector T cells. The existence of such tumor cell variants has been very well documented (130). The loss of antigen can be due to a defect in the presentation system. General loss of expression of HLA class I molecules, often due to mutation in the β-2 microglobulin gene or to loss of expression of one HLA haplotype or of a single HLA allele, has also been observed on many samples of tumors of different histological types (131). HLA loss or downregulation were observed in a signiﬁcant fraction of melanoma primary and metastatic samples (132). A clear example of partial HLA loss followed by evolution of the T cell response to target antigens presented by other HLA molecules has been reported recently (133). Antigen loss can also result from a mutation or deletion in the gene coding for the antigenic peptide. This has been well documented in mouse systems but to our knowledge not often in human tumors. One apparent exception is the switching off of the genes coding for melanoma differentiation antigens. However, we have observed that such genes can be readily switched on and off in cultured cells, and it is therefore difﬁcult to assign their arrest to immunoselection. Is antigen loss a major mechanism of tumor escape from spontaneous T cell response, and is it a major source of ineffec-

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tiveness of immunotherapy? In our opinion, it is not presently the major cause of failure of antitumoral vaccination strategies in melanoma. When we isolate tumor cell lines from metastatic samples either before or after a vaccine therapy, we ﬁnd in more than 90% of instances that these cells are capable of restimulating in vitro autologous T lymphocytes and are sensitive to speciﬁc lysis by the resulting CTL. This does not exclude, however, that if at one point antitumoral vaccination becomes effective in causing complete tumor regressions in a large fraction of the patients, relapses will be observed that result from antigen loss due to immunoselection. If one admits that the spontaneous antitumoral T cell response becomes blocked, a ﬁrst issue is whether there is systemic anergy or whether the anergy is a local phenomenon occurring at the tumor site. Are antitumoral T cells present in the blood less readily reactivatable in vivo than memory T cells directed against, e.g., viral antigens? Do they show signs of impaired lytic functionality? This issue can be analyzed by capturing with tetramers blood T cells that have the appropriate speciﬁcity and then studying their properties. Contradictory observations have been reported. Lee et al. (119) reported that antityrosinase CD8+ T cells in the blood of a melanoma patient were not readily reactivatable in vitro. In contrast, Zippelius et al. (134) found that CCR7− CD8+ T cells directed against the MelanA.A2 antigen and present in the blood expressed IFN-γ upon antigenic stimulations and contained perforin and granzyme. This ﬁnding was in sharp contrast to ﬁndings with TILs collected from tumor sites: Anti-MelanA.A2 CCR7− CD8+ T cells did not synthesize IFN-γ upon short-term restimulation with antigen, whereas anti-cytomegalovirus T cells present in metastases could be stimulated (134). There is additional evidence that antitumoral T cells present in metastases are anergic. Recently, Bronte et al. (135) showed that T cells present in prostatic carcinoma

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locations did not upregulate CD25, CD69, or CD137 upon in situ phytohemagglutinin- or phorbol myristate acetate–ionomycin stimulation. In contrast, T cells present in normal prostate could be stimulated. Interestingly, T cells inside prostatic carcinomas appeared to lack polarization of their cytotoxic granules, which ought to have prevented their lytic ability (135). This phenomenon had been previously described in mouse tumors (136). To conclude, we feel that there is credible evidence for local T cell anergy in tumor locations, whereas more evidence should be obtained to establish if there exists some degree of systemic anergy. What is the cause of local anergy? There seem to be two main possibilities. Either the tumor cells, or the stroma inﬂuenced by the tumor cells, anergize the potential responder T cells directly, or they generate suppressive T cells, now called regulatory T cells (Tregs), that in turn anergize the responder T cells. During the past few years, the concept of Tregs has been revisited, largely owing to the work of Sakaguchi (137, 138). In humans, CD4+ CD25+ T cells present in the blood can exert an inhibitory effect in vitro on the CD4+ CD25− T cell population and T cell clones (139–141). This ﬁnding is leading to a reexploration of the concept that antitumoral immune responses might be blocked by such Tregs. T cell clones that were derived from TILs and directed against a LAGE and a mutated antigen were found to have the functional proﬁle of Treg cells (110, 142). Another systemic suppressive mechanism has been proposed: Observations in renal carcinoma patients showed that a subset of human polymorphonuclear granulocytes produced arginase, resulting in a low arginine level in the blood and depressed T cell functions (143). There is no lack of candidate agents that tumor cells could use to directly or indirectly anergize responder T cells. In mouse systems, TGF-β could play a major role in inhibiting antitumoral immune responses, but, to our knowledge, ﬁrm evidence that it plays a

determining role in the evolution of human tumors is not yet available (144). Many human tumor cells express IDO (indoleamine 2,3-dioxygenase), an enzyme also expressed in placenta, which degrades tryptophan (145). Cells expressing IDO serve as a tryptophan sink that deprives surrounding T lymphocytes of tryptophan and impairs their function. Bronte et al. (135) observed that inhibitors of arginase and nitric oxide synthase restored the functionality of TILs. Human melanoma cells secrete galectin-1, which inhibits T cells (146), and also produce IL-10, which could inhibit production of proinﬂammatory cytokines such as TNF-α, IFN-γ, and IL-2 (147). Human melanoma cells, particularly in the presence of IFN-γ, express the B7-H1 surface molecule (148). This molecule, which binds to the PD-1 receptor on T cells, causes increased apoptosis.

IDO: indoleamine 2,3-dioxygenase

T CELL RESPONSES AGAINST VACCINE ANTIGENS Quantitative Evaluation of T Cell Responses The efﬁcacy of the T cell response to a tumor antigen vaccine ought to depend on the number of activated T lymphocytes and on their functional properties. Even though it appears very plausible that T cells with highafﬁnity receptors might be more effective and might even be required for an antitumoral effect (149), there is no evidence regarding the set of functional properties that are required for antivaccine T lymphocytes to be effective. This leaves us with only a quantitative evaluation of the T cell responses to vaccines. The evaluation of the frequency of vaccine-induced CD8+ T cells can be performed ex vivo, that is, with T cells that are tested immediately upon their collection from the blood. Alternatively, it can be performed after an in vitro restimulation period of several days, which results in the proliferation of the antivaccine T cells. The lymphocytes are usually drawn from the blood, but they can also be collected from tumor sites.

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CD8 responses. Ex vivo frequency assays allow for a direct evaluation of the frequency of antivaccine T cells. Three assays are commonly used. The tetramer assay measures the fraction of cells expressing TCRs speciﬁc for a given antigen, regardless of the functional characteristics of these T cells. The lymphocytes can then be counted and sorted while staying alive (150). Tetramers are soluble complexes of recombinant MHC molecules folded in the presence of an antigenic peptide and biotinylated. When these complexes are bound to ﬂuorescinated avidin, which can bind four biotin moieties, the name tetramer is appropriate. But when the ﬂuorescent reagent is phycoerythrin, which spontaneously polymerizes, it brings several avidin molecules together so that the reagent is really a multimer. Nevertheless, we use the word tetramer throughout. With most HLA class I tetramers, the lower detection limit of the test is ∼3 × 10−4 of the CD8+ cells. A second assay that is frequently used is the ELISPOT assay for IFN-γ secretion. Antibodies are used to detect the cytokine produced by individual cells after a short (6–48 h) period of restimulation with the antigen. The test has been adapted to the study of the response of melanoma patients by Wolfel and colleagues (151), who greatly simpliﬁed and standardized the reading procedure by using computer-assisted image analysis (152). When the assay is performed with unfractionated peripheral blood mononuclear cells (PBMC), the sensitivity of the assay is about 5 × 10−4 of the CD8+ T cells. A third assay also evaluates the number of cells producing IFN-γ upon contact with the antigen. During the last 4–18 h of stimulation, brefeldin is added to block the export of the cytokine, which then accumulates in the cell. The lymphocytes are ﬁxed, permeabilized, and stained with antibody. The fraction of positive cells is evaluated by ﬂow cytometry. The sensitivity of this assay is similar to that of ELISPOT. As with ELISPOT, the lymphocytes are lost during the assay, preventing any further assessment of their speciﬁcity.

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Because the frequency of antivaccine responder T cells is often lower than the detection limit of the assays just mentioned, it is usually necessary to submit the collected lymphocytes to an in vitro restimulation with the vaccine antigen during 5–14 days. During this restimulation, the precursor lymphocytes (CTLp) that recognize the antigen can multiply by several hundredfold. This ampliﬁcation can be followed, as for the ex vivo procedure, by a tetramer, ELISPOT, or intracellular cytokine assay. After in vitro restimulation, it is also possible to use a lysis assay on a target cell presenting the antigen. For a proper evaluation of the frequency of antivaccine T cell precursors in the original sample, it is essential to conduct the restimulation process under limiting dilution conditions, for all that can be deduced from the positive scoring of a restimulated culture is that it contained at least one relevant T cell precursor to start with. We have used a limiting dilution restimulation followed by a tetramer assay and the cloning of the positive microcultures. A lytic assay is then performed with each T cell clone, not only on peptide-pulsed cells but also on target cells expressing the appropriate antigen-encoding gene and HLA molecule. This approach is long and labor intensive, but it provides a stringent control of the speciﬁcity of the putative antivaccine T cells as well as the possibility of assessing their TCR sequence. This in turn provides the possibility of setting up clonotypic polymerase chain reaction (PCR) assays to reevaluate directly precursor frequencies in the blood and to compare these with the estimates obtained by the restimulation-tetramer procedure. The comparison has shown that, with a few exceptions, the results were remarkably close, leading to the conclusion that at least half of the antivaccine T cell precursors multiply in vitro under the restimulation conditions. However, if a precursor were totally refractory to our in vitro restimulation conditions, we would fail to notice its existence. We also observed that T cell precursors belonging to the same clonotype can proliferate at vastly different

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rates in independent microcultures, conﬁrming the necessity of the limiting dilution approach for a frequency evaluation. We believe that one can detect very lowfrequency antivaccine T cell responses that are barely above the naive precursor frequency by evaluating the presence of repeated clonotypes. As stated below, we ﬁnd that in a noncancerous individual, the frequency of precursors of T cells recognizing a MAGEA3 antigen presented by HLA-A1 is 4 × 10−7 CD8+ T cells, with a TCR repertoire of >100 clonotypes (100). On this basis, when sequence analysis indicates that at least 3/10 or 4/20 T cell clones derived from independent microcultures have the same TCR, we conclude that an ampliﬁed clonotype is present, which represents a response to the vaccine. CD4 responses. Tetramers of class II HLA molecules have proven much more arduous to produce than their class I equivalents, presumably because of the intrinsic structural instability of class II molecules in solution. A ﬁrst use of these tetramers was the evaluation of an anti-inﬂuenza response with a DR4 tetramer (153). We produced a DP4 tetramer that can be used for ex vivo staining of PBMC. The positive cells are sorted by ﬂow cytometry. The sorted cells are then cloned by limiting dilution. After multiplication, the speciﬁcity of the T cell clones is assessed by their ability to secrete IFN-γ upon contact with EBVtransformed B cells pulsed with the peptide or EBV-B cells transduced with a retrovirus carrying the protein coding sequence fused to a truncated invariant chain. This approach has a high sensitivity: Frequencies as low as 10−6 of CD4+ T cells have been ascertained (154). Ex vivo ELISPOT approaches involving release of IFN-γ, IL-2, and IL-4 following stimulation with the relevant peptide have also been used (155). Sensitivity of 3 × 10−4 of CD4+ T cells can be reached. The IFN-γELISPOT test can also be used after in vitro restimulation with a peptide, but here again,

a limiting dilution approach is required for a frequency evaluation. It is also important to analyze CD4+ T cell responses in patients vaccinated with protein under conditions in which the target antigenic peptides are unknown. One approach is a brief stimulation with DCs pulsed with protein, followed by an IFN-γ secretion assay involving the immediate recapture of the cytokine secreted by one lymphocyte with bispeciﬁc antibodies coating the cell. The cells are then stained with a ﬂuorescent anti-IFN-γ antibody and sorted by ﬂow cytometry. Cloning and functional analysis are performed as described above (156).

The Antivaccine Responses Responses against antigens encoded by cancer-germline genes. Early investigations regarding the CTL responses of patients vaccinated with MAGE antigens indicated that these responses were not of a high level (157). To evaluate weak responses, it appeared necessary ﬁrst to obtain a precise description of the anti-MAGE T cell precursors in noncancerous individuals. The precursor T cell frequency against the MAGE-A3168−176 peptide presented by HLA-A1 (MAGE-3.A1) was analyzed in detail (99, 100). The naive precursor frequency in the blood was estimated at 4 × 10−7 of CD8+ T cells and was found to be remarkably similar in different individuals. Independent T cell clones of one individual were tested for their TCRβ chain sequences, and only one repeat was found among 23 clones. The statistical analysis of this result led to the conclusion that the diversity of the naive repertoire against the MAGE-3.A1 antigen is very likely to exceed 100. Considering that the total number of CD8+ T cells in humans is about 4 × 1010 , one can therefore reasonably represent the naive CTLp as comprising about 100 clones of 160 cells each (Figure 1). Naive CTLp frequencies for another MAGE-A3 antigen were ﬁrst evaluated to be ∼10−6 of CD8 T cells (99), but later studies suggested somewhat lower

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Figure 1 Frequencies and diversity of anti-MAGE-3.A1 CTLs. Different clonotypes are represented by circled numbers.

frequencies of ∼5 × 10−7 . We have also observed frequencies of ∼5 × 10−7 for several other antigens, such as MAGE-A1278−286 , tyrosinase.A2, GNT-V.A2, MAGE-A1254−262 , and MAGE-C2336−344 , in prevaccination samples of melanoma patients (158; P.G. Coulie, unpublished data). Vaccines including antigen MAGE-3.A1 have been applied in various modalities to metastatic melanoma patients with detectable disease. In such patients, spontaneous T cell responses against the MAGE-3.A1 antigen are rare, as we observed only two such cases out of 45 (100, 101). Following vaccination with the peptide without adjuvant, seven patients showing evidence of tumor regression were analyzed, and a CTL response was observed in only two patients, with frequencies of 5 × 10−6 and 4 × 10−5 , respectively (159, 160). Other patients were vaccinated with a recombinant ALVAC (canarypox) virus containing the MAGE-3.A1 coding sequence. Among four patients showing evidence of tumor regression, anti-MAGE-3.A1 CTL responses were observed in three, with frequen188

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cies ranging from 3 × 10−3 to 7 × 10−7 , the latter being considered as a response because of the presence of a repeated clonotype (100, 161). For all the patients vaccinated with peptide or ALVAC, TCR analysis showed that the CTL response was monoclonal. Another constant feature of the anti-MAGE-3.A1 T cell responses is their stability: Once detected, the T cells appear to remain in the same frequency range, even several months following the last revaccination. Metastatic melanoma patients were also immunized with monocyte-derived mature DCs pulsed with the MAGE-3.A1 peptide (162). Analyses of three regressor patients showed a CTL response in all three, with frequencies ranging from 3 × 10−6 to 10−3 (163). In contrast with the results obtained with peptide or ALVAC, the responses of these three patients were polyclonal. No response could be detected in ﬁve vaccinated patients who did not show tumor regression. In another study involving vaccination with CD34+ progenitor-derived DCs pulsed with peptide MAGE-A3271−279 , ﬁve patients out of

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eight appeared to have a T cell response (164). On the basis of these results, one may conclude that when peptide-pulsed DCs stimulate a response against antigens encoded by cancer-germline genes, the responses are somewhat stronger than those elicited by peptide alone or by ALVAC, but there are still many patients who do not show a T cell response. Another possible method for increasing the immunogenicity of cancer-germlineencoded antigens is to combine them with immunological adjuvants. In a study in which peptide MAGE-A3271−279 was injected with IL-12 into four patients, ELISPOT assays after restimulation (which was not performed under limiting dilution conditions) suggested a response, as prevaccination tests were negative (165). No response was observed in two patients who received the peptide without IL-12. Using the same detection approach, responses were reported for peptides MAGE-A1243−258 and MAGE-A10254−262 injected with incomplete Freund’s adjuvant (IFA) and GM-CSF (166). In another study, no CTL response was observed in seven melanoma patients vaccinated with peptide MAGE-A12170−178 with IFA (167). Considering the strong T cell responses obtained with the Melan-A/MART-1 peptide combined with IFA and CpG (see below), it will be interesting to test this combination of adjuvants with MAGE peptides. Finally, the possibility of providing help to the CD8 response by vaccinating with antigens recognized by CD4+ T cells has been explored. We injected four patients with peptide MAGE-3.A1 together with peptide MAGEA3243−258 , presented by HLA-DP4, and observed no response (P. van der Bruggen, unpublished data). A recombinant MAGE-A3 protein mixed with the adjuvant AS02B was administered to 24 patients. All patients except one mounted an IgG response to the protein, indicating a stimulation of CD4+ T cells (168). When the protein was used without adjuvant, only 1/24 patients produced an anti-MAGE-A3 IgG response, conﬁrm-

ing the importance of the adjuvant for the in vivo stimulation of CD4+ T cells (169). It would be interesting to analyze the adjuvant effect of this protein-AS02B combination on CD8 responses against coadministered class I MAGE peptides. A few experiments were performed to document the CD4 responses to the MAGE-A3 protein. In a patient vaccinated with the protein alone, anti-MAGE-A3 CD4+ T cells were detected at a frequency of 1.5 × 10−5 of blood CD4+ cells, representing at least an 80-fold increase over the prevaccination frequency (156). Most of these CD4+ cells recognized the same peptide, presented by HLA-DR1. Class II MAGE peptides have also been combined with DCs. HLA-DP4restricted CD4+ T cells recognizing peptide MAGE-A3243−258 were found in 7 × 10−4 of CD4+ blood T cells after vaccination with peptide-pulsed DCs (154, 155). The evaluation of the responses of patients vaccinated with the NY-ESO-1.A2 peptide is complicated by the fact that, at the time of vaccination, about half the patients with NY-ESO-1+ tumors had already made a spontaneous T cell response against this antigen. Vaccination of patients with preexisting responses did not seem to lead to a clear increase in the CTL frequency. Vaccination of the other patients clearly produced T cell responses (102). A recent report suggests that melanoma patients with no detectable disease who were vaccinated with NY-ESO-1 protein and ISCOMATRIX, a saponin-based adjuvant, had CD8 and CD4 responses against a wide range of NY-ESO-1-encoded peptides (170).

IFA: incomplete Freund’s adjuvant

Vaccination with melanocyte differentiation antigens. Several groups reported the induction of CD8 T cell responses after vaccination of melanoma patients with the Melan-A/MART-126−35(27L) peptide administered with IFA (171–173). Speiser et al. (174) compared responses after vaccination with the peptide in IFA with or without CpG oligodeoxynucleotides. With CpG, more patients (8/8 versus 4/8) had a detectable

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increase in the frequency of anti-Melan-A CD8 T cells, and the percentages of antiMelan-A T cells among the CD8 were tenfold higher, reaching 1% in several patients. This is higher than the frequency observed after any of the following vaccinations: DC pulsed with a set of peptides including Melan-A26−35 (175); a recombinant vaccinia virus encoding antigenic peptides together with B7.1 and B7.2 molecules (176); a recombinant modiﬁed Ankara vaccinia virus encoding seven antigenic peptides (177); peptide and IL-12 (178); or peptides with IFA and an anti-CTLA-4 monoclonal antibody (179). Peptide gp100209−217(210M) has also been used in several vaccination trials. In one trial, the peptide was injected in IFA, and patients also received IL-2. Signiﬁcant T cell responses were observed (180). In another study in which a higher dose of peptide was combined with IFA, ex vivo tetramer analysis also indicated that almost all patients made a strong T cell response, with an average of 0.3% of CD8, whereas prevaccination frequencies were 0.02% (181). The peptide was also injected into patients with IL-12. Strong T cell responses with frequencies ranging from 0.05% to 2.5% of CD8 T cells were observed. In all patients, anti-gp100 T cells were undetectable in prevaccination samples (182). Slingluff and coworkers (183, 184) performed interesting studies in which the T cell responses observed in lymph nodes were compared with those observed in the blood following vaccination with a number of peptides, including gp100209−217 . T cell responses were observed more frequently in lymph nodes draining the vaccination site than in blood. In a study by Schaed et al. (185), ex vivo ELISPOT assays were used to compare postvaccination frequencies of T cells against peptide tyrosinase369−377 administered with either IFA, the puriﬁed saponin QS-21, or GM-CSF. None of the nine patients immunized with IFA developed a detectable response, whereas 3/9 and 2/8 did so after vaccination with QS-

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21 and GM-CSF, respectively. Immunization with DCs pulsed with various differentiation peptides produced T cell responses that appeared to be similar to those obtained with the modalities described above (155, 164, 186). Thus, it seems that the highest responses obtained against differentiation antigens are higher than the highest responses obtained against antigens encoded by cancer-germline genes.

RELATIONSHIP BETWEEN T CELL RESPONSES AND TUMOR REGRESSIONS Vaccination of metastatic melanoma patients with deﬁned antigens has up to now produced only modest results in terms of beneﬁt to the patients. Only a small minority of patients (5%–10%) has shown an objective clinical response. We have vaccinated with MAGE-A3 antigens metastatic melanoma patients with detectable disease using various modalities ranging from the MAGE-3.A1 antigenic peptide alone, to peptide with adjuvant, protein with adjuvant, and recombinant ALVAC. Of a total of 111 patients included in these various studies, 8 have shown objective clinical responses (159, 168, 169, 187). But some evidence of tumor regression, usually mixed responses, has been observed in 12 additional patients. We accept that these regressions are not clinically signiﬁcant, but we feel that they must be taken into account for the elucidation of the process by which the vaccines are occasionally effective. The results obtained by many groups using other antigens encoded by cancer-germline genes (MAGE, NY-ESO-1), or differentiation antigens (tyrosinase, Melan-A/Mart-1, gp100), or even allogenic melanoma lysates appear to produce tumor responses in the same frequency range (165, 178, 184, 188, 189). Rosenberg and coworkers (190) reported an even lower average objective clinical response frequency of 2.6% out of 444 treated patients. Clinical trials with DCs pulsed with peptide do not appear to produce markedly

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better tumor response rates (162, 186, 191– 194). This appraisal should, however, be received with some caution, as the small number of patients included in most studies precludes concluding either that one vaccination modality is superior to others or that it is not. Melanoma tumors occasionally regress spontaneously. But in metastatic melanoma these regressions appear to occur rarely, at a frequency situated between 0.1% and 1% (195). Infrequent as they are, the objective clinical responses and the other tumor regressions that occur following vaccination are above this frequency of spontaneous regression. Moreover, they occur in the weeks or months following onset of vaccination. Effects can also be observed in an adjuvant setting: Keilholz and coworkers (196) observed that among nine patients who had presented more than three new metastases in the year before vaccination, two patients showed no relapse for more than two years (196). We tentatively conclude that the regressions observed following vaccination are produced by the vaccine. But it is difﬁcult to exclude the possibility that patients included in therapeutic vaccination trials represent a biased favorable subset. For instance, there may be a selection of patients with slowly progressing tumors. The low proportion of objective clinical responses does not appear to be due to the fact that 90%–95% of melanoma tumors are absolutely resistant to T cell attack. Rosenberg et al. (197) have shown that following adoptive transfer of a massive number of antitumor CTL (1011 ), objective clinical responses are observed in about 50% of the patients. Before the adoptive transfer, these patients received a treatment with cyclophosphamide aimed at eliminating the bulk of Tregs. It is also noteworthy that B. Dr´eno and coworkers (198) have shown rigorously in a randomized trial that adoptive transfer of TILs in an adjuvant setting was signiﬁcantly extending the duration of relapse-free survival of patients with only one invaded lymph node.

Is there a correlation between the occurrence of a detectable antivaccine T cell response and tumor regression? For patients vaccinated with ALVAC-MAGE-3.A1, we observed a deﬁnite correlation, as 3/4 of patients with evidence of tumor regression showed an antivaccine T cell response, in contrast to 1/17 patients without tumor regression (100, 187). In a small group of patients vaccinated with DCs, all patients with regressions showed a T cell response (162, 163). In contrast, for patients vaccinated with peptide alone, we see antivaccine T cell responses only in a minority of the patients showing a regression. If we consider all the patients having received various modalities of MAGE-3.A1 antigen, we obtain a total of 10/20 for responder patients and 1/30 for nonresponders. There is therefore a clear correlation between T cell responses and tumor regressions. Attempts have been made to improve correlations by including various parameters in addition to antivaccine T cell frequencies in the blood. Figdor et al. (193) observed in patients vaccinated with mature DC pulsed with gp100 and tyrosinase peptides that all the patients produced delayed-type hypersensitivity (DTH) following challenge with those pulsed DC. Very interestingly, only in the patients showing evidence of tumor regression were anti-gp100 or antityrosinase CTL precursors present at the DTH site (193). Banchereau et al. (186) used a compound score involving a statistical analysis of the results of direct and recall assays of all the vaccine antigens. Speiser et al. (199) found that tumor regression following vaccination with the Melan-A.A2 peptide in IFA correlated well with a compound score involving antivaccine T cell frequency increase in the blood and increase in CD45RA− or CD28− anti-Melan-A T cells. In many MAGE-vaccinated patients who show tumor regression, the frequency of antivaccine T cells is very low. One wonders how such a low number of T cells could provide the main component of the speciﬁc effectors destroying the tumor cells. This led us to

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evaluate in some vaccinated patients the total frequency of T cells that recognized the autologous tumor, hereafter labeled antitumor T cells. In addition, we analyzed not only the T cells present in the blood but also those present at tumor sites. A detailed analysis was carried out on one patient, EB81, who showed complete regression of all of a large number of skin metastases, but had a progressing lymph node removed in the course of vaccination. A tumor cell line was obtained from the lymph node and it was used to stimulate CD8 T cells. We observed that the vaccine had elicited a frequency of about 3 × 10−6 anti-MAGE-3.A1 T cells, and that after vaccination antitumor T cells (deﬁned by their ability to lyse autologous tumor cells and not EBV-B cells or NK target K562) were present at a frequency of 10−3 . Moreover these antitumor T cells were already present at a similar high frequency before vaccination. Five other patients were analyzed, and all presented the same high frequency of antitumor T cells before as well as after vaccination (97). The antitumor T cells of patient EB81 were cloned, and the antigens of 13/15 clones could be identiﬁed. Three were directed against gp100 antigens and ten were directed against various antigenic peptides, all encoded by gene MAGE-C2. Gene MAGE-C2 is a cancer-germline gene belonging to a different MAGE family than MAGE-A3, the source of the vaccine antigen. Thus, the spontaneous antitumor T cell response of this patient was directed against the type of melanoma antigens that are used in the vaccines. PCR assays speciﬁc for TCR sequences made it possible to establish the frequency of antivaccine and antitumor T cell clones in a number of metastases. Whereas antivaccine T cells showed a modest enrichment in tumor sites relative to the blood, some antitumor T cell clones showed an enrichment by several hundredfold, reaching frequencies in the tumor of several percent of CD8 T cells. It appears that at least 50% of the T cells present at tumor sites of patient EB81 belong to a small number of antitumor T cell clones. The same antitu-

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mor T cells were found at high frequency in different metastases (109). These results have recently been conﬁrmed with another vaccinated patient. On the basis of these observations, we propose the following scenario for the occasional tumor regressions that are observed following vaccination (Figure 2). The melanoma metastases contain highly enriched antitumor T cells before vaccination. These T cells have become ineffective, presumably because of immunosuppressive factors produced by the tumor. In some vaccinated patients, antivaccine T cells migrate to the tumor and are capable of resisting the immunosuppressive environment long enough to attack some tumor cells and reverse focally the immunosuppression. This provides conditions that enable a much larger number of antitumor T cells to be stimulated and to proliferate. These antitumor T cells then serve as speciﬁc effectors to eliminate the bulk of the tumor cells. Thus, the antivaccine T cells only serve as a spark that triggers the process leading to the activation of the antitumor T cells. These activated antitumor T cells could either be derived from the blocked T cells that were already present in the tumor or result from a new wave of activation of naive precursors. Our analysis of patient EB81 indicated that regressing tumors contained T cell clones that were already present before vaccination, but that the most enriched T cell clones were not. It is therefore possible that a new wave of antitumor T cells constitutes an important component of the effectors of the tumor rejection. It is noteworthy that most of the antigens recognized by the new wave of antitumor T cells were already recognized by the prevaccination set of antitumor T cells. If correct, this hypothetical scenario leads to a reconsideration of the correlation between tumor regression and antivaccine T cell response. Insofar as the regression process involves a new stimulation and expansion process of antitumor lymphocytes, this process can also apply to the antivaccine T cells that have migrated to the tumor. Accordingly, the

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Figure 2 Hypothetical scenario for tumor regression following vaccination. T lymphocytes are represented by circles, with different colors indicating different antigenic speciﬁcities. Inactivation of these T cells is represented by faded out colors.

frequency of antivaccine T cells that is measured following vaccination would be the result of a two-stage ampliﬁcation: a ﬁrst one stimulated by the vaccine and a second one by the tumor. Therefore, the observation of a higher level of antivaccine T cells in a regressing patient would not necessarily be due to fact that in this patient the vaccine produced a higher response and therefore his tumor regressed. The antivaccine response may well not have been higher in this patient, but somehow it was capable of initiating a regression process that in turn resulted in the restimulation that produced a higher level of antivaccine T cells. In conclusion, therapeutic success following vaccination may not depend on the number of T cells produced directly by the vaccine, but rather on the production of a T cell clone with functional properties that enable it to migrate to the tumor and resist the

local immunosuppressive environment long enough to initiate a regression process. The view that the main effectors of tumor regression are antitumor cells distinct from the antivaccine T cells has other implications. One relates to the issue of tumor escape by antigen-loss variants. If antivaccine T cells were the only effectors of rejection, the loss of expression of the vaccine antigen by some tumor cells would ensure escape of the tumor. However, if many other antitumor effectors come into play, a general loss of antigen expression is required for escape. Another implication relates to the collaboration between CD4+ T helper cells and CD8+ effector T cells. A vast body of evidence supports the importance of this collaboration, which seems particularly important to the production of memory T cells (200). Accordingly, it has been claimed that effective

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vaccination of cancer patients requires the combined use of antigens presented by MHC class II and class I molecules. However, in our experience vaccines containing peptide MAGE-A3243−258 , which is presented by HLA-DP4, in addition to peptide MAGE3.A1, did not prove to be clearly superior to the vaccine containing only MAGE-3.A1. Other groups have included helper peptides or proteins, such as keyhole limpet hemocyanin, tetanus toxoid, or the “PADRE” peptide, in their vaccines (164, 166, 193, 201). In our view, it is impossible to conclude unequivocally that this improved the clinical responses. One explanation may be that if a regression process can be triggered by the antivaccine CD8, then the antitumor T cells that come into play comprise several CD4+ as well as CD8+ T cell clones, thus providing the required helper effect. To achieve therapeutic success, investigators will probably need to understand the cause of the local immunosuppression in the tumors and ﬁnd counteracting agents. As stated above, the list of possible immunosuppressive agents present in tumors is considerable. But it will be important to ﬁnd whether, for each type of tumor, there is a prevalent immunosuppressive agent. Just as many types of tumors have preferred oncogenic pathways that differ from one type of tumor to another, each type of tumor may also have preferred immunosuppressive processes that we must identify to achieve therapeutic success. A combination of vaccination with agents that reduce this immunosuppressive process may considerably increase the proportion of patients showing clinical responses following immunization. Recent immunotherapy trials have included cyclophosphamide pretreatment aimed at eliminating the bulk of the Tregs (197). Vaccination com-

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bined with anti-CTLA-4 antibodies has been applied to a small number of patients. In one study, 3/14 patients showed a clinical response, but many patients have suffered from severe autoimmune side effects (202). In another study, the tumor response rate did not seem to be very different from that obtained with other modalities (179). Therapeutic vaccination of cancer has not yet proved to be effective enough to become a generally applied cancer treatment. But considering its remarkably low toxicity, even in patients who have shown clinical response, it would be regrettable to abandon the effort at this stage without trying to understand the mechanisms of tumor resistance and ﬁnd the ways to overcome them. We can envision two extreme situations. In the ﬁrst, all patients would have tumors with a similar degree of tumor resistance. The difference between the regressors and the others would be due to a more effective response to the vaccine. We do not believe that melanoma patients suffer from a degree of general immunosuppression, which we believe is restricted to very late-stage patients who are not included in most studies (203). Therefore, the difference in the quality of the response would be due to a chance event determining, for instance, the functional properties of the unique or the few responder T cell clones elicited by the vaccine. In that case, it will be essential to understand what this crucial functional property is. At the other extreme, the antivaccine T cell responses would be similar in all patients, but the level of resistance of the tumors would vary considerably. In that case, investigators would need to identify the main component of this resistance and ﬁnd ways to counteract it. Analysis of gene expression in tumors using microarrays may be an important way to achieve this goal (204).

DISCLOSURE STATEMENT The Ludwig Institute owns several patents relative to tumor antigens discovered by the authors. The authors are entitled to a share of the royalties received by the Ludwig Institute for the licensing of the patents. 194

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LITERATURE CITED 1. Pardoll D. 2003. Does the immune system see tumors as foreign or self? Annu. Rev. Immunol. 21:807–39 2. Haas GG Jr, D’Cruz OJ, De Bault LE. 1988. Distribution of human leukocyte antigenABC and -D/DR antigens in the unﬁxed human testis. Am. J. Reprod. Immunol. Microbiol. 18:47–51 3. De Plaen E, Arden K, Traversari C, Gaforio JJ, Szikora JP, et al. 1994. Structure, chromosomal localization, and expression of 12 genes of the MAGE family. Immunogenetics 40:360–69 4. van der Bruggen P, Traversari C, Chomez P, Lurquin C, De Plaen E, et al. 1991. A gene encoding an antigen recognized by cytolytic T lymphocytes on a human melanoma. Science 254:1643–47 5. Chomez P, De Backer O, Bertrand M, De Plaen E, Boon T, Lucas S. 2001. An overview of the MAGE gene family with the identiﬁcation of all human members of the family. Cancer Res. 61:5544–51 6. De Smet C, De Backer O, Faraoni I, Lurquin C, Brasseur F, Boon T. 1996. The activation of human gene MAGE-1 in tumor cells is correlated with genome-wide demethylation. Proc. Natl. Acad. Sci. USA 93:7149–53 7. Lucas S, Brasseur F, Boon T. 1999. A new MAGE gene with ubiquitous expression does not code for known MAGE antigens recognized by T cells. Cancer Res. 59:4100–3 8. Ruault M, van der Bruggen P, Brun ME, Boyle S, Roizes G, De Sario A. 2002. New BAGE (B melanoma antigen) genes mapping to the juxtacentromeric regions of human chromosomes 13 and 21 have a cancer/testis expression proﬁle. Eur. J. Hum. Genet. 10:833– 40 9. De Backer O, Arden KC, Boretti M, Vantomme V, De Smet C, et al. 1999. Characterization of the GAGE genes that are expressed in various human cancers and in normal testis. Cancer Res. 59:3157–65 10. Lethe B, Lucas S, Michaux L, De Smet C, Godelaine D, et al. 1998. LAGE-1, a new gene with tumor speciﬁcity. Int. J. Cancer 76:903–8 11. Chen YT, Scanlan MJ, Sahin U, Tureci O, Gure AO, et al. 1997. A testicular antigen aberrantly expressed in human cancers detected by autologous antibody screening. Proc. Natl. Acad. Sci. USA 94:1914–18 12. Gure AO, Tureci O, Sahin U, Tsang S, Scanlan MJ, et al. 1997. SSX: a multigene family with several members transcribed in normal testis and human cancer. Int. J. Cancer 72:965– 71 13. Gure AO, Wei IJ, Old LJ, Chen YT. 2002. The SSX gene family: characterization of 9 complete genes. Int. J. Cancer 101:448–53 14. Van den Eynde B, Lethe B, Van Pel A, De Plaen E, Boon T. 1991. The gene coding for a major tumor rejection antigen of tumor P815 is identical to the normal gene of syngeneic DBA/2 mice. J. Exp. Med. 173:1373–84 15. Brandle D, Bilsborough J, Rulicke T, Uyttenhove C, Boon T, Van den Eynde BJ. 1998. The shared tumor-speciﬁc antigen encoded by mouse gene P1A is a target not only for cytolytic T lymphocytes but also for tumor rejection. Eur. J. Immunol. 28:4010– 19 16. Gotter J, Brors B, Hergenhahn M, Kyewski B. 2004. Medullary epithelial cells of the human thymus express a highly diverse selection of tissue-speciﬁc genes colocalized in chromosomal clusters. J. Exp. Med. 199:155–66 www.annualreviews.org • Human T Cell Responses Against Melanoma

195

ARI

23 February 2006

15:18

17. Derbinski J, Schulte A, Kyewski B, Klein L. 2001. Promiscuous gene expression in medullary thymic epithelial cells mirrors the peripheral self. Nat. Immunol. 2:1032–39 18. Van den Eynde B, van der Bruggen P. 2005. Peptide database of T-cell deﬁned tumor antigens. Cancer Immun. http://www.cancerimmunity.org/peptidedatabase/Tcellepitopes.htm 19. Wolfel T, Hauer M, Schneider J, Serrano M, Wolfel C, et al. 1995. A p16INK4ainsensitive CDK4 mutant targeted by cytolytic T lymphocytes in a human melanoma. Science 269:1281–84 20. Zuo L, Weger J, Yang Q, Goldstein AM, Tucker MA, et al. 1996. Germline mutations in the p16INK4a binding domain of CDK4 in familial melanoma. Nat. Genet. 12:97–99 21. Linard B, Bezieau S, Benlalam H, Labarriere N, Guilloux Y, et al. 2002. A ras-mutated peptide targeted by CTL inﬁltrating a human melanoma lesion. J. Immunol. 168:4802–8 22. Davies H, Bignell GR, Cox C, Stephens P, Edkins S, et al. 2002. Mutations of the BRAF gene in human cancer. Nature 417:949–54 23. Sharkey MS, Lizee G, Gonzales MI, Patel S, Topalian SL. 2004. CD4+ T-cell recognition of mutated B-RAF in melanoma patients harboring the V599E mutation. Cancer Res. 64:1595–99 24. Ikeda H, Lethe B, Lehmann F, van Baren N, Baurain JF, et al. 1997. Characterization of an antigen that is recognized on a melanoma showing partial HLA loss by CTL expressing an NK inhibitory receptor. Immunity 6:199–208 25. Kessler JH, Beekman NJ, Bres-Vloemans SA, Verdijk P, van Veelen PA, et al. 2001. Efﬁcient identiﬁcation of novel HLA-A(∗ )0201-presented cytotoxic T lymphocyte epitopes in the widely expressed tumor antigen PRAME by proteasome-mediated digestion analysis. J. Exp. Med. 193:73–88 26. Schmitz M, Diestelkoetter P, Weigle B, Schmachtenberg F, Stevanovic S, et al. 2000. Generation of survivin-speciﬁc CD8+ T effector cells by dendritic cells pulsed with protein or selected peptides. Cancer Res. 60:4845–49 27. Schmidt SM, Schag K, Muller MR, Weck MM, Appel S, et al. 2003. Survivin is a shared tumor-associated antigen expressed in a broad variety of malignancies and recognized by speciﬁc cytotoxic T cells. Blood 102:571–76 28. Vonderheide RH, Hahn WC, Schultze JL, Nadler LM. 1999. The telomerase catalytic subunit is a widely expressed tumor-associated antigen recognized by cytotoxic T lymphocytes. Immunity 10:673–79 29. Parkhurst MR, Riley JP, Igarashi T, Li Y, Robbins PF, Rosenberg SA. 2004. Immunization of patients with the hTERT:540–548 peptide induces peptide-reactive T lymphocytes that do not recognize tumors endogenously expressing telomerase. Clin. Cancer Res. 10:4688– 98 30. Ayyoub M, Migliaccio M, Guillaume P, Lienard D, Cerottini JC, et al. 2001. Lack of tumor recognition by hTERT peptide 540–548-speciﬁc CD8+ T cells from melanoma patients reveals inefﬁcient antigen processing. Eur. J. Immunol. 31:2642–51 31. Brichard V, Van Pel A, Wolfel T, Wolfel C, De Plaen E, et al. 1993. The tyrosinase gene codes for an antigen recognized by autologous cytolytic T lymphocytes on HLA-A2 melanomas. J. Exp. Med. 178:489–95 32. Coulie PG, Brichard V, Van Pel A, Wolfel T, Schneider J, et al. 1994. A new gene coding for a differentiation antigen recognized by autologous cytolytic T lymphocytes on HLAA2 melanomas. J. Exp. Med. 180:35–42 33. Bakker AB, Schreurs MW, de Boer AJ, Kawakami Y, Rosenberg SA, et al. 1994. Melanocyte lineage-speciﬁc antigen gp100 is recognized by melanoma-derived tumorinﬁltrating lymphocytes. J. Exp. Med. 179:1005–9

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

196

Boon et al.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

ARI

23 February 2006

15:18

34. Kawakami Y, Eliyahu S, Sakaguchi K, Robbins PF, Rivoltini L, et al. 1994. Identiﬁcation of the immunodominant peptides of the MART-1 human melanoma antigen recognized by the majority of HLA-A2-restricted tumor inﬁltrating lymphocytes. J. Exp. Med. 180:347– 52 35. Yee C, Thompson JA, Roche P, Byrd DR, Lee PP, et al. 2000. Melanocyte destruction after antigen-speciﬁc immunotherapy of melanoma: direct evidence of T cell-mediated vitiligo. J. Exp. Med. 192:1637–44 36. Kawakami Y, Robbins PF, Wang RF, Parkhurst M, Kang X, Rosenberg SA. 1998. The use of melanosomal proteins in the immunotherapy of melanoma. J. Immunother. 21:237–46 37. Morel S, Levy F, Burlet-Schiltz O, Brasseur F, Probst-Kepper M, et al. 2000. Processing of some antigens by the standard proteasome but not by the immunoproteasome results in poor presentation by dendritic cells. Immunity 12:107–17 38. Schultz ES, Chapiro J, Lurquin C, Claverol S, Burlet-Schiltz O, et al. 2002. The production of a new MAGE-3 peptide presented to cytolytic T lymphocytes by HLA-B40 requires the immunoproteasome. J. Exp. Med. 195:391–99 39. Van den Eynde BJ, Morel S. 2001. Differential processing of class-I-restricted epitopes by the standard proteasome and the immunoproteasome. Curr. Opin. Immunol. 13:147–53 40. Macagno A, Gilliet M, Sallusto F, Lanzavecchia A, Nestle FO, Groettrup M. 1999. Dendritic cells up-regulate immunoproteasomes and the proteasome regulator PA28 during maturation. Eur. J. Immunol. 29:4037–42 41. Macagno A, Kuehn L, de Giuli R, Groettrup M. 2001. Pronounced up-regulation of the PA28α/β proteasome regulator but little increase in the steady-state content of immunoproteasome during dendritic cell maturation. Eur. J. Immunol. 31:3271–80 42. Guilloux Y, Lucas S, Brichard VG, Van Pel A, Viret C, et al. 1996. A peptide recognized by human cytolytic T lymphocytes on HLA-A2 melanomas is encoded by an intron sequence of the N-acetylglucosaminyltransferase V gene. J. Exp. Med. 183:1173–83 43. Van den Eynde BJ, Gaugler B, Probst-Kepper M, Michaux L, Devuyst O, et al. 1999. A new antigen recognized by cytolytic T lymphocytes on a human kidney tumor results from reverse strand transcription. J. Exp. Med. 190:1793–800 44. Coulie PG, Lehmann F, Lethe B, Herman J, Lurquin C, et al. 1995. A mutated intron sequence codes for an antigenic peptide recognized by cytolytic T lymphocytes on a human melanoma. Proc. Natl. Acad. Sci. USA 92:7976–80 45. Lupetti R, Pisarra P, Verrecchia A, Farina C, Nicolini G, et al. 1998. Translation of a retained intron in tyrosinase-related protein (TRP) 2 mRNA generates a new cytotoxic T lymphocyte (CTL)-deﬁned and shared human melanoma antigen not expressed in normal cells of the melanocytic lineage. J. Exp. Med. 188:1005–16 46. Robbins PF, El-Gamil M, Li YF, Fitzgerald EB, Kawakami Y, Rosenberg SA. 1997. The intronic region of an incompletely spliced gp100 gene transcript encodes an epitope recognized by melanoma-reactive tumor-inﬁltrating lymphocytes. J. Immunol. 159: 303–8 47. Harada M, Li YF, El-Gamil M, Ohnmacht GA, Rosenberg SA, Robbins PF. 2001. Melanoma-Reactive CD8+ T cells recognize a novel tumor antigen expressed in a wide variety of tumor types. J. Immunother. 24:323–33 48. Wang RF, Appella E, Kawakami Y, Kang X, Rosenberg SA. 1996. Identiﬁcation of TRP-2 as a human tumor antigen recognized by cytotoxic T lymphocytes. J. Exp. Med. 184:2207– 16 49. Ronsin C, Chung-Scott V, Poullion I, Aknouche N, Gaudin C, Triebel F. 1999. A nonAUG-deﬁned alternative open reading frame of the intestinal carboxyl esterase mRNA www.annualreviews.org • Human T Cell Responses Against Melanoma

197

ANRV270-IY24-06

ARI

23 February 2006

50.

51.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

52.

53.

54.

55.

56. 57. 58. 59.

60.

61.

62.

63.

64.

198

15:18

generates an epitope recognized by renal cell carcinoma-reactive tumor-inﬁltrating lymphocytes in situ. J. Immunol. 163:483–90 Probst-Kepper M, Stroobant V, Kridel R, Gaugler B, Landry C, et al. 2001. An alternative open reading frame of the human macrophage colony-stimulating factor gene is independently translated and codes for an antigenic peptide of 14 amino acids recognized by tumor-inﬁltrating CD8 T lymphocytes. J. Exp. Med. 193:1189–98 Rosenberg SA, Tong-On P, Li Y, Riley JP, El-Gamil M, et al. 2002. Identiﬁcation of BING-4 cancer antigen translated from an alternative open reading frame of a gene in the extended MHC class II region using lymphocytes from a patient with a durable complete regression following immunotherapy. J. Immunol. 168:2402–7 Aarnoudse CA, van den Doel PB, Heemskerk B, Schrier PI. 1999. Interleukin-2-induced, melanoma-speciﬁc T cells recognize CAMEL, an unexpected translation product of LAGE-1. Int. J. Cancer 82:442–48 Wang RF, Johnston SL, Zeng G, Topalian SL, Schwartzentruber DJ, Rosenberg SA. 1998. A breast and melanoma-shared tumor antigen: T cell responses to antigenic peptides translated from different open reading frames. J. Immunol. 161:3598–606 Slager EH, van der Minne CE, Goudsmit J, van Oers JM, Kostense S, et al. 2004. Induction of CAMEL/NY-ESO-ORF2-speciﬁc CD8+ T cells upon stimulation with dendritic cells infected with a modiﬁed Ad5 vector expressing a chimeric Ad5/35 ﬁber. Cancer Gene Ther. 11:227–36 Skipper JC, Hendrickson RC, Gulden PH, Brichard V, Van Pel A, et al. 1996. An HLA-A2restricted tyrosinase antigen on melanoma cells results from posttranslational modiﬁcation and suggests a novel pathway for processing of membrane proteins. J. Exp. Med. 183:527– 34 Hanada K, Yewdell JW, Yang JC. 2004. Immune recognition of a human renal cancer antigen through post-translational protein splicing. Nature 427:252–56 Vigneron N, Stroobant V, Chapiro J, Ooms A, Degiovanni G, et al. 2004. An antigenic peptide produced by peptide splicing in the proteasome. Science 304:587–90 Ruiter DJ, Mattijssen V, Broecker EB, Ferrone S. 1991. MHC antigens in human melanomas. Semin. Cancer Biol. 2:35–45 Shaw AR, Chan JK, Reid S, Seehafer J. 1985. HLA-DR synthesis induction and expression in HLA-DR-negative carcinoma cell lines of diverse origins by interferon-γ but not by interferon-β. J. Natl. Cancer Inst. 74:1261–68 Chen BP, Madrigal A, Parham P. 1990. Cytotoxic T cell recognition of an endogenous class I HLA peptide presented by a class II HLA molecule. J. Exp. Med. 172:779– 88 Chicz RM, Urban RG, Gorga JC, Vignali DA, Lane WS, Strominger JL. 1993. Speciﬁcity and promiscuity among naturally processed peptides bound to HLA-DR alleles. J. Exp. Med. 178:27–47 Weiss S, Bogen B. 1989. B-lymphoma cells process and present their endogenous immunoglobulin to major histocompatibility complex-restricted T cells. Proc. Natl. Acad. Sci. USA 86:282–86 Dengjel J, Schoor O, Fischer R, Reich M, Kraus M, et al. 2005. Autophagy promotes MHC class II presentation of peptides from intracellular source proteins. Proc. Natl. Acad. Sci. USA 102:7922–27 Nimmerjahn F, Milosevic S, Behrends U, Jaffee EM, Pardoll DM, et al. 2003. Major histocompatibility complex class II-restricted presentation of a cytosolic antigen by autophagy. Eur. J. Immunol. 33:1250–59

Boon et al.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

ARI

23 February 2006

15:18

65. Paludan C, Schmid D, Landthaler M, Vockerodt M, Kube D, et al. 2005. Endogenous MHC class II processing of a viral nuclear antigen after autophagy. Science 307:593–96 66. Vijayasaradhi S, Xu Y, Bouchard B, Houghton AN. 1995. Intracellular sorting and targeting of melanosomal membrane proteins: identiﬁcation of signals for sorting of the human brown locus protein, gp75. J. Cell Biol. 130:807–20 67. Wang S, Bartido S, Yang G, Qin J, Moroi Y, et al. 1999. A role for a melanosome transport signal in accessing the MHC class II presentation pathway and in eliciting CD4+ T cell responses. J. Immunol. 163:5820–26 68. Chaux P, Vantomme V, Stroobant V, Thielemans K, Corthals J, et al. 1999. Identiﬁcation of MAGE-3 epitopes presented by HLA-DR molecules to CD4+ T lymphocytes. J. Exp. Med. 189:767–78 69. Manici S, Sturniolo T, Imro MA, Hammer J, Sinigaglia F, et al. 1999. Melanoma cells present a MAGE-3 epitope to CD4+ cytotoxic T cells in association with histocompatibility leukocyte antigen DR11. J. Exp. Med. 189:871–76 70. Schultz ES, Lethe B, Cambiaso CL, Van Snick J, Chaux P, et al. 2000. A MAGE-A3 peptide presented by HLA-DP4 is recognized on tumor cells by CD4+ cytolytic T lymphocytes. Cancer Res. 60:6272–75 71. Zhang Y, Chaux P, Stroobant V, Eggermont AM, Corthals J, et al. 2003. A MAGE-3 peptide presented by HLA-DR1 to CD4+ T cells that were isolated from a melanoma patient vaccinated with a MAGE-3 protein. J. Immunol. 171:219–25 72. Lin KY, Guarnieri FG, Staveley-O’Carroll KF, Levitsky HI, August JT, et al. 1996. Treatment of established tumors with a novel vaccine that enhances major histocompatibility class II presentation of tumor antigen. Cancer Res. 56:21–26 73. Lin X, Dashti A, Schinazi RF, Tang J. 1993. Intracellular diversion of glycoprotein GP160 of human immunodeﬁciency virus to lysosomes as a strategy of AIDS gene therapy. FASEB J. 7:1070–80 74. Rowell JF, Ruff AL, Guarnieri FG, Staveley-O’Carroll K, Lin X, et al. 1995. Lysosomeassociated membrane protein-1-mediated targeting of the HIV-1 envelope protein to an endosomal/lysosomal compartment enhances its presentation to MHC class II-restricted T cells. J. Immunol. 155:1818–28 75. Ruff AL, Guarnieri FG, Staveley-O’Carroll K, Siliciano RF, August JT. 1997. The enhanced immune response to the HIV gp160/LAMP chimeric gene product targeted to the lysosome membrane protein trafﬁcking pathway. J. Biol. Chem. 272:8671–78 76. Thomson SA, Burrows SR, Misko IS, Moss DJ, Coupar BE, Khanna R. 1998. Targeting a polyepitope protein incorporating multiple class II-restricted viral epitopes to the secretory/endocytic pathway facilitates immune recognition by CD4+ cytotoxic T lymphocytes: a novel approach to vaccine design. J. Virol. 72:2246–52 77. Wu TC, Guarnieri FG, Staveley-O’Carroll KF, Viscidi RP, Levitsky HI, et al. 1995. Engineering an intracellular pathway for major histocompatibility complex class II presentation of antigens. Proc. Natl. Acad. Sci. USA 92:11671–75 78. Bonehill A, Heirman C, Tuyaerts S, Michiels A, Breckpot K, et al. 2004. Messenger RNAelectroporated dendritic cells presenting MAGE-A3 simultaneously in HLA class I and class II molecules. J. Immunol. 172:6649–57 79. Yewdell JW, Anton LC, Bennink JR. 1996. Defective ribosomal products (DRiPs): a major source of antigenic peptides for MHC class I molecules? J. Immunol. 157:1823–26 80. Yu Z, Theoret MR, Touloukian CE, Surman DR, Garman SC, et al. 2004. Poor immunogenicity of a self/tumor antigen derives from peptide-MHC-I instability and is independent of tolerance. J. Clin. Invest. 114:551–59 www.annualreviews.org • Human T Cell Responses Against Melanoma

199

ARI

23 February 2006

15:18

81. Parkhurst MR, Salgaller ML, Southwood S, Robbins PF, Sette A, et al. 1996. Improved induction of melanoma-reactive CTL with peptides from the melanoma antigen gp100 modiﬁed at HLA-A∗ 0201-binding residues. J. Immunol. 157:2539–48 82. Pass HA, Schwarz SL, Wunderlich JR, Rosenberg SA. 1998. Immunization of patients with melanoma peptide vaccines: immunologic assessment using the ELISPOT assay. Cancer J. Sci. Am. 4:316–23 83. Clay TM, Custer MC, McKee MD, Parkhurst M, Robbins PF, et al. 1999. Changes in the ﬁne speciﬁcity of gp100209 –217 -reactive T cells in patients following vaccination with a peptide modiﬁed at an HLA-A2.1 anchor residue. J. Immunol. 162:1749–55 84. Valmori D, Fonteneau JF, Lizana CM, Gervois N, Lienard D, et al. 1998. Enhanced generation of speciﬁc tumor-reactive CTL in vitro by selected Melan-A/MART-1 immunodominant peptide analogues. J. Immunol. 160:1750–58 85. Rivoltini L, Squarcina P, Loftus DJ, Castelli C, Tarsini P, et al. 1999. A superagonist variant of peptide MART1/Melan A27–35 elicits anti-melanoma CD8+ T cells with enhanced functional characteristics: implication for more effective immunotherapy. Cancer Res. 59:301–6 86. Valmori D, Levy F, Miconnet I, Zajac P, Spagnoli GC, et al. 2000. Induction of potent antitumor CTL responses by recombinant vaccinia encoding a melan-A peptide analogue. J. Immunol. 164:1125–31 87. Dutoit V, Rubio-Godoy V, Pittet MJ, Zippelius A, Dietrich PY, et al. 2002. Degeneracy of antigen recognition as the molecular basis for the high frequency of naive A2/Melan-A peptide multimer+ CD8+ T cells in humans. J. Exp. Med. 196:207–16 88. Ayyoub M, Zippelius A, Pittet MJ, Rimoldi D, Valmori D, et al. 2003. Activation of human melanoma reactive CD8+ T cells by vaccination with an immunogenic peptide analog derived from Melan-A/melanoma antigen recognized by T cells. Clin. Cancer Res. 9:669–77 89. Chen JL, Dunbar PR, Gileadi U, Jager E, Gnjatic S, et al. 2000. Identiﬁcation of NYESO-1 peptide analogues capable of improved stimulation of tumor-reactive CTL. J. Immunol. 165:948–55 90. Romero P, Dutoit V, Rubio-Godoy V, Lienard D, Speiser D, et al. 2001. CD8+ T-cell response to NY-ESO-1: relative antigenicity and in vitro immunogenicity of natural and analogue sequences. Clin. Cancer Res. 7:S766–72 91. Rosenberg SA, Packard BS, Aebersold PM, Solomon D, Topalian SL, et al. 1988. Use of tumor-inﬁltrating lymphocytes and interleukin-2 in the immunotherapy of patients with metastatic melanoma. A preliminary report. N. Engl. J. Med. 319:1676–80 92. Yannelli JR, Hyatt C, McConnell S, Hines K, Jacknin L, et al. 1996. Growth of tumorinﬁltrating lymphocytes from human solid cancers: summary of a 5-year experience. Int. J. Cancer 65:413–21 93. Rosenberg SA. 2001. Progress in the development of immunotherapy for the treatment of patients with cancer. J. Intern. Med. 250:462–75 94. Coulie PG, Somville M, Lehmann F, Hainaut P, Brasseur F, et al. 1992. Precursor frequency analysis of human cytolytic T lymphocytes directed against autologous melanoma cells. Int. J. Cancer 50:289–97 95. Mazzocchi A, Belli F, Mascheroni L, Vegetti C, Parmiani G, Anichini A. 1994. Frequency of cytotoxic T lymphocyte precursors (CTLp) interacting with autologous tumor via the T-cell receptor: limiting dilution analysis of speciﬁc CTLp in peripheral blood and tumor-invaded lymph nodes of melanoma patients. Int. J. Cancer 58:330– 39

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

200

Boon et al.

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23 February 2006

15:18

96. Sensi M, Parmiani G. 1995. Analysis of TCR usage in human tumors: a new tool for assessing tumor-speciﬁc immune responses. Immunol. Today 16:588–95 97. Germeau C, Ma W, Schiavetti F, Lurquin C, Henry E, et al. 2005. High frequency of antitumor T cells in the blood of melanoma patients before and after vaccination with tumor antigens. J. Exp. Med. 201:241–48 98. Letsch A, Keilholz U, Schadendorf D, Nagorsen D, Schmittel A, et al. 2000. High frequencies of circulating melanoma-reactive CD8+ T cells in patients with advanced melanoma. Int. J. Cancer 87:659–64 99. Chaux P, Vantomme V, Coulie P, Boon T, van der Bruggen P. 1998. Estimation of the frequencies of anti-MAGE-3 cytolytic T-lymphocyte precursors in blood from individuals without cancer. Int. J. Cancer 77:538–42 100. Lonchay C, van der Bruggen P, Connerotte T, Hanagiri T, Coulie P, et al. 2004. Correlation between tumor regression and T cell responses in melanoma patients vaccinated with a MAGE antigen. Proc. Natl. Acad. Sci. USA 101(Suppl. 2):14631–38 101. Hanagiri T, van Baren N, Neyns B, Boon T, Coulie PG. 2006. Analysis of one of the rare melanoma patients with a spontaneous CTL response to a MAGE-A3 peptide presented by HLA-A1. Cancer Immunol. Immunother. 55:178–84 102. Jager E, Nagata Y, Gnjatic S, Wada H, Stockert E, et al. 2000. Monitoring CD8 T cell responses to NY-ESO-1: correlation of humoral and cellular immune responses. Proc. Natl. Acad. Sci. USA 97:4760–65 103. Jager E, Gnjatic S, Nagata Y, Stockert E, Jager D, et al. 2000. Induction of primary NY-ESO-1 immunity: CD8+ T lymphocyte and antibody responses in peptidevaccinated patients with NY-ESO-1+ cancers. Proc. Natl. Acad. Sci. USA 97:12198– 203 104. Valmori D, Dutoit V, Lienard D, Rimoldi D, Pittet MJ, et al. 2000. Naturally occurring human lymphocyte antigen-A2 restricted CD8+ T-cell response to the cancer testis antigen NY-ESO-1 in melanoma patients. Cancer Res. 60:4499–506 105. Grifﬁoen M, Borghi M, Schrier PI, Osanto S. 2001. Detection and quantiﬁcation of CD8+ T cells speciﬁc for HLA-A∗ 0201-binding melanoma and viral peptides by the IFN-γ-ELISPOT assay. Int. J. Cancer 93:549–55 106. Zorn E, Hercend T. 1999. A MAGE-6-encoded peptide is recognized by expanded lymphocytes inﬁltrating a spontaneously regressing human primary melanoma lesion. Eur. J. Immunol. 29:602–7 107. Ferradini L, Mackensen A, Genevee C, Bosq J, Duvillard P, et al. 1993. Analysis of T cell receptor variability in tumor-inﬁltrating lymphocytes from a human regressive melanoma. Evidence for in situ T cell clonal expansion. J. Clin. Invest. 91:1183–90 108. Mackensen A, Ferradini L, Carcelain G, Triebel F, Faure F, et al. 1993. Evidence for in situ ampliﬁcation of cytotoxic T-lymphocytes with antitumor activity in a human regressive melanoma. Cancer Res. 53:3569–73 109. Lurquin C, Lethe B, De Plaen E, Corbiere V, Theate I, et al. 2005. Contrasting frequencies of antitumor and anti-vaccine T cells in metastases of a melanoma patient vaccinated with a MAGE tumor antigen. J. Exp. Med. 201:249–57 110. Wang HY, Lee DA, Peng G, Guo Z, Li Y, et al. 2004. Tumor-speciﬁc human CD4+ regulatory T cells and their ligands: implications for immunotherapy. Immunity 20:107– 18 111. Zorn E, Hercend T. 1999. A natural cytotoxic T cell response in a spontaneously regressing human melanoma targets a neoantigen resulting from a somatic point mutation. Eur. J. Immunol. 29:592–601 www.annualreviews.org • Human T Cell Responses Against Melanoma

201

ARI

23 February 2006

15:18

112. Novellino L, Renkvist N, Rini F, Mazzocchi A, Rivoltini L, et al. 2003. Identiﬁcation of a mutated receptor-like protein tyrosine phosphatase κ as a novel, class II HLA-restricted melanoma antigen. J. Immunol. 170:6363–70 113. Sensi M, Nicolini G, Zanon M, Colombo C, Molla A, et al. 2005. Immunogenicity without immunoselection: a mutant but functional antioxidant enzyme retained in a human metastatic melanoma and targeted by CD8+ T cells with a memory phenotype. Cancer Res. 65:632–40 114. Robbins PF, El-Gamil M, Li YF, Zeng G, Dudley M, Rosenberg SA. 2002. Multiple HLA class II-restricted melanocyte differentiation antigens are recognized by tumor-inﬁltrating lymphocytes from a patient with melanoma. J. Immunol. 169:6036–47 115. Topalian SL, Gonzales MI, Parkhurst M, Li YF, Southwood S, et al. 1996. Melanomaspeciﬁc CD4+ T cells recognize nonmutated HLA-DR-restricted tyrosinase epitopes. J. Exp. Med. 183:1965–71 116. Romero P, Valmori D, Pittet MJ, Zippelius A, Rimoldi D, et al. 2002. Antigenicity and immunogenicity of Melan-A/MART-1 derived peptides as targets for tumor reactive CTL in human melanoma. Immunol. Rev. 188:81–96 117. Zippelius A, Pittet MJ, Batard P, Rufer N, de Smedt M, et al. 2002. Thymic selection generates a large T cell pool recognizing a self-peptide in humans. J. Exp. Med. 195:485–94 118. Dunbar PR, Smith CL, Chao D, Salio M, Shepherd D, et al. 2000. A shift in the phenotype of melan-A-speciﬁc CTL identiﬁes melanoma patients with an active tumor-speciﬁc immune response. J. Immunol. 165:6644–52 119. Lee PP, Yee C, Savage PA, Fong L, Brockstedt D, et al. 1999. Characterization of circulating T cells speciﬁc for tumor-associated antigens in melanoma patients. Nat. Med. 5:677–85 120. Palermo B, Campanelli R, Mantovani S, Lantelme E, Manganoni AM, et al. 2001. Diverse expansion potential and heterogeneous avidity in tumor-associated antigen-speciﬁc T lymphocytes from primary melanoma patients. Eur. J. Immunol. 31:412–20 121. Halpern AC, Schuchter LM. 1997. Prognostic models in melanoma. Semin. Oncol. 24:S2–7 122. Clemente CG, Mihm MC Jr, Bufalino R, Zurrida S, Collini P, Cascinelli N. 1996. Prognostic value of tumor inﬁltrating lymphocytes in the vertical growth phase of primary cutaneous melanoma. Cancer 77:1303–10 123. Mihm MC Jr, Clemente CG, Cascinelli N. 1996. Tumor inﬁltrating lymphocytes in lymph node melanoma metastases: a histopathologic prognostic indicator and an expression of local immune response. Lab. Invest. 74:43–47 124. van Oijen M, Bins A, Elias S, Sein J, Weder P, et al. 2004. On the role of melanoma-speciﬁc CD8+ T-cell immunity in disease progression of advanced-stage melanoma patients. Clin. Cancer Res. 10:4754–60 125. Vesalainen S, Lipponen P, Talja M, Syrjanen K. 1994. Histological grade, perineural inﬁltration, tumour-inﬁltrating lymphocytes and apoptosis as determinants of long-term prognosis in prostatic adenocarcinoma. Eur. J. Cancer 30A:1797–803 126. Naito Y, Saito K, Shiiba K, Ohuchi A, Saigenji K, et al. 1998. CD8+ T cells inﬁltrated within cancer cell nests as a prognostic factor in human colorectal cancer. Cancer Res. 58:3491–94 127. Schumacher K, Haensch W, Roefzaad C, Schlag PM. 2001. Prognostic signiﬁcance of activated CD8+ T cell inﬁltrations within esophageal carcinomas. Cancer Res. 61:3932–36 128. Zhang L, Conejo-Garcia JR, Katsaros D, Gimotty PA, Massobrio M, et al. 2003. Intratumoral T cells, recurrence, and survival in epithelial ovarian cancer. N. Engl. J. Med. 348:203–13

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

202

Boon et al.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

ARI

23 February 2006

15:18

129. Molldrem JJ, Lee PP, Wang C, Felio K, Kantarjian HM, et al. 2000. Evidence that speciﬁc T lymphocytes may participate in the elimination of chronic myelogenous leukemia. Nat. Med. 6:1018–23 130. Marincola FM, Jaffee EM, Hicklin DJ, Ferrone S. 2000. Escape of human solid tumors from T-cell recognition: molecular mechanisms and functional signiﬁcance. Adv. Immunol. 74:181–273 131. Garcia-Lora A, Algarra I, Garrido F. 2003. MHC class I antigens, immune surveillance, and tumor immune escape. J. Cell. Physiol. 195:346–55 132. Kageshita T, Ishihara T, Campoli M, Ferrone S. 2005. Selective monomorphic and polymorphic HLA class I antigenic determinant loss in surgically removed melanoma lesions. Tissue Antigens 65:419–28 133. Yamshchikov GV, Mullins DW, Chang CC, Ogino T, Thompson L, et al. 2005. Sequential immune escape and shifting of T cell responses in a long-term survivor of melanoma. J. Immunol. 174:6863–71 134. Zippelius A, Batard P, Rubio-Godoy V, Bioley G, Lienard D, et al. 2004. Effector function of human tumor-speciﬁc CD8 T cells in melanoma lesions: a state of local functional tolerance. Cancer Res. 64:2865–73 135. Bronte V, Kasic T, Gri G, Gallana K, Borsellino G, et al. 2005. Boosting antitumor responses of T lymphocytes inﬁltrating human prostate cancers. J. Exp. Med. 201:1257– 68 136. Radoja S, Saio M, Schaer D, Koneru M, Vukmanovic S, Frey AB. 2001. CD8+ tumorinﬁltrating T cells are deﬁcient in perforin-mediated cytolytic activity due to defective microtubule-organizing center mobilization and lytic granule exocytosis. J. Immunol. 167:5042–51 137. Sakaguchi S, Fukuma K, Kuribayashi K, Masuda T. 1985. Organ-speciﬁc autoimmune diseases induced in mice by elimination of T cell subset. I. Evidence for the active participation of T cells in natural self-tolerance; deﬁcit of a T cell subset as a possible cause of autoimmune disease. J. Exp. Med. 161:72–87 138. Sakaguchi S. 2004. Naturally arising CD4+ regulatory T cells for immunologic selftolerance and negative control of immune responses. Annu. Rev. Immunol. 22:531– 62 139. Dieckmann D, Plottner H, Berchtold S, Berger T, Schuler G. 2001. Ex vivo isolation and characterization of CD4+ CD25+ T cells with regulatory properties from human blood. J. Exp. Med. 193:1303–10 140. Levings MK, Sangregorio R, Roncarolo MG. 2001. Human CD25+ CD4+ T regulatory cells suppress naive and memory T cell proliferation and can be expanded in vitro without loss of function. J. Exp. Med. 193:1295–302 141. Jonuleit H, Schmitt E, Stassen M, Tuettenberg A, Knop J, Enk AH. 2001. Identiﬁcation and functional characterization of human CD4+ CD25+ T cells with regulatory properties isolated from peripheral blood. J. Exp. Med. 193:1285–94 142. Wang HY, Peng G, Guo Z, Shevach EM, Wang RF. 2005. Recognition of a new ARTC1 peptide ligand uniquely expressed in tumor cells by antigen-speciﬁc CD4+ regulatory T cells. J. Immunol. 174:2661–70 143. Zea AH, Rodriguez PC, Atkins MB, Hernandez C, Signoretti S, et al. 2005. Arginaseproducing myeloid suppressor cells in renal cell carcinoma patients: a mechanism of tumor evasion. Cancer Res. 65:3044–48 144. Gorelik L, Flavell RA. 2001. Immune-mediated eradication of tumors through the blockade of transforming growth factor-β signaling in T cells. Nat. Med. 7:1118–22 www.annualreviews.org • Human T Cell Responses Against Melanoma

203

ARI

23 February 2006

15:18

145. Uyttenhove C, Pilotte L, Theate I, Stroobant V, Colau D, et al. 2003. Evidence for a tumoral immune resistance mechanism based on tryptophan degradation by indoleamine 2,3-dioxygenase. Nat. Med. 9:1269–74 146. Rubinstein N, Alvarez M, Zwirner NW, Toscano MA, Ilarregui JM, et al. 2004. Targeted inhibition of galectin-1 gene expression in tumor cells results in heightened T cellmediated rejection: a potential mechanism of tumor-immune privilege. Cancer Cell 5:241– 51 147. Chen Q, Daniel V, Maher DW, Hersey P. 1994. Production of IL-10 by melanoma cells: examination of its role in immunosuppression mediated by melanoma. Int. J. Cancer 56:755–60 148. Dong H, Strome SE, Salomao DR, Tamura H, Hirano F, et al. 2002. Tumor-associated B7-H1 promotes T-cell apoptosis: a potential mechanism of immune evasion. Nat. Med. 8:793–800 149. Speiser DE, Kyburz D, Stubi U, Hengartner H, Zinkernagel RM. 1992. Discrepancy between in vitro measurable and in vivo virus neutralizing cytotoxic T cell reactivities. Low T cell receptor speciﬁcity and avidity sufﬁcient for in vitro proliferation or cytotoxicity to peptide-coated target cells but not for in vivo protection. J. Immunol. 149:972– 80 150. Altman JD, Moss PA, Goulder PJ, Barouch DH, McHeyzer-Williams MG, et al. 1996. Phenotypic analysis of antigen-speciﬁc T lymphocytes. Science 274:94–96 151. Herr W, Schneider J, Lohse AW, Meyer zum Buschenfelde KH, Wolfel T. 1996. Detection and quantiﬁcation of blood-derived CD8+ T lymphocytes secreting tumor necrosis factor α in response to HLA-A2.1-binding melanoma and viral peptide antigens. J. Immunol. Methods 191:131–42 152. Herr W, Linn B, Leister N, Wandel E, Meyer zum Buschenfelde KH, Wolfel T. 1997. The use of computer-assisted video image analysis for the quantiﬁcation of CD8+ T lymphocytes producing tumor necrosis factor α spots in response to peptide antigens. J. Immunol. Methods 203:141–52 153. Novak EJ, Liu AW, Nepom GT, Kwok WW. 1999. MHC class II tetramers identify peptide-speciﬁc human CD4+ T cells proliferating in response to inﬂuenza A antigen. J. Clin. Invest. 104:R63–67 154. Zhang Y, Renkvist N, Sun Z, Schuler-Thurner B, Glaichenhaus N, et al. 2005. A polyclonal anti-vaccine CD4 T cell response detected with HLA-DP4 multimers in a melanoma patient vaccinated with MAGE-3.DP4-peptide-pulsed dendritic cells. Eur. J. Immunol. 35:1066–75 155. Schuler-Thurner B, Schultz ES, Berger TG, Weinlich G, Ebner S, et al. 2002. Rapid induction of tumor-speciﬁc type 1 T helper cells in metastatic melanoma patients by vaccination with mature, cryopreserved, peptide-loaded monocyte-derived dendritic cells. J. Exp. Med. 195:1279–88 156. Zhang Y, Sun Z, Nicolay H, Meyer RG, Renkvist N, et al. 2005. Monitoring of antivaccine CD4 T cell frequencies in melanoma patients vaccinated with a MAGE-3 protein. J. Immunol. 174:2404–11 157. Marchand M, Weynants P, Rankin E, Arienti F, Belli F, et al. 1995. Tumor regression responses in melanoma patients treated with a peptide encoded by gene MAGE-3. Int. J. Cancer 63:883–85 158. Ottaviani S, Zhang Y, Boon T, van der Bruggen P. 2005. A MAGE-1 antigenic peptide recognized by human cytolytic T lymphocytes on HLA-A2 tumor cells. Cancer Immunol. Immunother. 54:1214–20

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

204

Boon et al.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

ARI

23 February 2006

15:18

159. Marchand M, van Baren N, Weynants P, Brichard V, Dreno B, et al. 1999. Tumor regressions observed in patients with metastatic melanoma treated with an antigenic peptide encoded by gene MAGE-3 and presented by HLA-A1. Int. J. Cancer 80:219–30 160. Coulie PG, Karanikas V, Colau D, Lurquin C, Landry C, et al. 2001. A monoclonal cytolytic T-lymphocyte response observed in a melanoma patient vaccinated with a tumorspeciﬁc antigenic peptide encoded by gene MAGE-3. Proc. Natl. Acad. Sci. USA 98:10290– 95 161. Karanikas V, Lurquin C, Colau D, van Baren N, De Smet C, et al. 2003. Monoclonal anti-MAGE-3 CTL responses in melanoma patients displaying tumor regression after vaccination with a recombinant canarypox virus. J. Immunol. 171:4898–904 162. Thurner B, Haendle I, Roder C, Dieckmann D, Keikavoussi P, et al. 1999. Vaccination with mage-3A1 peptide-pulsed mature, monocyte-derived dendritic cells expands speciﬁc cytotoxic T cells and induces regression of some metastases in advanced stage IV melanoma. J. Exp. Med. 190:1669–78 163. Godelaine D, Carrasco J, Lucas S, Karanikas V, Schuler-Thurner B, et al. 2003. Polyclonal CTL responses observed in melanoma patients vaccinated with dendritic cells pulsed with a MAGE-3.A1 peptide. J. Immunol. 171:4893–97 164. Paczesny S, Banchereau J, Wittkowski KM, Saracino G, Fay J, Palucka AK. 2004. Expansion of melanoma-speciﬁc cytolytic CD8+ T cell precursors in patients with metastatic melanoma vaccinated with CD34+ progenitor-derived dendritic cells. J. Exp. Med. 199:1503–11 165. Gajewski TF, Fallarino F, Ashikari A, Sherman M. 2001. Immunization of HLA-A2+ melanoma patients with MAGE-3 or MelanA peptide-pulsed autologous peripheral blood mononuclear cells plus recombinant human interleukin 12. Clin. Cancer Res. 7:S895–901 166. Chianese-Bullock KA, Pressley J, Garbee C, Hibbitts S, Murphy C, et al. 2005. MAGEA1-, MAGE-A10-, and gp100-derived peptides are immunogenic when combined with granulocyte-macrophage colony-stimulating factor and montanide ISA-51 adjuvant and administered as part of a multipeptide vaccine for melanoma. J. Immunol. 174:3080–86 167. Bettinotti MP, Panelli MC, Ruppe E, Mocellin S, Phan GQ, et al. 2003. Clinical and immunological evaluation of patients with metastatic melanoma undergoing immunization with the HLA-Cw∗ 0702-associated epitope MAGE-A12:170–178. Int. J. Cancer 105:210– 16 168. Marchand M, Punt CJ, Aamdal S, Escudier B, Kruit WH, et al. 2003. Immunisation of metastatic cancer patients with MAGE-3 protein combined with adjuvant SBAS-2: a clinical report. Eur. J. Cancer 39:70–77 169. Kruit WH, van Ojik HH, Brichard VG, Escudier B, Dorval T, et al. 2005. Phase 1/2 study of subcutaneous and intradermal immunization with a recombinant MAGE-3 protein in patients with detectable metastatic melanoma. Int. J. Cancer 117:596–604 170. Davis ID, Chen W, Jackson H, Parente P, Shackleton M, et al. 2004. Recombinant NY-ESO-1 protein with ISCOMATRIX adjuvant induces broad integrated antibody and CD4+ and CD8+ T cell responses in humans. Proc. Natl. Acad. Sci. USA 101:10697– 702 171. Wang F, Bade E, Kuniyoshi C, Spears L, Jeffery G, et al. 1999. Phase I trial of a MART-1 peptide vaccine with incomplete Freund’s adjuvant for resected high-risk melanoma. Clin. Cancer Res. 5:2756–65 172. Pittet MJ, Speiser DE, Lienard D, Valmori D, Guillaume P, et al. 2001. Expansion and functional maturation of human tumor antigen-speciﬁc CD8+ T cells after vaccination with antigenic peptide. Clin. Cancer Res. 7:S796–803 www.annualreviews.org • Human T Cell Responses Against Melanoma

205

ARI

23 February 2006

15:18

173. Cormier JN, Salgaller ML, Prevette T, Barracchini KC, Rivoltini L, et al. 1997. Enhancement of cellular immunity in melanoma patients immunized with a peptide from MART-1/Melan A. Cancer J. Sci. Am. 3:37–44 174. Speiser DE, Lienard D, Rufer N, Rubio-Godoy V, Rimoldi D, et al. 2005. Rapid and strong human CD8+ T cell responses to vaccination with peptide, IFA, and CpG oligodeoxynucleotide 7909. J. Clin. Invest. 115:739–46 175. Mackensen A, Herbst B, Chen JL, Kohler G, Noppen C, et al. 2000. Phase I study in melanoma patients of a vaccine with peptide-pulsed dendritic cells generated in vitro from CD34+ hematopoietic progenitor cells. Int. J. Cancer 86:385–92 176. Oertli D, Marti WR, Zajac P, Noppen C, Kocher T, et al. 2002. Rapid induction of speciﬁc cytotoxic T lymphocytes against melanoma-associated antigens by a recombinant vaccinia virus vector expressing multiple immunodominant epitopes and costimulatory molecules in vivo. Hum. Gene Ther. 13:569–75 177. Smith CL, Dunbar PR, Mirza F, Palmowski MJ, Shepherd D, et al. 2005. Recombinant modiﬁed vaccinia Ankara primes functionally activated CTL speciﬁc for a melanoma tumor antigen epitope in melanoma patients with a high risk of disease recurrence. Int. J. Cancer 113:259–66 178. Cebon J, Jager E, Shackleton MJ, Gibbs P, Davis ID, et al. 2003. Two phase I studies of low dose recombinant human IL-12 with Melan-A and inﬂuenza peptides in subjects with advanced malignant melanoma. Cancer Immun. 3:7–12 179. Sanderson K, Scotland R, Lee P, Liu D, Groshen S, et al. 2005. Autoimmunity in a phase I trial of a fully human anti-cytotoxic T-lymphocyte antigen-4 monoclonal antibody with multiple melanoma peptides and Montanide ISA 51 for patients with resected stages III and IV melanoma. J. Clin. Oncol. 23:741–50 180. Rosenberg SA, Yang JC, Schwartzentruber DJ, Hwu P, Marincola FM, et al. 1998. Immunologic and therapeutic evaluation of a synthetic peptide vaccine for the treatment of patients with metastatic melanoma. Nat. Med. 4:321–27 181. Smith JW 2nd, Walker EB, Fox BA, Haley D, Wisner KP, et al. 2003. Adjuvant immunization of HLA-A2-positive melanoma patients with a modiﬁed gp100 peptide induces peptide-speciﬁc CD8+ T-cell responses. J. Clin. Oncol. 21:1562–73 182. Lee P, Wang F, Kuniyoshi J, Rubio V, Stuges T, et al. 2001. Effects of interleukin-12 on the immune response to a multipeptide vaccine for resected metastatic melanoma. J. Clin. Oncol. 19:3836–47 183. Yamshchikov GV, Barnd DL, Eastham S, Galavotti H, Patterson JW, et al. 2001. Evaluation of peptide vaccine immunogenicity in draining lymph nodes and peripheral blood of melanoma patients. Int. J. Cancer 92:703–11 184. Slingluff CL Jr, Petroni GR, Yamshchikov GV, Hibbitts S, Grosh WW, et al. 2004. Immunologic and clinical outcomes of vaccination with a multiepitope melanoma peptide vaccine plus low-dose interleukin-2 administered either concurrently or on a delayed schedule. J. Clin. Oncol. 22:4474–85 185. Schaed SG, Klimek VM, Panageas KS, Musselli CM, Butterworth L, et al. 2002. T-cell responses against tyrosinase 368–376 (370D) peptide in HLA∗ A0201+ melanoma patients: randomized trial comparing incomplete Freund’s adjuvant, granulocyte macrophage colony-stimulating factor, and QS-21 as immunological adjuvants. Clin. Cancer Res. 8:967– 72 186. Banchereau J, Palucka AK, Dhodapkar M, Burkeholder S, Taquet N, et al. 2001. Immune and clinical responses in patients with metastatic melanoma to CD34+ progenitor-derived dendritic cell vaccine. Cancer Res. 61:6451–58

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

206

Boon et al.

Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-06

ARI

23 February 2006

15:18

187. van Baren N, Bonnet M-C, Dr´eno B, Khammari A, Dorval T, et al. 2006. Tumoral and immunological responses following vaccination of metastatic melanoma patients with an ALVAC virus encoding two Mage antigens recognized by T cells. J. Clin. Oncol. In press 188. Slingluff CL Jr, Petroni GR, Yamshchikov GV, Barnd DL, Eastham S, et al. 2003. Clinical and immunologic results of a randomized phase II trial of vaccination using four melanoma peptides either administered in granulocyte-macrophage colony-stimulating factor in adjuvant or pulsed on dendritic cells. J. Clin. Oncol. 21:4016–26 189. Mitchell MS, Harel W, Kempf RA, Hu E, Kan-Mitchell J, et al. 1990. Active-speciﬁc immunotherapy for melanoma. J. Clin. Oncol. 8:856–69 190. Rosenberg SA, Yang JC, Restifo NP. 2004. Cancer immunotherapy: moving beyond current vaccines. Nat. Med. 10:909–15 191. Hersey P, Menzies SW, Halliday GM, Nguyen T, Farrelly ML, et al. 2004. Phase I/II study of treatment with dendritic cell vaccines in patients with disseminated melanoma. Cancer Immunol. Immunother. 53:125–34 192. Butterﬁeld LH, Ribas A, Dissette VB, Amarnani SN, Vu HT, et al. 2003. Determinant spreading associated with clinical response in dendritic cell-based immunotherapy for malignant melanoma. Clin. Cancer Res. 9:998–1008 193. de Vries IJ, Lesterhuis WJ, Scharenborg NM, Engelen LP, Ruiter DJ, et al. 2003. Maturation of dendritic cells is a prerequisite for inducing immune responses in advanced melanoma patients. Clin. Cancer Res. 9:5091–100 194. Lau R, Wang F, Jeffery G, Marty V, Kuniyoshi J, et al. 2001. Phase I trial of intravenous peptide-pulsed dendritic cells in patients with metastatic melanoma. J. Immunother. 24:66– 78 195. Baldo M, Schiavon M, Cicogna PA, Boccato P, Mazzoleni F. 1992. Spontaneous regression of subcutaneous metastasis of cutaneous melanoma. Plast. Reconstr. Surg. 90:1073– 76 196. Letsch A, Keilholz U, Fluck M, Nagorsen D, Asemissen AM, et al. 2005. Peptide vaccination after repeated resection of metastases can induce a prolonged relapse-free interval in melanoma patients. Int. J. Cancer 114:936–41 197. Rosenberg SA, Dudley ME. 2004. Cancer regression in patients with metastatic melanoma after the transfer of autologous antitumor lymphocytes. Proc. Natl. Acad. Sci. USA 101(Suppl. 2):14639–45 198. Dr´eno B, Nguyen JM, Khammari A, Pandolﬁno MC, Tessier MH, et al. 2002. Randomized trial of adoptive transfer of melanoma tumor-inﬁltrating lymphocytes as adjuvant therapy for stage III melanoma. Cancer Immunol. Immunother. 51:539–46 199. Lienard D, Rimoldi D, Marchand M, Dietrich PY, van Baren N, et al. 2004. Ex vivo detectable activation of Melan-A-speciﬁc T cells correlating with inﬂammatory skin reactions in melanoma patients vaccinated with peptides in IFA. Cancer Immun. 4:4 200. Bevan MJ. 2004. Helping the CD8+ T-cell response. Nat. Rev. Immunol. 4:595–602 201. Weber JS, Hua FL, Spears L, Marty V, Kuniyoshi C, Celis E. 1999. A phase I trial of an HLA-A1 restricted MAGE-3 epitope peptide with incomplete Freund’s adjuvant in patients with resected high-risk melanoma. J. Immunother. 22:431–40 202. Phan GQ, Yang JC, Sherry RM, Hwu P, Topalian SL, et al. 2003. Cancer regression and autoimmunity induced by cytotoxic T lymphocyte-associated antigen 4 blockade in patients with metastatic melanoma. Proc. Natl. Acad. Sci. USA 100:8372–77 203. Marincola FM, Rivoltini L, Salgaller ML, Player M, Rosenberg SA. 1996. Differential anti-MART-1/MelanA CTL activity in peripheral blood of HLA-A2 melanoma patients www.annualreviews.org • Human T Cell Responses Against Melanoma

207

ANRV270-IY24-06

ARI

23 February 2006

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Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

in comparison to healthy donors: evidence of in vivo priming by tumor cells. J. Immunother. Emphasis Tumor Immunol. 19:266–77 204. Wang E, Miller LD, Ohnmacht GA, Mocellin S, Perez-Diez A, et al. 2002. Prospective molecular proﬁling of melanoma metastases suggests classiﬁers of immune responsiveness. Cancer Res. 62:3581–86

208

Boon et al.

Contents

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:175-208. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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FOXP3: Of Mice and Men Annu. Rev. Immunol. 2006.24:209-226. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Steven F. Ziegler Immunology Program, Benaroya Research Institute and Department of Immunology, University of Washington School of Medicine, Seattle, Washington 98101; email: [email protected]

Annu. Rev. Immunol. 2006. 24:209–26 First published online as a Review in Advance on December 1, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090547 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0209$20.00

Key Words regulatory T cell, autoimmunity, forkhead, transcription factor

Abstract The immune system has evolved mechanisms to recognize and eliminate threats, as well as to protect against self-destruction. Tolerance to self-antigens is generated through two fundamental mechanisms: (a) elimination of self-reactive cells in the thymus during selection and (b) generation of a variety of peripheral regulatory cells to control self-reactive cells that escape the thymus. It is becoming increasing apparent that a population of thymically derived CD4+ regulatory T cells, exempliﬁed by the expression of the IL-2Rα chain, is essential for the maintenance of peripheral tolerance. Recent work has shown that the forkhead family transcription factor Foxp3 is critically important for the development and function of the regulatory T cells. Lack of Foxp3 leads to development of fatal autoimmune lymphoproliferative disease; furthermore, ectopic Foxp3 expression can phenotypically convert effector T cells to regulatory T cells. This review focuses on Foxp3 expression and function and highlights differences between humans and mice regarding Foxp3 regulation.

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INTRODUCTION Foxp3: forkhead box protein P3

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Tregs: regulatory T cells CD4+ CD25+ Treg: a subset of CD4+ T cells that is capable of suppressing the proliferation and cytokine production of naive or memory T cells Forkhead family: a large family of transcriptional regulators named after the founding member, which was found to be the gene responsible for the forkhead mutation in Drosophila. All members of the family have a closely conserved motif, known as a forkhead box, or Fox, that is involved in DNA binding. The family is further subdivided based on homology outside the forkhead box.

Immunological tolerance to self-antigens is a tightly regulated process. A primary mechanism for self-tolerance is deletion of selfreactive cells in the thymus. However, this mechanism is not perfect, and autoreactive clones do escape into the periphery. Tolerance is maintained in the periphery through a variety of mechanisms, including a population of regulatory T cells that actively suppress the function of autoreactive T cells. These T cells, identiﬁed by their expression of CD4, the IL-2Rα chain (CD25), and the forkhead family transcription factor Foxp3, are know as regulatory T cells, or Tregs. They have the ability to inhibit the development of autoimmunity when transferred into the appropriate host. Recent work has shown that the forkhead/winged-helix protein Foxp3 is expressed predominantly in Tregs and is both necessary and sufﬁcient for their development and function. Several excellent reviews have been written recently on the development and function of CD4+ CD25+ Tregs (1–5). Thus, this review does not directly address these issues. Instead, I review what is known concerning the expression and function of Foxp3, which is critical for the development and function of Tregs, especially in the mouse. Although the overall importance of Foxp3 is obvious from the phenotype of humans and mice that lack this protein, very little is actually known about how it functions and what controls its expression. This review highlights the similarities and differences in Foxp3 expression function in human and mouse, with an emphasis on the differences.

IDENTIFICATION AND CHARACTERIZATION OF MOUSE Foxp3 Characterization of Scurfy (sf ) Mutant Mice The scurfy (sf ) mutation arose spontaneously in the partially inbred MR strain in 1949 at the 210

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Oak Ridge National Laboratory. Subsequent studies showed the gene to be X-linked, with only Xsf /Y males affected (here, these mice are referred to as sf/Y). Grossly, affected sf/Y males exhibit several external markers of disease, including ear thickening; scaling of the ears, eyelids, feet, and tail; and severe runting. Internally, the mice exhibit lymphadenopathy, splenomegaly, hepatomegaly, and massive lymphocytic inﬁltrates in the skin and liver. These lesions closely resemble those seen in a graft-versus-host reaction. Mice display anemia and have a positive direct Coombs’s test, but they lack evidence of autoantibodies against double-strand DNA or small ribonuclear protein antigens (6). Taken together, these gross assessments suggest that sf is a mutation that causes an autoimmune-like disease in affected animals. In this regard, sf/Y mice resemble mice bearing targeted mutations in the ctla-4 or tgf-β1 genes. These mice die at approximately three weeks of age from a massive lymphoproliferative disease, with peripheral lymphocyte levels up to 20-fold greater than normal mice (7, 8). Phenotypically, scurfy disease is most consistent with a diagnosis of autoimmune lymphoproliferative disease. This idea is supported by the ﬁnding that mice expressing a transgenic T cell receptor (TCR) survive signiﬁcantly longer in the absence of antigen stimulation (60 days compared with 20– 24 days) and that they live a normal lifespan and are free of sf disease when they are also rag-2-null (sf/ova/rag mice). Finally, T cells from sf/Y mice display reduced sensitivity to inhibitors of T cell activation, suggesting that TCR signaling is dysregulated in sf/Y mice. Taken together, these data suggest the sf disease results from an inability to properly regulate antigen-driven T cell activation. One of the in vitro hallmarks of sf disease is the spontaneous proliferation and cytokine production exhibited by T cells isolated from sf/Y mice. Initial studies showed that cytokine production in vitro by ConAstimulated sf/Y splenocytes was greatly elevated (9). More detailed studies, using puriﬁed

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CD4+ T cells from sf/Y and normal littermate control (NLC) mice, showed that these cells were capable of proliferation and cytokine production when examined directly ex vivo. However, to achieve maximal T cell stimulation, TCR engagement was required, suggesting that the defect in sf mice is not nonspeciﬁc T cell activation. CD4+ T cells from sf/Y mice were hyperresponsive to TCR-mediated signals, responding to TCR engagement with crosslinking antibodies at concentrations 10-fold lower than their wild-type counterparts (10). Consistent with the increased expression of costimulatory molecules in sf/Y mice, CD4+ T cells from these mice responded to CD28 crosslinking (10). Recent studies have shown that the transcription factors NFAT and AP-1 are constitutively activated in CD4+ T cells from sf/Y mice (11). Also, these same T cells have a marked decrease in their sensitivity to cyclosporin A, with an IC50 15-fold higher than NLCderived T cells (10). Both observations are indicative of an alteration in TCR-mediated signal transduction in CD4+ T cells from sf/Y mice and may account for the increased cytokine production exhibited by these cells.

Consequence of Foxp3 Expression on CD4+ T Cell Function The gene responsible for scurfy disease was molecularly cloned using a standard positional cloning approach (12). Sequence analysis of the cloned gene showed that it encoded a novel member of the forkhead family of transcriptional regulators, Foxp3 (see below for more details). To verify this gene as the gene responsible for the scurfy mutation, transgenic mice were generated using a cosmid clone containing mouse Foxp3. When bred to sf/Y mice, expression of the transgene was capable of rescuing the mice from scurfy disease and demonstrated that a mutation in Foxp3 was indeed responsible for the scurfy phenotype (12). The Foxp3 transgenic mice (referred to as Foxp3-Tg hereafter) also provided a model

system for examining the in vivo consequences of Foxp3 expression. Several individual Foxp3-Tg lines were established, each with differing levels of transgene expression (12, 13). Each of these lines, when bred into an otherwise wild-type background, resulted in a reduction in peripheral CD4+ and CD8+ T cell numbers, with the extent of the reduction directly correlated with transgene copy number and expression (12). However, thymic cellularity was unaffected, as was positive and negative selection. Thus, levels of Foxp3 determined the number of peripheral T cells, while having little effect on the number and differentiation of thymocytes. To examine the function of T cells from Foxp3-Tg mice, a single transgenic line was chosen. This line (2826) had approximately 16 copies of the transgene and had approximately ﬁvefold higher levels of Foxp3 expression (13, 13a). Although T cell development appears to be normal in these mice, peripheral T cell numbers and representation in secondary lymphoid organs are dramatically decreased. Using a variety of in vitro and in vivo assays, the function of T cells from the Foxp3Tg mice was shown to have severely decreased responses when activated through the TCR. For example, puriﬁed CD4+ T cells displayed dramatically reduced proliferative responses when stimulated in vitro (13). Similarly, these T cells produced virtually no IL-2 upon stimulation. In fact, by any measure, CD4+ T cells from Foxp3-Tg mice were functionally inert. Taken together, these data reﬂect a generalized deﬁcit in cellular activation of CD4+ T cells that express Foxp3. The failure of T cells from Foxp3-Tg mice to function in vitro is also reﬂected in the response of these mice to immunologic challenge in vivo. The Foxp3-Tg mice failed to mount an antigen-mediated contact sensitivity response (13). Also, recent data demonstrate that Foxp3-Tg mice have dramatically depressed responses to T-dependent antigens (14). The latter response appears to be due to a reduced ability to produce cytokines and to www.annualreviews.org • FOXP 3

NFAT: nuclear factor of activated T cells

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elaborate CD40L on their cell surface. Thus, overexpression of Foxp3 in CD4+ T cells in vivo results in lowered overall numbers of T cells, with impaired functionality of those T cells that remain.

Foxp3: Necessary and Sufficient for Mouse CD4+ CD25+ Treg Development and Function

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Although it is clear that expression of Foxp3 in CD4+ T cells has deleterious effects on their ability to respond normally to TCR-mediated signals (13, 14), the fact that a population of Foxp3+ T cells exists suggests a role for this protein in CD4+ T cell function. Clues for this function come from an initial analysis of the cells that express Foxp3 in the mouse. Several groups simultaneously reported that the predominant cell type that expressed Foxp3 was CD4+ CD25+ T cells (15–18), the population of CD4+ T cells that can suppress the proliferation and cytokine production of TCR-stimulated conventional CD4+ T cells (19, 20). The role of Foxp3 in these cells has been elucidated in mice through a combination of genetic and direct functional studies. Taken as a whole, these data demonstrate that Foxp3 expression in mouse CD4+ T cells is sufﬁcient to mark these T cells as regulatory. CD25, the IL-2Rα chain, had previously been shown to be the only reliable marker for these cells (19); however, it is also a marker of activated CD4+ T cells. Thus, the identiﬁcation of Foxp3 as a marker for this population of T cells was critical for the further analysis of these cells and their role in peripheral tolerance. In addition to the correlation of Foxp3 expression and CD4+ CD25+ Tregs, the ability of Foxp3 to reprogram CD4+ T cell function has recently been reported (15–17). Several lines of evidence demonstrate a role for Foxp3 in the development and/or function of CD4+ CD25+ Tregs. First, as described earlier, CD4+ CD25+ T cells display Foxp3 expression, whereas other T cell subsets in the mouse do not have detectable expression (15, 212

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16). In fact, using a knockin allele of Foxp3 consisting of an in-frame fusion of green ﬂuorescent protein (GFP) and Foxp3, Fontenot et al. (21) showed that αβTCR+ CD4+ T cells are the predominant cell population that expresses Foxp3. They also found that the relative number of CD25+ and CD25− cells that were Foxp3+ varied according to the source of the T cells. The ability of the Foxp3+ T cells to act as suppressors in vitro was not dependent on expression of CD25. These data were conﬁrmed using a second knockin mouse containing an IRES-RFP (internal ribosome entry site–red ﬂuorescent protein) cassette in the 3 untranslated region of the Foxp3 gene (22). The second line of evidence that Foxp3 expression is necessary and sufﬁcient for mouse Treg development and function comes from studies using ectopic expression of Foxp3 in otherwise conventional T cells. Infection of CD4+ CD25− T cells with a retrovirus expressing Foxp3 converted those cells to a regulatory phenotype, with the infected cells capable of suppressing the proliferation of uninfected CD4+ CD25− T cells (16). The infected cells can also function as Tregs in vivo. In a wide variety of adoptive transfer models of autoimmune disease, cotransfer of CD4+ CD25+ T cells has been shown to protect the host from disease development in these model systems (23–26). Using naive CD4+ CD25− T cells expressing Foxp3 following retroviral gene transfer, Hori et al. (16) showed that cotransfer of cells infected with the Foxp3 retrovirus also protects host mice from autoimmune gastritis. Similarly, CD4+ CD25− T cells expressing Foxp3 also protects against colitis (15). These results have been conﬁrmed by several groups, demonstrating that expression of Foxp3 can convert cells to a Treg-like phenotype. Recent studies have suggested a potential therapeutic use of ectopic Foxp3 expression. BDC2.5 TCR transgenic T cells, transduced with a Foxp3-expressing retrovirus, were capable of ameliorating disease when transferred into nonobese diabetic (NOD) mice with recent

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onset disease (27). However, transfer of polyclonal NOD CD4+ T cells infected with the Foxp3 retrovirus did not protect, suggesting that the frequency of antigen-speciﬁc Tregs is a major factor in the ability of Tregs to control autoimmunity. Similar results have been seen with transfer of puriﬁed CD4+ CD25+ Tregs (25, 28). Complimentary to the experiments using retroviral-mediated delivery of Foxp3 to T cells, Treg function in Foxp3-Tg animals has also been studied. As described above, these mice contain a transgene consisting of cosmid clone containing the mouse Foxp3 gene (12), with approximately 80% of CD4+ T cells in these animals expressing Foxp3 (D.J. Kasprowicz & S.F. Ziegler, unpublished observation). An examination of the function of the CD4+ T cells from these mice showed that the entire population had in vitro regulatory activity (13a, 17). Thus, similar to what was seen with retroviral-mediated introduction of Foxp3, transgenic expression also converted conventional CD4+ T cells to a regulatory phenotype. Interestingly, CD8+ in these mice also displayed in vitro regulatory activity, demonstrating that Foxp3 expression in nonCD4+ T cells was also capable of phenotypic conversion. Just as ectopic Foxp3 expression has been shown to drive Treg function, lack of Foxp3 has been correlated with a lack of Treg cells. For example, mice that lack functional Foxp3, either via the scurfy mutation or a targeted mutation, lack Treg activity (15, 17, 21). Importantly, a conditional deletion of Foxp3 in CD4+ T cells also led to a lymphoproliferative disease indistinguishable from that seen in scurfy males (21). Further evidence for the critical role of Foxp3 in determining the Treg lineage comes from mixed bone marrow chimera experiments. Lethally irradiated mice were reconstituted with a mixture of bone marrow from wild-type and Foxp3− mice, congenically marked to allow the contribution of each to the reconstituted animal. Only bone marrow from the wild-type donor contributed to the CD4+ CD25+ compartment, demonstrat-

ing that Foxp3 is needed in the development of Tregs and that the role of Foxp3 is cell intrinsic (15). Taken as a whole, the data described in this section demonstrate the absolute need for Foxp3 in the development and function of Tregs. They also show that ectopic expression of Foxp3 can cause a phenotypic conversion of T cells to a regulatory phenotype. Thus, in the mouse, Foxp3 is both necessary and sufﬁcient for Treg development and function.

TGF-β: transforming growth factor-β Stat: signal transduction activator of transcription

Regulation of Foxp3 expression. Although obviously important to the understanding of Treg development, the factors that regulate Foxp3 expression remain elusive. The primary issue that has confounded these studies is that the factors that appear to affect Foxp3 expression also affect the survival and expansion of Tregs. However, some signaling pathways, including CD28, IL-2, and TGF-β, are emerging that appear to have an affect on the expression of Foxp3. For example, recent work has shown that CD28-mediated signals during thymic development are required for proper Foxp3 expression (29). However, other studies have implicated CD28 in Treg survival and expansion (30, 31), complicating an interpretation of the role of CD28 in Foxp3 expression. Similarly, mice lacking the IL-2 pathway owing to targeted mutations also display a deﬁcit in CD4+ CD25+ Tregs. This is true for mice with mutations in il-2, il-2rα, and il-2rβ, as well as for mice lacking the downstream mediator of IL-2 signaling, Stat5 (32–40). However, similar to the situation with CD28, distinguishing between a direct affect on transcription and survival is not possible. Recent studies have shown that IL-2 is absolutely required for Treg expansion in the periphery (41, 42). Furthermore, Malek et al. (39, 40) showed that thymus-restricted expression of IL-2Rβ-chain was sufﬁcient to rescue il-2rb−/− mice from fatal autoimmune disease, demonstrating a need for IL-2 signals during thymic development of Tregs. Finally, using the GFP-Foxp3 strain, Fontenot et al. (21) have shown that thymic expression of Foxp3 www.annualreviews.org • FOXP 3

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is absolutely dependent on TCR-MHC interactions and independent of commitment to either the CD4 or CD8 lineage. A more deﬁnitive assessment of the factors that control Foxp3 expression await an analysis of its cis-regulatory region. The availability of the knockin strains described above, as well as a characterization of the Foxp3 cis-regulatory region, will be invaluable in sorting out direct and indirect effects of these and other factors on Foxp3 expression. Perhaps the most controversial aspect of Foxp3 gene regulation is the role of TGF-β. Although TGF-β can have effects on Treg expansion and survival (43, 44), its actual role is not at all clear. Several groups have shown that in vitro culture of CD4+ CD25− T cells with a cocktail that includes TGF-β and IL2, in combination with TCR engagement and costimulation through CD28, can lead to induction of Foxp3 expression and acquisition of repressor activity (45–47). However, there are several issues with these experiments. The cultures used superphysiological concentrations of TGF-β, and investigators have shown that TGF-β can inhibit the proliferative affects of IL-2, at least in conventional T cells (48). Thus, one explanation for the in vitro results is that TGF-β inhibits the proliferation of CD4+ CD25− Foxp3− T cells in the cultures, allowing the expansion of contaminating Foxp3+ cells and an apparent increase in Foxp3 mRNA. Also, the culture systems used are very likely to produce a complex combination of signals in responding cells, making a deﬁnitive role for any of the signaling molecules difﬁcult to discern. In support of an indirect role for TGF-β in Foxp3 expression and function, Tregs from mice either lacking TGF-β1 or expressing a dominant-negative TGF-βRII are indistinguishable from their counterparts from wild-type mice in in vitro suppression assays (49). The data on the in vivo role of TGFβ are equally contradictory. In support of a role for TGF-β in Treg function and Foxp3 expression, several groups have shown that TGF-β is capable of expanding the in vivo

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pool of Tregs (18). For example, T cell– speciﬁc expression of a dominant-active form of TGF-β leads to an elevation in the number of CD4+ CD25+ Tregs and increased Foxp3 expression (50, 51). However, these studies do not differentiate between expansion and de novo generation of Tregs. Using mice with an inducible TGF-β transgene, Peng et al. (44) showed that transient TGF-β expression speciﬁcally in the pancreatic islets was sufﬁcient to protect against onset of diabetes. They found that this transient expression increased the number of CD4+ T cells with a regulatory phenotype in the islets, consistent with a role for TGF-β in the expansion of Tregs in vivo. Studies using mice incapable either of producing TGF-β or of responding to it provide the most contradictory data. These mice have been used to demonstrate that TGF-β is either absolutely required for Treg expansion and function (50, 51) or completely dispensable (49). In addition, recent work has suggested that the effector T cell needs to be TGF-β responsive in order to be suppressed, at least in vivo (52). However, this appears not to be the case in vitro (49; K. Newton & S.F. Ziegler, manuscript in preparation). The recent development of Foxp3 reporter mice should help clarify this issue. In fact, using mice with a reporter for Foxp3 mRNA, Wan & Flavell (22) showed that TGF-β may directly inﬂuence Foxp3 expression.

FUNCTIONAL CHARACTERIZATION OF Foxp3 As described above, Foxp3 is a member of the forkhead/winged-helix family of transcriptional regulators. A common feature of members of this family is the FKH domain (Figure 1), which has been shown to be necessary and sufﬁcient for DNA binding (53, 54). DNA-binding analyses from a number of Fox family proteins have deﬁned a core DNA sequence (5 -A(A/T)TRTT(G/T)R-3 , where R = pyrimidine) surrounded by less conserved sequences (55). Experimentally, two

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Figure 1 Schematic of human FOXP3 and FOXP3 with location of known IPEX mutations. Top schematic is FOXP3, with locations of missense mutations indicated by arrow. Bottom schematic is FOXP3, with location of mutations predicted to affect splicing or mRNA stability (red arrows) and to generated frameshifts (blue arrows) indicated. Exons are color coded according to the protein schematic, with pale blue indicating coding region of unknown function (FKH: forkhead).

canonical FKH-binding sequences [from the transthyretin promoter (TTR-S) and the immunoglobulin variable regions V1 promoter (V1P) (56)] have been used as templates to analyze the DNA-binding properties of Fox proteins. Similar to other members of the Fox family, Foxp3 can bind to each of these FKHbinding sites (57). Thus, as predicted from the sequence analysis, Foxp3 is a DNA-binding protein capable of speciﬁc binding to a canonical FKH DNA-binding site. Members of the Fox family are both transcriptional activators and transcriptional repressors. To assess the role of Foxp3 in transcriptional regulation, a reporter plasmid was constructed that contained three copies of the V1P FKH site linked to a minimal SV40 promoter and the ﬁreﬂy luciferase gene. Cotransfection of this reporter plasmid with a Foxp3 expression plasmid into Cos-7 cells resulted in a dramatic reduction of luciferase activity, suggesting that Foxp3 acts as a transcriptional repressor (57). The ability of Foxp3 to act as a transcriptional repressor required the presence of the FKH domain of Foxp3 and the V1P-binding site, showing that binding of Foxp3 to the FKH site is responsible for the observed effect of Foxp3 on the expression of the reporter. Several lines of evidence suggest that a major target of Foxp3-mediated transcriptional regulation is cytokine genes. For ex-

Other Forkhead Family Proteins and Immune Regulation In addition to Foxp3, several other members of the forkhead family have been implicated in regulating immune system development and function (90). For example, mutations in the Foxn1 gene are responsible for the phenotype seen in nude (nu) mice, rats, and humans (91, 92). These mice are characterized by abnormal development of the epidermis, lack of hair, and absence of a thymus. Mutations in Foxj1 lead to embryonic lethality, but fetal liver chimeras have been used to study its role in the immune system. These mice displayed multisystemic cellular autoimmunity, as well as T cell hyperproliferation and hyperactivity. CD4+ T cells from these animals were skewed toward a Th1 cytokine proﬁle, could be activated solely by IL-2 (CD3 and CD28 stimulation were not required), and exhibited decreased levels of the IκKβ regulatory subunit (93). These results are consistent with a role for Foxj1 in the negative regulation of NF-κB activity. Finally, Foxo3a has been implicated in cellular survival, where it is involved in the regulation of NF-κB (94, 95). Upon Aktdependent phosphorylation, Foxo3a is shuttled to cytoplasm and rendered inactive as a transcriptional regulator. Mice deﬁcient in Foxo3a develop T cell hyperactivity and multiorgan lymphocytic inﬁltrates with age. ample, ectopic expression of Foxp3 in Jurkat cells resulted in a marked reduction of IL2 production following stimulation (57). In addition, CD4+ T cells from mice expressing www.annualreviews.org • FOXP 3

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a Foxp3 transgene were unable to produce IL2, IL-4, or IFN-γ following TCR-mediated stimulation in vitro and had severely reduced ability to express cytokines in vivo following immunization (13, 14). Taken as whole, these data demonstrate a role for Foxp3 in the negative regulation of cytokine gene expression in CD4+ T cells. Inspection of composite NFAT/AP-1 sites upstream of several cytokine genes revealed a possible explanation for the ability of Foxp3 to regulate these genes. Either overlapping or directly adjacent to these DNA-binding sites was a potential FKH-binding site. Provocatively, oligonucleotide probes containing the composite sites from the human IL-2 and GM-CSF genes were capable of competing with the V1P site for Foxp3 binding in a gel shift assay (57). Conﬁrmation of the ability of Foxp3 to repress NFAT-mediated transcription comes from experiments using a reporter plasmid consisting of the –280 composite NFAT/AP-1 site from the mouse IL2 gene. NFAT activation was inhibited when the Foxp3 expression vector was cotransfected (57). Furthermore, work from our laboratory, using a bipartite reporter containing both NFAT and GAL4 DNA-binding sites, has shown that Foxp3 can inhibit NFAT-mediated transcription while binding to a more distant DNA site ( J. Lopes, T. Torgerson, L.A. Schubert, H. Ochs, and S.F. Ziegler, manuscript submitted). These experiments took advantage of fusion proteins consisting of the yeast GAL4 DNA-binding domain fused to the non-FKH sequences from FOXP3, which was found to be capable of inhibiting activation-induced expression of the NFAT reporter. Much effort is currently being used to deﬁne functional domains within FOXP3. As shown in Figure 1, FOXP3 contains at least three distinct structural domains: FKH, Leucine zipper, and C2H2 zinc ﬁnger. As described above, the FKH domain is critical for both DNA binding and for nuclear localization. Conﬁrmation of its role in nuclear localization comes from site-directed mutation of

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two lysine residues (K415 and K416), at the carboxy end of the FKH domain, to glutamic acid. This mutant form of FOXP3, when expressed in cell lines, is localized to the cytoplasm ( J. Lopes, T. Torgerson, L.A. Schubert, H. Ochs, and S.F. Ziegler, manuscript submitted). The functional properties of the remainder of the protein are not well understood. However, studies on other members of the FOXP family may be relevant to the analysis of FOXP3 function. For example, the Leucine zipper domain of Foxp1 and Foxp2 is critical for homo- and heterodimer formation (58, 59). Additionally, deletion of the Leucine zipper domain from Foxp1 and Foxp2 abrogated their ability to act as transcriptional repressors. This domain of FOXP3 is likely also to be involved in dimerization (see below). The amino terminal half of FOXP3 contains no obvious identiﬁable functional domains, although it does contain a fairly high proportion of proline residues (Figure 1). However, we have shown that a fusion protein containing the DNA-binding domain of the yeast transcription factor GAL4 and the amino terminal half of human FOXP3 (amino acids 1–198) was functional as a transcriptional repressor, whereas a fusion of the zinc ﬁnger and Leucine zipper domains was nonfunctional (the GAL4 DNA-binding domains also contain nuclear localization and dimerization sequences). Using infection of primary T cells with FOXP3 deletion mutants, Bettelli et al. (11) showed that there were two functional domains within the amino terminal 200 amino acids of Foxp3, one between amino acids 1 and 150, and one between amino acids 150 and 200. Our lab has also deﬁned two functional domains with the amino terminus of Foxp3. One domain, between amino acids 67 and 132, is involved in general transcriptional repression by FOXP3. The second, between amino acids 135 and 198, is speciﬁcally required for repression of NFAT-mediated transcription. Bettelli et al. (11) also showed that Foxp3 can directly interact with NFAT

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and the NF-κB subunit P65. Although the region of interaction with Foxp3 was not identiﬁed, that data described above would predict that the region between amino acids 135 and 198 is involved.

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HUMAN FOXP3: NOT QUITE THE SAME AS MOUSE The identiﬁcation of Foxp3 as the gene mutated in scurfy mice led to an investigation of its possible role in a human syndrome with clinical features similar to those seen in scurfy animals. This syndrome, known as IPEX (immune dysfunction/polyendocrinopathy/ enteropathy/X-linked), was ﬁrst characterized in 1982 and subsequently mapped to the X chromosome (60, 61). It is characterized by watery, sometimes bloody, diarrhea in very early infancy. In addition, nearly all affected males develop dermatitis (usually eczematous in nature), and most cases develop autoimmune endocrinopathies, with type 1 diabetes and thyroiditis the most common. In general, affected individuals develop symptoms very early in infancy and usually die in the ﬁrst two years of life (61–64). The similarity of the phenotypes of scurfy mice and IPEX humans suggested a common cause for the two diseases. Indeed, mutations in FOXP3 have been identiﬁed in over 20 families with affected offspring (62, 65–69). Many of these are missense mutations in the FKH domain, but mutations have been found across the length of the gene (Figure 1). However, not all patients diagnosed with IPEX have FOXP3 mutations. Sequencing of the FOXP3 gene in a large cohort of individuals diagnosed with IPEX revealed identiﬁable coding region mutations in 60%, with an additional 10% having dramatically reduced mRNA levels (65). One interpretation of these data is that a mutation in FOXP3 leads to development of IPEX, and those IPEX patients without FOXP3 mutations may be mutations in genes whose products interact with FOXP3, either physically or functionally. Indeed, a recent study has found a patient with IPEX-like

symptoms for whom FOXP3 was ruled out as a candidate gene (66). As shown in Figure 1, FOXP3 mutations in IPEX patients have been identiﬁed throughout the coding region. We have begun to analyze the affect of these IPEX mutations on FOXP3 function. Accordingly, mutations in the FKH domain ablate the ability of FOXP3 to inhibit transcription of a reporter construct containing canonical forkhead-binding sites. Consistent with the prevailing view of the FKH domain, mutations in FOXP3 that are predicted to affect DNA binding fail to inhibit transcription of a FKH-dependent reporter ( J. Lopes, T. Torgerson, L. Schubert, H. Ochs, and S.F. Ziegler, manuscript submitted). Interestingly, two IPEX patients were shown to have single codon mutations in the Leucine zipper domain of FOXP3(K250 and E251) (70, 71). We have now found that introducing the E251 mutation into FOXP3 results in a failure to homodimerize and in reduced ability to repress transcription ( J. Lopes, T. Torgerson, L.A. Schubert, H. Ochs, and S.F. Ziegler, manuscript submitted). These data strongly suggest that the Leucine zipper domain of FOXP3 is critical for proper function. Additional mutations have been identiﬁed in the amino terminal domain identiﬁed as critical for FOXP3 function (see previous section); these mutations are now being analyzed to determine their impact on FOXP3 transcriptional repression.

IPEX: immune dysfunction/polyendocrinopathy/ enteropathy/ X-linked

Human FOXP3: Regulation of Expression and Multiple Isoforms A population of CD4+ CD25+ T cells has also been identiﬁed and characterized from human peripheral blood. These cells, when assayed in vitro, are anergic, have the ability to suppress the proliferation and cytokine production of cocultured CD4+ CD25− T cells, and express FOXP3 (18, 72–75). Thus, as has been demonstrated in the mouse, human CD4+ CD25+ T cells express FOXP3 and act as suppressors. www.annualreviews.org • FOXP 3

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However, further examination of FOXP3 expression showed several differences between humans and mice. When FOXP3 expression was monitored by Western blotting, it became apparent that the protein actually ran as a closely spaced doublet (18). RT-PCR analysis has shown that the upper isoform represents the ortholog to mouse Foxp3, whereas the lower isoform is encoded from an mRNA lacking exon 2 (amino acids 71–105) (76). This splicing variant has not been reported in mouse CD4+ CD25+ Tregs. Whether the same T cell expresses both isoforms simultaneously is currently not known, although all sources of FOXP3 in humans have both isoforms. In addition, it is unclear if there is a functional difference between the two isoforms. However, expression of the exon2 isoform in CD4+ CD25− FOXP3− human T cells leads to functional anergy, but not to the same extent as expression of fulllength FOXP3 (76). Human T cells expressing FOXP3exon2 have intermediate proliferative responses to TCR stimulation and make slightly more IL-2 than cells expressing only the full length. Interestingly, the sequences encoded by exon 2 fall within one of the functional domains of FOXP3 deﬁned above. Thus, it is possible that the lower isoform represents a natural dominant-negative form of FOXP3. Our lab has recently shown that FOXP3 and the orphan retinoic acid nuclear receptor RORα interact and that the region of FOXP3 involved in the interaction is encoded by exon 2 ( J. Du & S.F. Ziegler, manuscript in preparation). We are currently characterizing the molecular basis for this interaction. Another clue that the regulation of Foxp3 expression was not conserved between humans and mice comes from an analysis of FOXP3 expression in stimulated human CD4+ CD25− T cells. The starting population of T cells lacked detectable FOXP3 expression, but within 24 h of stimulation antiCD3+ CD28 FOXP3 expression was detected, peaking at 72 h (18). Similar experiments using mouse CD4+ CD25− T cells did not

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demonstrate upregulation of Foxp3 (15, 16). Further analysis with human CD4+ T cells showed that the subset of T cells that upregulated CD25 also expressed FOXP3 (18). Thus, in humans, FOXP3 was behaving like an activation-induced gene in CD4+ cells. These initial ﬁndings have been conﬁrmed by other groups (76–78) and provide the ﬁrst clue that human and mouse Foxp3 are regulated differently. Consistent with the idea that FOXP3 expression in human CD4+ T cells is linked to TCR-mediated stimulation, CD4+ human T cell clones are also FOXP3+ (77–79; J.H. Buckner & S.F. Ziegler, unpublished observations). Mouse T cell clones have not been found to express Foxp3. As described earlier, ectopic expression of Foxp3 in conventional mouse CD4+ T cells converted those cells to Tregs (15, 16). Several groups have now attempted similar experiments with human FOXP3 using either retroviral or Lentiviral transduction of CD4+ CD25− T cells. As is seen in the mouse system, the transduced cells display anergy when stimulated through the TCR and are capable of suppressing responder CD4+ T cells (76, 80, 81). However, the level of suppression is not at the levels seen in the mouse experiments. Also, these experiments are somewhat confounded by the fact that the CD4+ T cells are stimulated prior to infection, which induces the expression of endogenous FOXP3 (18, 76, 77). As described above, stimulation of human CD4+ CD25− T cells leads to FOXP3 expression. An obvious question is what is the functional outcome for the CD4+ T cell that induces FOXP3 expression? Recent work from our group has shown that these cells develop the capability to act as Tregs (18). When the CD25+ cells are isolated following the 10-day culture period, they have the ability to suppress the proliferation of freshly isolated responder T cells. One possible explanation for this phenomenon is that a preexisting pool of CD4+ CD25− FOXP3+ cells preferentially expands in the cultures. However, this does not appear to be the case.

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There is no detectable FOXP3 expression in the starting population of cells. In addition, when the CD4+ CD25− cells, isolated after the 10-day culture period, are restimulated, the same ratio of CD25+ FOXP3+ / CD25− FOXP3− cells results (M.R. Walker, P. Putheti, J.H. Buckner, and S.F. Ziegler, unpublished data). Further support comes from the ﬁnding that naive CD4+ T cells (CD45RA+ ) and CD4+ CD25− T cells from cord blood are also capable of inducing FOXP3 and becoming Tregs, suggesting that antigen experience is not required for this process (82). Taken as a whole, these data support a model in which the ability of a CD4+ T cell to induce FOXP3 expression is not a developmentally determined event. Rather, it is dependent on the conditions present at the time of antigen stimulation of a given T cell. The underlying molecular mechanism that governs this development remains to be determined. Another piece of evidence regarding the ability to generate Tregs in vitro using human CD4+ T cells comes from studies using speciﬁc antigens, rather than antibody crosslinking, as stimulus. For example, Walker et al. (82) showed that coculture of CD4+ CD25− T cells with allo-antigen-presenting cells (APCs) resulted in the development of Tregs speciﬁc for the stimulating allo-APCs. Although these experiments support a model of de novo Treg generation, the actual stimulating antigen is not known. However, they do demonstrate the speciﬁcity of the in vitro– generated Tregs in that they were unable to suppress responses stimulated by third-party APCs. The ﬁnal piece of evidence that induction of FOXP3 expression and de novo Treg generation can occur following antigen stimulation comes from studies using human MHC class II tetramers to isolate antigen-speciﬁc cells following in vitro stimulation. CD4+ CD25− T cells from DR ∗ 0401 (DR4) individuals were stimulated with the immunodominant peptide from inﬂuenza virus, HA(306–319), in the presence of ir-

radiated APCs. After 10 days of culture, tetramer+ CD25+ cells were isolated from the cultures. These cells expressed FOXP3 and inhibited the subsequent proliferation of freshly isolated responder CD4+ T cells from the same donor (82). However, these HA(306–319)+ CD25+ FOXP3+ T cells did not affect the proliferation of T cells when stimulated with an unrelated antigen, tetanus toxoid, unless they also were given the HA peptide. Thus, they required cognate antigen for activation, but once activated they were capable of suppressing responses in an antigennonspeciﬁc manner. The data outlined above suggest that there are at least two distinct populations of Tregs in humans. One population is generated in the thymus, is self-reactive, and is involved in protection from autoimmune responses. These Tregs are referred to as natural Tregs and would be analogous to those seen in the mouse that arise during thymic selection (83). The role of this population of Tregs is to protect against self-reactive effector T cells in the periphery. These cells comprise the CD4+ CD25+ T cell population in human peripheral blood and are equivalent to those Tregs described in the mouse. This population of Tregs is also likely to be most affected by the mutations in FOXP3 seen in IPEX patients. These mutations can be expected to result either in the development of self-reactive T cells that lack suppressor activity or in the complete lack of this population altogether. In either instance, the result would be a failure in the ability to control self-reactive effector T cells. In this respect, the IPEX patients resemble the scurfy mice. A second population of Tregs is represented by CD4+ CD25+ FOXP3+ cells that are generated upon in vitro stimulation of CD4+ CD25− FOXP3− cells. By all criteria measured, these cells are indistinguishable from natural Tregs (18, 82; J.H. Buckner & S.F. Ziegler, unpublished data). As described above, they can be generated following polyclonal or antigen-speciﬁc stimulation. Also, similar to natural Tregs, their ability to www.annualreviews.org • FOXP 3

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suppress is IL-10 and TGF-β independent, but is contact dependent (18). In addition, transcript array analysis has shown that the same set of genes regulated in natural Tregs appears to be similarly regulated in the in vitro–generated cells (P. Putheti & S.F. Ziegler, unpublished data). Taken together, the data demonstrate that CD4+ T cells with the same functional properties as thymically derived Tregs can be generated from naive conventional CD4+ T cells following TCR stimulation. Although the in vivo function of these cells is not known, we propose that they play a role in controlling immune responses, possibly serving to limit bystander activation at the site of the response. In addition, there are reports that CD4+ CD25+ Tregs can induce infectious tolerance through the local generation of classes of regulatory T cells (e.g., Tr1 cells) (84), further extending this paradigm. An unanswered question at this point is the fate of the induced Tregs. Because these cells appear to respond poorly to proliferative signals and are impaired in cytokine production, it seems likely that they will not persist once the response has ended and antigen is no longer available.

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FOXP3 AND THERAPEUTIC INTERVENTION FOR AUTOIMMUNITY As described above, following retroviralmediated introduction of Foxp3, conventional CD4+ T cells acquire a regulatory-like phenotype and are capable of suppressing immune responses both in vitro and in vivo (15–17). The in vivo experiments involved cotransfer of Foxp3-transduced T cells with pathogenic CD4+ T cells in a model of colitis. These studies suggest the possibility of using ectopic Foxp3 expression to convert T cells to take on a regulatory phenotype and then using these cells in adoptive cellular immunotherapy. Coupled with the ability to identify and isolate antigen-speciﬁc human CD4+ T cells using MHC class II tetramers (85–87), ex vivo generation of antigen-speciﬁc 220

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Tregs has great promise. Indeed, some recent reports have suggested that this approach may be viable. For example, Jaeckel et al. (27) used Foxp3-transduced T cells from BDC2.5 TCR transgenic mice to treat NOD mice with recent onset diabetes. They found that the antigen-speciﬁc BDC2.5 T cells, expressing Foxp3, were capable of reversing disease. However, they also showed that transfer of Foxp3-transduced polyclonal T cells from NOD mice failed to rescue. These data suggest that the critical feature for this type of therapy may be getting enough antigenspeciﬁc Tregs to the correct site, and that the transduced polyclonal T cells did not contain sufﬁcient islet-antigen-speciﬁc cells to have a therapeutic effect. In contrast to that study, Loser et al. (88) have shown that Foxp3-transduced polyclonal naive T cells, when transferred to sensitized hosts, could inhibit contact hypersensitivity responses. These polyclonal T cells were also used to treat mice expressing a keratin 14-CD40L transgene. These mice develop chronic skin inﬂammation and systemic autoimmunity (89). Transfer of Foxp3transduced CD4+ T cells into these animals protected against both the local skin inﬂammation and the systemic autoimmunity (88). Transferring these types of therapies to humans presents unique problems. As described above, stimulation of human CD4+ CD25− T cells induces FOXP3 expression, and most methods for introducing foreign genes into T cells using viral vectors require some level of stimulation. Thus, the very cells one may want to transduce and convert to Tregs may be less likely to be infected owing to expression of FOXP3 and a lowered overall activation level. However, the work from Walker et al. (82) suggests that deriving antigen-speciﬁc CD4+ T cells in vitro with self-antigens, followed by isolation of CD25+ that are also MHC class II tetramer+ , may be a solution to this problem. The obvious limitation to this approach is the availability of the appropriate MHC class II tetramer for a given HLA haplotype and speciﬁc epitope.

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In summary, the reemergence of suppressor T cells and the discovery of FOXP3 as a critical player in their development and function hold great promise for the treatment of autoimmune diseases in humans. The differences between humans and mice may present

obstacles in the transfer from mouse models to actual human disease. Thus, a more concerted effort to develop better human in vitro and in vivo systems is required to understand better the potential therapeutic beneﬁts of manipulating FOXP3.

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ACKNOWLEDGMENTS The author thanks Drs. Megan K. Levings (University of British Columbia) and Troy R. Torgerson (University of Washington) for sharing information and data prior to publication, and Drs. Jane H. Buckner, Daniel J. Campbell, and Gerald T. Nepom (Benaroya Research Institute) for critical reading of the manuscript prior to submission. The author is supported by grants from the NIH (AI48779, AI059926, DK068312), American Diabetes Association, Juvenile Diabetes Research Foundation, and DANA Foundation.

LITERATURE CITED 1. Baecher-Allan C, Viglietta V, Haﬂer DA. 2004. Human CD4+ CD25+ regulatory T cells. Semin. Immunol. 16:89–98 2. Piccirillo CA, Thornton AM. 2004. Cornerstone of peripheral tolerance: naturally occurring CD4+ CD25+ regulatory T cells. Trends Immunol. 25:374–80 3. Shevach EM. 2002. CD4+ CD25+ suppressor T cells: more questions than answers. Nat. Rev. Immunol. 2:389–400 4. Sakaguchi S. 2005. Naturally arising Foxp3-expressing CD25+ CD4+ regulatory T cells in immunological tolerance to self and non-self. Nat. Immunol. 6:345–52 5. Fontenot JD, Rudensky AY. 2005. A well adapted regulatory contrivance: regulatory T cell development and the forkhead family transcription factor Foxp3. Nat. Immunol. 6:331–37 6. Godfrey V, Rouse BT, Wilkinson JE. 1994. Transplantation of T cell-mediated lymphoreticular disease from the scurfy (sf ) mouse. Am. J. Pathol. 145:281–86 7. Waterhouse P, Penninger JM, Timms E, Wakeham A, Shahinian A, et al. 1995. Lymphoproliferative disorders with early lethality in mice deﬁcient in Ctla-4. Science 270:985–88 8. Tivol EA, Borriello F, Schweitzer AN, Lynch WP, Bluestone JA, Sharpe AH. 1995. Loss of CTLA-4 leads to massive lymphoproliferation and fatal multiorgan tissue destruction, revealing a critical negative regulatory role of CTLA-4. Immunity 3:541–47 9. Kanangat S, Blair P, Reddy R, Deheshia M, Godfrey V, et al. 1996. Disease in the scurfy (sf ) mouse is associated with overexpression of cytokine genes. Eur. J. Immunol. 26:161– 65 10. Clark LB, Appleby MW, Brunkow ME, Wilkinson JE, Ziegler SF, Ramsdell F. 1999. Cellular and molecular characterization of the scurfy mouse mutant. J. Immunol. 162:2546–54 11. Bettelli E, Dastrange M, Oukka M. 2005. Foxp3 interacts with nuclear factor of activated T cells and NF-κB to repress cytokine gene expression and effector functions of T helper cells. Proc. Natl. Acad. Sci. USA 102:5138–43 12. Brunkow ME, Jeffery EW, Hjerrild KA, Paeper B, Clark LB, et al. 2001. Disruption of a new forkhead/winged-helix protein, scurfin, results in the fatal lymphoproliferative disorder of the scurfy mouse. Nat. Genet. 27:68–73 13. Khattri R, Kasprowicz DJ, Cox T, Yasayko S-A, Ziegler SF, Ramsdell F. 2001. The www.annualreviews.org • FOXP 3

12. This paper describes the identification of Foxp3 as the gene mutated in scurfy mice and describes the transgenic animals used to study Foxp3’s role in CD4 T cell function.

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14. 15, 16, and 17. These seminal papers demonstrate conclusively Foxp3’s critical role in Treg development and function. 18. This was the first paper to show that, in humans, FOXP3 expression can be induced by TCR stimulation, that cells with regulatory capability can be generated de novo, and that regulation of human and mouse Foxp3 have key differences. 19. This paper “rediscovered” Tregs as a distinct population of cells. Much subsequent work on Tregs and control of peripheral tolerance derives from the work described here. 21 and 22. These two papers describe the Foxp3 reporter mice that are vital to the study of Tregs as, in the mouse, Foxp3 is the only marker that appears to be exclusive to these cells.

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22. 23.

24. 25.

26.

27. 28. 29.

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amount of scurﬁn protein determines peripheral T cell number and responsiveness. J. Immunol. 167:6312–20 Kasprowicz DJ, Droin N, Ramsdell F, Green DR, Ziegler SF. 2006. Elevated FoxP3 expression leads to activation-induced cell death, while effector cell differentiation correlates with loss of FoxP3 expression. Eur J. Immunol. In press Kasprowicz DJ, Smallwood PS, Tyznik AJ, Ziegler SF. 2003. Scurﬁn (FoxP3) controls T-dependent immune responses in vivo through regulation of CD4+ T cell effector function. J. Immunol. 171:1216–23 Fontenot JD, Gavin MA, Rudensky AY. 2003. FoxP3 programs the development and function of CD4+ CD25+ regulatory T cells. Nat. Immunol. 4:330–36 Hori S, Nomura T, Sakaguchi S. 2003. Control of regulatory T cell development by the transcription factor FoxP3. Science 299:1057–61 Khattri R, Cox T, Yasayko S-A, Ramsdell F. 2003. An essential role for Scurfin in CD4+ CD25+ T regulatory cells. Nat. Immunol. 4:337–42 Walker MR, Kasprowicz DJ, Gersuk VH, Benard A, Van Landeghen M, et al. 2003. Induction of FoxP3 and acquisition of T regulatory activity by stimulated human CD4+ CD25− T cells. J. Clin. Invest. 112:1437–43 Sakaguchi S, Sakaguchi N, Asano M, Itoh M, Toda M. 1995. Immunologic selftolerance maintained by activated T cells expressing IL-2 receptor alpha-chains (CD25). Breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J. Immunol. 155:1151–64 Suri-Payer E, Amar AZ, Thornton AM, Shevach EM. 1998. CD4+ CD25+ T cells inhibit both the induction and effector function of autoreactive T cells and represent a unique lineage of immunoregulatory cells. J. Immunol. 160:1212–18 Fontenot JD, Rasmussen JP, Williams LM, Dooley JL, Farr AG, Rudensky AY. 2005. Regulatory T cell lineage specification by the forkhead transcription factor foxp3. Immunity 22:329–41 Wan YY, Flavell RA. 2005. Identifying Foxp3-expressing suppressor T cells with a bicistronic reporter. Proc. Natl. Acad. Sci. USA 102:5126–31 Kohm AP, Carpentier PA, Anger HA, Miller SD. 2002. Cutting edge: CD4+ CD25+ regulatory T cells suppress antigen-speciﬁc autoreactive immune responses and central nervous system inﬂammation during active experimental autoimmune encephalomyelitis. J. Immunol. 169:4712–16 Wraith DC, Nicolson KS, Whitely NT. 2004. Regulatory CD4+ T cells and the control of autoimmune disease. Curr. Opin. Immunol. 16:695–701 Tang Q, Henriksen KJ, Bi M, Finger EB, Szot G, et al. 2004. In vitro-expanded antigen-speciﬁc regulatory T cells suppress autoimmune diabetes. J. Exp. Med. 199:1455– 65 Chai JG, Xue SA, Coe D, Addey C, Bartok I, et al. 2005. Regulatory T cells, derived from naive CD4+ CD25− T cells by in vitro Foxp3 gene transfer, can induce transplantation tolerance. Transplantation 79:1310–16 Jaeckel E, von Boehmer H, Manns MP. 2005. Antigen-speciﬁc FoxP3-transduced T-cells can control established type 1 diabetes. Diabetes 54:306–10 Lundsgaard D, Holm TL, Hornum L, Markholst H. 2005. In vivo control of diabetogenic T-cells by regulatory CD4+ CD25+ T-cells expressing Foxp3. Diabetes 54:1040–47 Tai X, Cowan M, Feigenbaum L, Singer A. 2005. CD28 costimulation of developing thymocytes induces Foxp3 expression and regulatory T cell differentiation independently of interleukin 2. Nat. Immunol. 6:152–62

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30. Tang Q, Henriksen KJ, Boden EK, Tooley AJ, Ye J, et al. 2003. Cutting edge: CD28 controls peripheral homeostasis of CD4+ CD28+ regulatory T cells. J. Immunol. 171:3348–52 31. Boden E, Tang Q, Bour-Jordan H, Bluestone JA. 2003. The role of CD28 and CTLA4 in the function and homeostasis of CD4+ CD25+ regulatory T cells. Novartis Found. Symp. 252:55–63 ¨ ¨ 32. Scheffold A, Huhn J, Hofer T. 2005. Regulation of CD4+ CD25+ regulatory T cell activity: it takes (IL-)two to tango. Eur. J. Immunol. 35:1336–41 33. Nelson BH. 2004. IL-2, regulatory T cells, and tolerance. J. Immunol. 172:3983–88 34. Malek TR, Bayer AL. 2004. Tolerance, no immunity, critically depends on IL-2. Nat. Rev. Immunol. 4:665–74 35. Nelson BH. 2002. Interleukin-2 signaling and the maintenance of self-tolerance. Curr. Dir. Autoimmun. 5:92–112 36. Schimpl A, Berberich I, Kneitz B, Kramer S, Santner-Nanan B, et al. 2002. IL-2 and autoimmune disease. Cytokine Growth Factor Rev. 13:369–78 37. Snow JW, Abraham N, Ma MC, Herndier BG, Pastuszak AW, Goldsmith MA. 2003. Loss of tolerance and autoimmunity affecting multiple organs in STAT5A/5B-deﬁcient mice. J. Immunol. 171:5042–50 38. Burchill MA, Goetz CA, Prlic M, O’Neil JJ, Harmon IR, et al. 2003. Distinct effects of STAT5 activation on CD4+ and CD8+ T cell homeostasis: development of CD4+ CD25+ regulatory T cells versus CD8+ memory T cells. J. Immunol. 171:5853–64 39. Malek TR, Porter BO, Codias EK, Scibelli P, Yu A. 2000. Normal lymphoid homeostasis and lack of lethal autoimmunity in mice containing mature T cells with severely impaired IL-2 receptors. J. Immunol. 164:2905–14 40. Malek TR, Yu A, Vincek V, Scibelli P, Kong L. 2002. CD4 regulatory T cells prevent lethal autoimmunity in IL-2Rβ-deﬁcient mice: implications for the nonredundant function of IL-2. Immunity 17:167–78 41. Bayer AL, Yu A, Adeegbe D, Malek TR. 2005. Essential role for interleukin-2 for CD4+ CD25+ T regulatory cell development during the neonatal period. J. Exp. Med. 201:769–77 42. Setoguchi R, Hori S, Takahashi T, Sakaguchi S. 2005. Homeostatic maintenance of natural Foxp3+ CD25+ CD4+ regulatory T cells by interleukin (IL)-2 and induction of autoimmune disease by IL-2 neutralization. J. Exp. Med. 201:723–35 43. Marie JC, Letterio JJ, Gavin M, Rudensky AY. 2005. TGF-β1 maintains suppressor function and Foxp3 expression in CD4+ CD25+ regulatory T cells. J. Exp. Med. 201:1061– 67 44. Peng Y, Laouar Y, Li MO, Green EA, Flavell RA. 2004. TGF-β regulates in vivo expansion of Foxp3-expressing CD4+ CD25+ regulatory T cells responsible for protection against diabetes. Proc. Natl. Acad. Sci. USA 101:4572–77 45. Chen W, Jin W, Hardegen N, Lei KJ, Li L, et al. 2003. Conversion of peripheral CD4+ CD25− T cells to CD4+ CD25+ regulatory T cells by TGF-β induction of transcription factor FoxP3. J. Exp. Med. 198:1875–86 46. Fantini MC, Becker C, Monteleone G, Pallone F, Galle PR, Neurath MF. 2004. Cutting edge: TGF-β induces a regulatory phenotype in CD4+ CD25− T cells through FoxP3 induction and down-regulation of Smad7. J. Immunol. 172:5149–53 47. Fu S, Zhang N, Yopp AC, Chen D, Mao M, et al. 2004. TGF-β induces Foxp3 + Tregulatory cells from CD4+ CD25− precursors. Am. J. Transplant. 4:1614–27 48. Nelson BH, Martyak TP, Thompson LJ, Moon JJ, Wang T. 2003. Uncoupling of www.annualreviews.org • FOXP 3

223

ANRV270-IY24-07

ARI

13 February 2006

49.

50.

51.

Annu. Rev. Immunol. 2006.24:209-226. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

52.

53. 54. 55. 56. 57.

58.

59.

60. 61.

62.

63.

64.

65. 66. 224

Ziegler

13:5

promitogenic and antiapoptotic functions of IL-2 by Smad-dependent TGF-β signaling. J. Immunol. 170:5563–70 Piccirillo CA, Letterio JJ, Thornton AM, McHugh RS, Mamura M, et al. 2002. CD4+ CD25+ regulatory T cells can mediate suppressor function in the absence of transforming growth factor β1 production and responsiveness. J. Exp. Med. 196:237–46 Schramm C, Huber S, Protschka M, Czochra P, Burg J, et al. 2004. TGFβ regulates the CD4+ CD25+ T-cell pool and the expression of Foxp3 in vivo. Int. Immunol. 16:1241– 49 Huber S, Schramm C, Lehr HA, Mann A, Schmitt S, et al. 2004. Cutting edge: TGFβ signaling is required for the in vivo expansion and immunosuppressive capacity of regulatory CD4+ CD25+ T cells. J. Immunol. 173:6526–31 Fahlen L, Read S, Gorelik L, Hurst SD, Coffman R, et al. 2005. T cells that cannot respond to TGF-β escape control by CD4+ CD25+ regulatory T cells. J. Exp. Med. 201:737–46 ¨ Kaestner K, Knochel W, Martinez D. 2000. Uniﬁed nomenclature for the wingedhelix/forkhead transcription factors. Genes Dev. 14:142–46 ¨ Kaufmann E, Knochel W. 1996. Five years on the wings of fork head. Mech. Dev. 57:3–20 ¨ ¨ Kaufmann E, Muller D, Knochel W. 1995. DNA recognition site analysis of Xenopus winged helix proteins. J. Mol. Biol. 248:239–54 Li C, Tucker PW. 1993. DNA-binding properties and secondary structural model of the hepatocyte nuclear factor 3/fork head domain. Proc. Natl. Acad. Sci. USA 90:11583–87 Schubert LA, Jeffery EW, Zhang Y, Ramsdell F, Ziegler SF. 2001. Scurﬁn (FOXP3) acts as a repressor of transcription and regulates T cell activation. J. Biol. Chem. 276:37672– 79 Li S, Weidenfeld J, Morrisey EE. 2004. Transcriptional and DNA binding activity of the Foxp1/2/4 family is modulated by heterotypic and homotypic protein interactions. Mol. Cell. Biol. 24:809–22 Wang B, Lin D, Li C, Tucker PW. 2003. Multiple domains deﬁne the expression and regulatory properties of Foxp1 forkhead transcriptional repressors. J. Biol. Chem. 278:24259– 68 Powell BR, Buist N, Stenzel P. 1982. An X-linked syndrome of diarrhea, polyendocrinopathy, and fatal infection in infancy. J. Pediatr. 100:731–37 Bennett CL, Ochs HD. 2001. IPEX is a unique X-linked syndrome characterized by immune dysfunction, polyendocrinopathy, enteropathy, and a variety of autoimmune phenomena. Curr. Opin. Pediatr. 13:533–38 Gambineri E, Torgerson TR, Ochs HD. 2003. Immune dysregulation, polyendocrinopathy, enteropathy, and X-linked inheritance (IPEX), a syndrome of systemic autoimmunity caused by mutations of FOXP3, a critical regulator of T cell homeostasis. Curr. Opin. Rheumatol. 15:430–35 Torgerson TR, Ochs HD. 2002. Immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: a model of immune dysregulation. Curr. Opin. Allergy Clin. Immunol. 2:481–87 Nieves DS, Phipps RP, Pollock SJ, Ochs HD, Zhu Q, et al. 2004. Dermatologic and immunologic ﬁndings in the immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome. Arch. Dermatol. 140:466–72 Ochs HD, Ziegler SF, Torgerson TR. 2005. FOXP3 acts as a rheostat of the immune response. Immunol. Rev. 203:156–64 Owen CJ, Jennings CE, Imrie H, Lachaux A, Bridges NA, et al. 2003. Mutational analysis

ANRV270-IY24-07

67.

68.

69.

Annu. Rev. Immunol. 2006.24:209-226. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

70.

71.

72.

73.

74. 75.

76.

77.

78.

79. 80.

81.

82.

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of the FOXP3 gene and evidence for genetic heterogeneity in the immunodysregulation, polyendocrinopathy, enteropathy syndrome. J. Clin. Endocrinol. Metab. 88:6034–39 Kobayashi I, Shiari R, Yamada M, Kawamura N, Okano M, et al. 2001. Novel mutations of FOXP3 in two Japanese patients with immune dysregulation, polyendocrinopathy, enteropathy, X linked syndrome (IPEX). J. Med. Genet. 38:874–76 Wildin RS, Ramsdell F, Peake J, Faravelli F, Casanova J-L, et al. 2001. X-linked neonatal diabetes mellitus, enteropathy and endocrinopathy syndrome is the human equivalent of mouse scurfy. Nat. Genet. 27:18–20 Bennett CL, Christie J, Ramsdell F, Brunkow ME, Ferguson PF, et al. 2001. The immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX) is caused by mutation of FOXP3. Nat. Genet. 27:20–21 Chatila TA, Blaeser F, Ho N, Lederman HM, Voulgaropoulos C, et al. 2000. JM2, encoding a fork head-related protein, is mutated in X-linked autoimmunity-allergic disregulation syndrome. J. Clin. Invest. 106:R75–81 Wilden RS, Smyk-Pearson S, Filipovich AH. 2002. Clinical and molecular features of the immunodysregulation, polyendocrinopathy, enteropathy X linked (IPEX) syndrome. J. Med. Genet. 39:537–45 Jonuleit H, Schmitt E, Stassen M, Tuttenberg A, Knop J, Enk AH. 2001. Identiﬁcation and functional characterization of human CD4+ CD25+ T cells with regulatory properties isolated from peripheral blood. J. Exp. Med. 193:1285–94 Dieckmann D, Plottner H, Berchtold S, Berger T, Schuler G. 2001. Ex vivo isolation and characterization of CD4+ CD25+ T cells with regulatory properties from human blood. J. Exp. Med. 193:1303–10 Baecher-Allan C, Brown JA, Freeman GJ, Haﬂer DA. 2001. CD4+ CD25high regulatory cells in human peripheral blood. J. Immunol. 167:1245–53 Levings MK, Sangregorio R, Roncarolo MG. 2001. Human CD25+ CD4+ T regulatory cells suppress naive and memory T cell proliferation and can be expanded in vitro without loss of function. J. Exp. Med. 193:1295–302 Allan SE, Passerini L, Bacchetta R, Crellin N, Da M, et al. 2006. The role of FOXP3, and an isoform lacking exon 2, in the generation of human CD4+ T regulatory cells. J. Clin. Invest. In press Morgan ME, van Bilsen JH, Bakker AM, Heemskerk B, Schilham MW, et al. 2005. Expression of FOXP3 mRNA is not conﬁned to CD4+ CD25+ T regulatory cells in humans. Hum. Immunol. 66:13–20 Roncador G, Brown PJ, Maestre L, Hue S, Martinex-Torrecuadrada JL, et al. 2005. Analysis of FOXP3 protein expression in human CD4+ CD25+ regulatory T cells at the single-cell level. Eur. J. Immunol. 18:1681–91 Malone R, Kochik SA, Reijonen H, Carson BD, Ziegler SF, et al. 2006. Functional avidity directs T-cell fate in autoreactive CD4+ T-cells. Blood. In press Yagi H, Nomura T, Nakamura K, Yamazaki S, Kitawaki T, et al. 2004. Crucial role of FOXP3 in the development and function of human CD25+ CD4+ regulatory T cells. Int. Immunol. 16:1643–56 Oswald-Richter K, Grill SM, Shariat N, Leelawong M, Sundrud MS, et al. 2004. HIV infection of naturally occurring and genetically reprogrammed human regulatory T cells. PLoS Biol. 2:955–66 Walker MR, Carson BD, Nepom GT, Ziegler SF, Buckner JH. 2005. De novo generation of antigen-speciﬁc CD4+ CD25+ regulatory T cells from human CD4+ . Proc. Natl. Acad. Sci. USA 102:4103–8 www.annualreviews.org • FOXP 3

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83. Bluestone JA, Abbas AK. 2003. Natural versus adapted regulatory T cells. Nat. Rev. Immunol. 3:253–57 84. Moore KJ, de Waal MR, Coffman R, O’Garra A. 2001. Interleukin-10 and the interleukin10 receptor. Annu. Rev. Immunol. 19:683–765 85. Novak EJ, Masewics SA, Liu AW, Lernmark A, Kwok WW, Nepom GT. 2001. Activated human epitope-speciﬁc T cells identiﬁed by class II tetramers reside within a CD4high , proliferating subset. Int. Immunol. 13:799–806 86. Buckner JH, Holzer U, Novak EJ, Reijonen H, Kwok WW, Nepom GT. 2002. Deﬁning antigen-speciﬁc responses with human MHC class II tetramers. J. Allergy. Clin. Immunol. 110:199–208 87. Nepom GT, Buckner JH, Novak EJ, Reichstetter S, Reijonen H, et al. 2002. HLA class II tetramers. Tools for direct analysis of antigen-speciﬁc CD4+ T cells. Arthritis Rheum. 46:5–12 88. Loser K, Hansen W, Apelt J, Balkow S, Buer J, Beissert S. 2005. In vitro-generated regulatory T cells induced by FoxP3-retrovirus infection control murine contact allergy and systemic auroimmunity. Gene Ther. In press 89. Mehling A, Loser K, Varga G, Metze D, Luger TA, et al. 2001. Overexpression of CD40 ligand in murine epidermis results in chronic skin inﬂammation and systemic autoimmunity. J. Exp. Med. 194:615–28 90. Cofffer PJ, Burgering BM. 2004. Forkhead box transcription factors and their role in the immune system. Nat. Rev. Immunol. 4:889–99 91. Pignata C, Gaetaniello L, Masci AM, Frank J, Christiano A, et al. 2001. Human equivalent of the mouse Nude/SCID phenotype: long-term evaluation of immunologic reconstitution after bone marrow transplantation. Blood 97:880–85 92. Nehls M, Pfeifer D, Schorpp M, Hedrich H, Boehm T. 1994. New member of the winged-helix protein family disrupted in mouse and rat nude mutations. Nature 372:103– 7 93. Lin L, Spoor MS, Gerth AJ, Body SL, Peng SL. 2004. Modulation of Th1 activation and inﬂammation by the NF-κB repressor Foxj1. Science 303:1017–20 94. Lin L, Hron JD, Peng SL. 2004. Regulation of NF-κB, Th activation, and autoinﬂammation by the forkhead transcription factor Foxo3a. Immunity 21:203–13 95. Burgering BM, Kops GJ. 2002. Cell cycle and death control: long live Forkheads. Trends Biochem. Sci. 27:352–60

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:209-226. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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HIV Vaccines Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Andrew J. McMichael MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford OX3 9DS UK; email: [email protected]

Annu. Rev. Immunol. 2006. 24:227–55 First published online as a Review in Advance on December 5, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090605 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0227$20.00

Key Words HIV-1, neutralizing antibody, T cell immunity, vaccination

Abstract A prophylactic vaccine for HIV-1 is badly needed. Despite 20 years of effort, it is still a long way off. However, considerable progress has been made in understanding the problem. The virus envelope has evolved to evade neutralizing antibodies in an extraordinary way, yet a vaccine that can stimulate such antibodies remains the best hope. Anti-HIV-1 T cell responses are evaded by continuous mutation of the virus. Vaccine strategies that concentrate on stimulating T cell immunity will at best generate broadly reactive and persisting T cell responses that can suppress virus without preventing infection, limiting or preventing the damage the virus causes. The SIV macaque models give encouragement that this is possible, but they need further understanding. Therapeutic vaccination should also be considered.

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INTRODUCTION SIV: simian immunodeﬁciency virus

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GALT: gut-associated lymphoid tissue

The human immunodeﬁciency virus type 1 (HIV-1) pandemic shows no sign of slowing. WHO/UNAIDS estimates that 39.4 million people were living with HIV-1 infection at the end of 2004, with 5 million new infections each year. The introduction of potent antiretroviral drugs has lessened the impact of AIDS in developed countries, but their use in developing countries is patchy and is unlikely to slow the pandemic. Although the rates of infection appear to have fallen in Uganda and Senegal, possibly because of vigorous education campaigns, changes in social behavior, and increased condom use, the infection rates are still high in these countries. There is therefore a desperate need for a vaccine. Yet despite increasingly intense efforts, development of a vaccine is still a long way off. This review explores this difﬁcult problem, arguing that progress is being made and that investigators are still hopeful that a vaccine is possible.

THE NATURAL HISTORY OF HIV INFECTION The commonest route of transmission of HIV-1 is through heterosexual intercourse. Estimates of the rate of infection in very high risk sex worker cohorts show that ∼1 in 100 sexual contacts with an infected person results in infection (1–3); notably, 25% of adults in sub-Saharan Africa are in regular sexual contact with an infected partner in a stable monogamous relationship, and 5% become infected per year (4, 5). Studies of very early mucosal infection of rhesus macaques with simian immunodeﬁciency virus (SIV) have detailed what happens in early infection (6, 7). Despite deliberate insertion of virus at a high titer, infected cells cannot be detected in the mucosa for 1–3 days. The ﬁrst infected cells that become detectable are resting memory CD4 T cells that express CCR5, the virus coreceptor. Over the following 3 days, the number of infected cells increases. 228

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At ∼1 week, virus becomes detectable in local lymph nodes; Langerhans cells or dendritic cells (DCs) are involved in carrying virus to these sites. HIV can infect plasmacytoid DCs (pDCs) in peripheral tissue by binding to CD4 and mannose-binding proteins; after internalization of virus, expression of CCR7 is induced, which directs pDCs to lymph nodes (8, 9). HIV can also attach to myeloid DCs by binding to DC-SIGN, and this can target virus to HIV-speciﬁc CD4 T cells without infecting the DC (10, 11). Virus production may thus be ampliﬁed in draining lymph nodes when T cells come in contact with DC and are simultaneously activated and infected (12); HIV-speciﬁc CD4 T cells are particularly susceptible (13). By day 7, virus begins hematogenous spread and is seeded to other sites, particularly to gut-associated lymphoid tissue (GALT) where large numbers of CD4+ CCR5+ memory T cells reside. This seeding to other sites results in an exponential expansion of the infection, with up to 20% of gut-associated CD4 T cells infected and ∼80% destroyed, probably by Fas-mediated bystander killing (14). Virus appears in the blood, with viremia peaking at around day 21, often >107 virus particles per ml of plasma. Then the level of HIV-1 starts to fall, either because of saturation of the main target cell population (memory CD4+ CCR5+ T cells) (15) or because of the appearance of speciﬁc immune responses, or both (16, 17). Without therapeutic intervention, virus levels fall 10- to 100-fold and level out to a relatively stable set point between 2 and 6 months after infection. The virus set point is inversely related to the rate of progression to AIDS (18). After this acute phase, virus levels generally stay around the set point for many months. Blood CD4 T cell levels, which dip in acute infection, usually recover as set point is reached, but then gradually decline. However, this gradual decline masks what is really happening in the whole CD4+ T cell population. As blood CD4 count falls below 200– 400, virus levels rise as AIDS progresses.

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Virus load

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CD8 T cell control Expansion in lymph node blood GALT

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Early phase: infection of mucosal memory T cells CD4+/CCR5+ LC/DC 5

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Abortive infection

Damage limitation

Figure 1 The natural history of HIV-1 infection and options for control by a vaccine-induced immune response. In the upper part of the ﬁgure, the change in virus load is shown over time as the virus spreads. Below is shown the time of action of different types of immune response and their effect on the virus. Abbreviations: LC, Langerhans cell; DC, dendritic cell; GALT, gut-associated lymphoid tissue; NK, natural killer cells; Neut Ab, neutralizing antibody.

IMMUNE RESPONSES DURING HIV-1 INFECTION These events, summarized in Figure 1, should be viewed in the context of the concomitant immune responses. In very early infection, innate immunity may impede virus spread, although that it does so is uncertain. It is clear that activated DC and NK cells can be found as the viremia becomes apparent (9, 19– 21); plasma IFN-α levels may rise (9). How these early events inﬂuence the early adaptive immune response and contribute to early control of virus needs further study. The ﬁrst adaptive immune responses that appear are the CD8 T cell response and a humoral antibody response that is non-

neutralizing (17). The HIV-speciﬁc CD8 T cell response appears and expands to ∼10% of all circulating CD8+ T cells as the viremia peaks, and this T cell response is probably responsible for much of the reduction in virus load (16, 22). In support of this hypothesis, depletion of CD8+ cells by anti-CD8 antibody infusion during primary SIV infection in macaques results in sustained high viremia that is only controlled when the antibody infusion ceases (23). However, a subpopulation of NK cells expresses CD8. Additional evidence that CD8+ T cells are responsible for virus control comes from cases in macaques and humans in which virus populations with escape mutations in immunodominant epitopes www.annualreviews.org • HIV Vaccines

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recognized by CD8+ T cells are selected during primary HIV infection (24–27). This ﬁnding implies strong selective pressure on the virus as the peak viremia is controlled. There is a CD4+ T cell response at the same time as the CD8+ T cell response (28, 29). As in other acute virus infections, the CD4+ T cell response is smaller than the CD8+ response (28, 30). The CD4+ T cell response has not been well studied in acute HIV-1 infection, but it may well be compromised by the susceptibility of CD4+ T cells to HIV-1 infection (13). Nonetheless, the CD8+ T cell response appears normal (31). Studies in mice have indicated that the quality of the CD8+ T cell response is inﬂuenced by T cell help. When CD4+ T cells were depleted by infusion of anti-CD4 antibody or were absent in CD4-null mice, CD8 T cell memory development was impaired (32–34); in the most extreme case, the acutely responding T cells apoptosed, and the clones were deleted (35). However, these studies involved complete deletion of CD4 T cells, including T regulatory cells, which is rather different from the partial impairment of CD4+ T cells in HIV-1 infection. Once virus set point is reached, the CD8+ T cell response continues to suppress virus. This suppression has been shown in macaques by using anti-CD8 antibody infusion to deplete CD8 T cells, which caused a rise in virus load for as long as the antibody-mediated CD8 T cell depletion was present (23, 36). The continuing selection of cytotoxic T lymphocyte escape mutants implies continuous selective pressure (37–40). The T cell response evolves over time, often with different patterns of immunodominance during the chronic phase compared with acute infection (41), which may in part reﬂect the selection of virus escape mutants and consequent new T cell responses. The phenotypic features of HIV-speciﬁc T cells, such as CD27, CD28, CCR7, CD57, and CD38 expression, differ between acute and chronic infection and differ from the phenotypes of CD8 T cells responding to other

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chronically infecting viruses (42). The differentiation state of HIV-speciﬁc T cells may be abnormally skewed (43) or may simply reﬂect subtle differences in antigen stimulation by different cell populations in different virus infections. Functional assays have also revealed heterogeneity in expression of perforin, granzymes A and B, IFN-γ, IL-2, MIP-1α, MIP-1β, and RANTES (43, 44). Expression of surface glycoproteins and secreted functional mediators can differ within single clones. It is not clear how these phenotypes and functions relate to the ability of CD8 T cells to suppress HIV-1 and whether impaired CD4 T cell help adversely affects the real function of CD8 T cells. Expression of IL-2 does have some role in slowing progression of HIV infection in both CD4+ and CD8+ T cells; antigen stimulation of IL-2 secretion correlates with slow progression to AIDS (45). Two to three months after primary infection, neutralizing antibody appears (17). However, the antibody fails to neutralize contemporary virus variants (46) and is always chasing the evolving virus so that it probably contributes little to control of the infection (47, 48). There must, however, be a strong enough antiviral force to select the escape variants, but the changes in envelope epitopes seem to have little cost to the virus, possibly because many of the epitopes are in ﬂexible parts of the envelope such as hypervariable loops of gp120 that have no other function. This contrasts with CD8 T cells, for which many escape variants come with a cost to the virus ﬁtness, so that virus escape may actually select less-virulent virus (49, 50).

WHAT COULD VACCINES DO? The problem with the natural immune response to HIV is that it is always chasing the persisting, variable virus. In these circumstances, the virus will always escape. Could this be prevented by ensuring that the immune response is there before the virus infects?

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Vaccination has been a major cornerstone of disease prevention and control since 1798. However, vaccinologists often argue that for every successful vaccine, researchers knew that people who recovered from natural infection were subsequently protected against infection with the same pathogen. As no one has ever been known to clear HIV-1 infection, protection against this infection by an immune response may not be possible. Nonetheless, inability of those infected to clear the virus naturally is not unique to HIV1. For several persisting viruses, including Epstein Barr virus (EBV) and cytomegalovirus, natural clearance and subsequent protective immunity is unknown. This is also true of tuberculosis. However, in these infections lifelong immune control is regularly achieved, with no illness. Such a status for HIV-1 may be possible, particularly as it occurs naturally for HIV-2 in ∼80% of those infected (51). The best vaccine-induced immune response would be neutralizing antibody. If present at high enough titer in blood and mucosa at the time of ﬁrst infection, true sterilizing protection could be achieved (Figure 1). However, even this might fail if some part of the infection was caused by cell-cell spread of virus, for instance from macrophages in seminal ﬂuid to female lymphocytes, when epithelial integrity is breached. Once CD4+ CCR5+ memory T cells become infected, a vaccine-induced cellular immune response becomes important. These T cells could, in theory, destroy HIV-1infected cells before release of new virus particles (52) and act with the neutralizing antibody to limit early spread and abort infection (Figure 1). This is probably how many vaccines to other viruses actually work. But for HIV-1 there is no evidence that abortive infection occurs in humans, although it has been suggested experimentally in chimpanzees (53). There is, however, one line of evidence that suggests it could occur in humans. Highly HIV-1-exposed and -uninfected people have been identiﬁed, possibly comprising up to 5% of people at risk (54, 55).

Several studies have shown that they make HIV-1-speciﬁc CD8+ T cell and/or CD4+ T cell responses (56–59), but at variable levels (60). HIV-1-speciﬁc T cells have also been found in cervical mucosal lavage samples from exposed uninfected female sex workers (61). Some studies have claimed that mucosal IgA speciﬁc for HIV-1 is present, but this antibody in the female genital tract can be hard to distinguish from contamination by seminal ﬂuid IgA from infected partners (62, 63). The level of T cell response is often variable, possibly related to ﬂuctuating levels of HIV-1 exposure. One study of sex workers who had been exposed but remained uninfected found that later reduction in exposure to HIV actually increased susceptibility to infection (64). There are several possible explanations for these ﬁndings. The exposed uninfected people may actually be infected with HIV-1 at a low level, e.g., in GALT (65), and this infection may protect them from superinfection, by either immune mechanisms or retroviral interference [where there is target cell depletion and/or receptor blockade (66)]. Alternatively, the immune response in exposed uninfected people may simply reﬂect their undoubted exposure to HIV-1, but they are protected from infection by some other mechanism. Protective genes such as the ∂32 allele of CCR5, which abrogates expression of this virus coreceptor in homozygotes, have been excluded (57), but there could be others yet to be discovered. Finally, it is possible that these uninfected people really have been immunized, either by defective virus or by competent virus that has been cleared, and that this has given them a protective immune response. If such an immune response is protective, it must be remarkably efﬁcient because quantity of HIV-speciﬁc T cells in the blood is nearly always very low; however, the ﬁnding of HIV-1-speciﬁc T cells in the genital mucosa might mediate this protective effect by being in the right location (61). Given the lack of neutralizing antibody in such people, protection may result from abortion of very early infection by the CD8+ T cell response. www.annualreviews.org • HIV Vaccines

EBV: Epstein Barr virus

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SHIV: chimeric simian-human immunodeﬁciency virus

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SCID: severe combined immunodeﬁciency

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These results are therefore very intriguing, but proving that the immune responses actually protect remains difﬁcult. Once infection is established and spreading with high viremia, CD8+ T cell responses induced by vaccines may be the best means of damage limitation (Figure 1). Although they may be incapable of clearing infection, they may contain it for a long time. A vaccine that could exert this level of control and make HIV-1 more like HIV-2, and no more threatening than chronic EBV infection, would be a great gain. There is a widespread view that this might be the best that can be offered by a prophylactic T cell–stimulating vaccine and that this is achievable (see below); hence, a large number of vaccines and clinical trials are addressing this hypothesis. These trials include not only prophylactic vaccine trials but also therapeutic vaccination aimed at enhancing CD8+ T cell–mediated control of HIV-1, usually combining a course of antiretroviral drug therapy (ART) and vaccination (Figure 1, and see below).

NEUTRALIZING ANTIBODY VACCINES Stimulation of neutralizing antibodies with broad speciﬁcity for all HIV variants by a vaccine is undoubtedly the best approach, if it can be achieved. Passive administration of one or more broadly neutralizing monoclonal HIV-1-speciﬁc antibodies to macaques immediately before infection with SIV-HIV hybrid virus (SHIV) that has an HIV-1 envelope prevents infection (67). Similar results were obtained in mice with severe combined immunodeﬁciency (SCID) that were reconstituted with human lymphoid tissue and then infected with HIV-1 (68). Vaginal administration of such antibodies to female macaques also protected against vaginal challenge (69, 70). In contrast, administration of these antibodies after infection is too late; the established virus simply escapes by mutation (68). The message is clear: The presence of neutral232

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izing antibody, at high titer, before infection protects. When HIV was ﬁrst identiﬁed and sequenced, many thought it would be straightforward to make the virus envelope protein gp120 and create a vaccine. This idea has been thwarted by the intrinsic variability of the virus and the differing susceptibility to neutralization of virus cultured on cell lines in vitro compared with primary isolate virus (71). One gp120 vaccine, made by VAXGEN, was taken to two phase III efﬁcacy trials in the United States and Thailand, but there was no protection against HIV-1 infection (72, 73) (see Table 2, below). The problem lies in the structure of the envelope. Two seminal contributions have been the solutions of two crystal structures of HIV-1 gp120. The ﬁrst HIV-1 gp120 crystal was bound to CD4, as well as to a Fab antibody that is speciﬁc for the CCD5/CXCR4binding sites (74, 75). The second structure, SIV gp120, was on its own, not bound to any ligands (76). Given the functional and sequence homologies of HIV and SIV envelopes, these structures clearly explain much of the difﬁculty in ﬁnding neutralizing antibodies and show that there are serious obstacles ahead. Figure 2 shows the key features of the structure of gp120 and the conformational changes it undergoes on binding its ligands (77). Gp120 and the linked gp41 are homotrimers. Thus, much of the surface of monomeric gp120 is hidden and not accessible to antibody (75). The exposed surface is heavily glycosylated (76); the sugars are put there by host enzymes so that the carbohydrate moieties themselves are very unlikely to be immunogenic in humans. The importance of the sugars in protection against neutralization by antibodies is illustrated by an experiment in which macaques were infected with SIV that had mutations designed to remove glycosylation sites around the V1/V2 loops; the mutations were designed to require more than one mutation to repair. When macaques were infected with this virus, there was good

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Figure 2 Changes in the structure of the HIV envelope on binding to target cells. (a) The heavily glycosylated envelope comprises gp120 and gp41 as a trimer. This structure was solved by Chen et al. (76) for SIV. (b) When the envelope binds to CD4, the gp120 undergoes a conformational change that forms and exposes the binding site (shown as red β sheet changing to green) for the coreceptor CCR5 or CXCR4. This structure was determined by Kwong et al. (74). (c) Subsequent to this event, the gp41 undergoes conformational changes to mediate fusion of the membranes so that the virus enters the target cell. [Figure from Kwong et al. (77), with permission of the author and the editors of Nature.]

initial control with potently neutralizing antibody, but after a few weeks virus escaped neutralization, and the sugars had been put back at the nearest site where this could be achieved by a point mutation (78). The CD4-binding site is deeply recessed (75) and is also guarded by the V1/V2 loops that can be varied in the virus without any ﬁtness penalty. The CCR5-binding site is completely absent in the unliganded structure and appears after a large conformational change in the structure after CD4 binding (76). In addition, this site is guarded by the V3 loop, which also can vary without cost to the virus, although the sequence does inﬂuence the choice of CCR5 or CXCR4 coreceptor. Thus, the virus has evolved in a remarkable way to be able to evade neutralizing antibody attack during the chronic phase of infection. Two potential Achilles heels have to be preserved, the CD4-binding site and the CCR5binding site, but both are well hidden. Most of the exposed surface where a binding antibody could sterically interfere with envelope function is protected by the sugars (75).

Many monoclonal antibodies speciﬁc for HIV envelope gp120 and gp41 have been made, both in mice and in humans, by immortalizing EBV-transformed B cells or by phage display. Out of these, only ﬁve have been found that are broadly neutralizing (79–83). These antibodies are summarized in Table 1. One antibody, b12, sees the CD4-binding site, having a long heavy chain CDR (complementarity-determining region) 3 loop. This good neutralizing antibody binds to gp120 with minimal entropic penalty, in contrast to poorly neutralizing antibodies to the same site that bind with an energetic handicap because of induced conformational changes (83a). Another antibody, X5, sees the CCR5/CXCR4-binding site, which is exposed only after CD4 binding, but this antibody only works as a Fab fragment. One extraordinary antibody, 2G12, sees a complex mannose determinant made up of mannans linked to the asparagines at positions 289, 332, and 396 (84). The spatial epitope dimensions are greater than normal for an antibody-binding site; this is explained by the www.annualreviews.org • HIV Vaccines

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The five monoclonal antibodies that broadly neutralize HIV-1

Antibody

Specificity

Special features

Reference

b12

CD4-binding site

Long CDR3 region

(165)

X5

CD4 induced (coreceptor-binding site)

Fab only effective

(166)

2G12

Complex mannose

Antibody has VH domain swap

(80)

2F5

Conserved membrane proximal domain of gp41

IgG3 with long CDR3; cross-reacts with cardiolipin

(85, 167)

4E10

Conserved membrane

IgG3 with long CDR3; proximal domain of gp41 cross-reacts with cardiolipin

(79, 85)

remarkable ﬁnding that 2G12 has mutations in the heavy chain hinge region that cause a crossover of the variable regions to match the opposite light chains, making a single, large antigen-binding site (80). Failed attempts to ﬁnd other similar antibodies imply that this is a very rare if not unique antibody. Two broadly neutralizing human monoclonal antibodies that bind to the membrane proximal region of gp41 have been found; the two adjacent epitopes can be represented by linear peptide sequences. Both antibodies 2F5 and 4E10 are unusual in having long CDR3 regions. A structural analysis of 2F5 binding to the gp41 peptide indicated that the hydrophobic tips of the heavy chain CDR3 region interact with membrane lipid, extending the binding interactions (84a). Some autoantibodies have similar features, and when these monoclonal antibodies were tested against clinically relevant autoantigens, both were found to bind with high afﬁnity to cardiolipin (85). Using the peptides to scan for the presence of such antibodies in large numbers of infected patients has shown that these speciﬁcities are very rarely present. Self-tolerance of B cells likely makes generating antibodies with these speciﬁcities after infection or vaccination almost impossible (85). Thus, although these antibodies are very potent, their special features ensure that they probably cannot be reproduced by vaccination. The VAXGEN gp120 was the last of a series of early envelope vaccines that failed. Although some trials in progress still use gp120 234

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as a boost for a recombinant virus or DNA prime, nearly all current trials are of T cell– stimulating vaccines. At the same time, much effort is being put into design of immunogens that represent the intermediate conformational variants or the liganded gp120 structure (Figure 2). There is also a search for further broadly neutralizing determinants that might be more amenable to vaccine design; however, such determinants may not exist. Characterization of very early virus isolates, obtained before the immune response has a chance to mold the virus envelope, might reveal further sites that could be targets for neutralization, if the earliest infecting virus that grows unimpeded by antibodies has novel structural properties in its envelope that are normally rapidly selected out.

VACCINES THAT STIMULATE T CELL RESPONSES As described above, in natural HIV-1 infection and experimental SIV infection the primary CD8 T cell response clearly follows the virus, peaking just after virus load starts to decline (16, 17, 22, 24). The best hope for a T cell–inducing vaccine is to ensure a more rapid secondary T cell response targeted to mucosal sites so as to act at the early stages of infection and to eliminate the virus at this time (Figure 1). However, despite the puzzle of the exposed uninfected people, there are as yet no studies of SIV vaccination and challenge in macaques that show that this can

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happen. A more likely scenario is that vaccinestimulated memory T cells will need (a) expansion by cell division, (b) differentiation to acquire the capacity to kill virus-infected cells, and (c) relocation to mucosal sites and draining lymph nodes. Thus, the window for being able to abort the initial infection may be too narrow. In this scenario, T cell–inducing vaccines will limit damage by controlling HIV-1 as effectively as EBV is controlled in most of us. Although there is good evidence that CD8 T cells suppress virus, how they do it and what functions will be required of vaccine-stimulated T cells is unclear. For long-term asymptomatic control of human T-lymphotropic virus type 1 (HTLV-1), T cell–mediated killing is important (86), but for hepatitis B virus, IFN-γ production is critical (87). For HIV-1, this information is badly needed, as different vaccine approaches, like different virus infections (42), are likely to stimulate T cells with different functional capabilities. In this context, we need to know what type of immune response to measure after vaccination. At present, all vaccine clinical trial teams use peptide antigenstimulated IFN-γ production as their main assays of vaccine immunogenicity, either using the ELISPOT method or intracellular cytokine production measured by ﬂow cytometry. However, this could be the wrong function to measure, although there is a chance that antigen-stimulated IFN-γ production could be a surrogate for the real protective functions. The critical information could come from studies of those already infected. Clearly, some T cell responses drive virus escape and must therefore be suppressive, whereas others appear to offer little protection, for example those that were not evaded by superinfecting virus (88). We do not know all T cell functions, and some of the less well deﬁned, such as the CD8+ cell antiviral (HIV-1) factor (CAF), could be important (89). We may not be able to design effective vaccines until we understand the most effective antiviral activities of CD8+ cells.

The CD8 T cell vaccine approach has been built on a series of inﬂuential studies in the rhesus macaque–SIV challenge model that demonstrate partial protection by vaccines that stimulate CD8 T cell immunity. Immunogens encoding or presenting internal virus proteins such as gag stimulate strong CD8 T cell responses. The vaccines include plasmid DNA encoding HIV genes, recombinant modiﬁed vaccinia virus Ankara (MVA), recombinant adenovirus-5, and recombinant vesicular stomatitis virus, as well as combinations of some of these (90–93). In all these experiments, the challenge virus was an aggressive SIV-HIV hybrid virus SHIV89.6P that expresses a CXCR4-speciﬁc HIV envelope (94) and causes very rapid loss of CD4+ T cells and onset of immunodeﬁciency. In no case did the vaccine give sterilizing immunity; all vaccinated animals were infected, but they did not lose CD4 T cells, and they remained healthy with virus loads 1000-times lower than the unvaccinated controls. However, there is a possible problem: Paradoxically, this virus appears to be easier to control by vaccination than some others, and the same vaccine approaches have been much less effective at controlling infection with the CCR5 using viruses SIVmac239 or SIVmac251 (95– 97), although temporary partial reductions in virus level have been seen (98). Furthermore, immunization of macaques with a phagedisplay peptide vaccine aimed at stimulating neutralizing antibody gave a similar level of protection against SHIV89.6P, suggesting that this aggressive virus may be relatively easy to control (94). Clearly, this issue needs further exploration, in particular to determine whether the apparent differences relate to the different coreceptors used by SHIV89.6P (CXCR4) and SIVmac239 (CCR5). The early evidence that the SHIV89.6P challenge virus is controlled by CD8 T cells was indirect; in some cases (92) the immunogens did not include envelope, and no neutralizing antibodies were stimulated. But now direct evidence exists (99): In an initially protected vaccinated animal that started to lose www.annualreviews.org • HIV Vaccines

MVA: modiﬁed vaccinia virus Ankara

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CD4 T cells with a rise in blood virus titer, a mutation was observed of a single amino acid in the immunodominant gag epitope, making it unable to bind the presenting MHC molecule (Figure 3). Thus, at least in this animal, the protection was likely mediated by T cells with this speciﬁcity. There are as yet no equivalent data in humans. In one clinical case, superinfection by a second B clade virus in an HIV-infected patient occurred despite a very strong pre-existing CD8+ T cell response, but the new virus was different in about half the epitopes, and the quality of the T cell response could have been impaired by poor CD4+ T cell function (88). Also, we do not know how often chronically HIV-1infected people resist superinfection. Stimulation of CD8+ T cell responses with vaccines to levels similar to those that protect macaques from SHIV is proving difﬁcult in humans. Vaccines that stimulate strong T cell responses in mice may fail completely in humans, and, although immune responses stimulated in primates should be closer to those seen in humans, they may not always be predictive. Key issues could be vaccine dose, different pattern speciﬁcity of Toll-like receptors (TLRs) in different species, degree of vector attenuation in different species, cross-reactivity with self-antigens or epitopes previously encountered, and unrepresentative T cell responses to highly immunodominant epitopes. For priming CD8 T cell responses,

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vaccine antigen needs to enter the MHC class I–processing pathway. There are three routes: (a) direct entry by using intracellular pathogens as vectors; (b) transfection for DNA plasmids; or (c) cross-priming by DC, which can take up protein antigen from outside the cell. The last is increasingly recognized as an important route of priming and, unlike the direct route, favors antigen that is stable rather than rapidly degraded (100). Guided by these principles, several vaccine delivery systems are under development. The key candidates are shown in Table 2, which indicates the considerable activity in the past two years, with 35 trials in progress and at least another 5 completed but not yet formally reported. Most experience so far has been obtained with plasmid DNA, MVA, canarypox, fowlpox, and adenovirus-5. More recent additions are adeno-associated virus (AAV), Venezuelan equine encephalitis virus, and lipopeptides. Some of these vaccine vectors (MVA, fowlpox, adenoviruses) are also being used for stimulation of T cell responses to other pathogens, including malaria, mycobacterium tuberculosis, human papilloma virus, and tumor antigens. The HIV genes/proteins most commonly include gag, pol, nef, and env, but all genes are being used. The subtypes are most commonly B or C, but also include A, D, and E strains. Because these vaccine candidates are used in clinical trials that are very highly

−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→ Figure 3 Escape of SHIV89.6P from T cell control in a vaccinated macaque shows that protection is mediated by CD8+ T cells. The top graph shows the time course of RNA virus load in the macaque that had been vaccinated by DNA-expressing gag and IL-2 and challenged with SHIV89.6P at week 0. Between weeks 10 and 20 there was good virus control with a low virus set point; the unvaccinated controls had virus loads ∼106 copies per ml (not shown here). After 20 weeks, virus load started to rise, and the CD4 T cell count fell dramatically (middle graph). At the same time, the strong CD8+ T cell response to the immunodominant gag epitope p11C declined, measured by tetramer staining of CD8 T cells (lower graph). These events after week 20 were associated with a selection of an escape mutant in the immunodominant epitope in gag p24 p11C 181–189 sequence shown at the top (single letter amino acid code). At week 20 and thereafter the threonine (T) at position 182, a residue that anchors the peptide to the presenting Mamu A∗ 01 MHC molecule, was mutated to isoleucine (I). This new peptide does not bind to Mamu A∗ 01 and is therefore an escape mutant that is represented in all virus sequences (indicated in parentheses as the number of variant sequences/number of sequences determined). [Figure adapted from Barouch et al. (99), reproduced with permission of the authors and editor of Nature.] 236

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regulated, even a simple phase I trial can often take two years, and the clinical work may then be followed by more than a year for data analysis. This lengthy process means that there are to date very limited published data on

many of the HIV candidates. Earlier studies of vaccine-stimulated T cell immunity generally used less quantitative assays, such as T cell killing after culture in vitro or proliferation in vitro measured by thymidine incorporation

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Gag p11C (181–189) week 0 week 14 week 20 week 24 week 28 week 36 week 44

CTPYDINQM

––––––––– ––––––––– – I ––––––– – I ––––––– – I ––––––– – I ––––––– – I –––––––

(15/15) (8/8) (10/10) (11/11) (11/11) (11/11) (10/10)

7.0 6.0

RNA (copies / ml)

5.0 4.0 3.0 2.0 1200

800

+ CD4 T cells / µl 400

0 20 15

p11C tetramer (percent)

10 5 0

0

10

20

30

40

50

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(101). Since 2002 the assays have been improved and validated by interlaboratory studies (e.g., 102), making them more reliable. Nevertheless, from the more recent studies, some general points have been reported at conferences, and some conclusions can also be drawn from studies with similar types of vaccine in other ﬁelds.

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1. All the vaccine candidates are strongly immunogenic for CD8 T cell responses in mice. 2. Most, or all, vaccine candidates stimulate measurable T cell responses in macaques (e.g., 92). 3. Some candidates, including DNA given together with IL-2, DNA and MVA in a DNA prime with MVA boost combination, adenovirus-5, and vesicular stomatitis virus, partially protect macaques against SHIV-89.6P virus challenge (see above). However, protection against SIVmac239 has been much harder to achieve and has usually been marginal (95). 4. DNA stimulates weak T cell responses in humans that are predominately CD4 T cells (103, 104). 5. MVA, canarypox, and fowlpox stimulate weak primary T cell responses in humans (104, 105). 6. The DNA prime with an MVA boost gives stronger responses than either alone in humans; responses are mostly CD4+ T cells. Responses peak at day 7 after MVA and are short lived (104; N. Goonetilleke, S. Moore, N. Winston, A. Mahmood, S. Pinheirho, J. Roberts, T. Hanke, A. McMichael, unpublished data). 7. MVA can boost well-primed CD8+ T cell responses (106). 8. Recombinant replication defective adenovirus-5 gives fairly strong and sustained CD8+ T cell responses in humans (107). 9. Pre-existing antibodies to adenovirus5, which are present at high frequencies 238

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in many populations, reduce immunogenicity of recombinant adenovirus-5 vaccines (107, 108). DNA-induced responses are dose dependent in macaques, needing 2–5 mg to obtain responses (91). This dose is close to the maximum that is practical, and humans may need much larger doses to see equivalent T cell responses. Adding adjuvants to DNA vaccines may enhance their immunogenicity, and various combinations of cytokines, chemokines, and TLR ligands are under study. Also, delivery devices such as the gene guns that ﬁre particles of gold coated with the vaccine into the dermis/epidermis may enhance immunogenicity (109). The pox virus vectors are probably at a disadvantage because ∼200 pox virus proteins may compete to stimulate a T cell response, but this may be less of a problem when used to boost after priming with a different vector recombinant for the same vaccine antigen. MVA may be too attenuated compared with adenovirus-5, which may persist sufﬁciently to give a longer lasting T cell response. The responses to some vaccines may be underestimated by the IFN-γ ELISPOT assay; more sensitive assays for which the response is measured after several days of culture with the antigen might better reﬂect the presence of central memory T cells (104). This approach has revealed many more T cell responses than the conventional assays in vaccine recipients. The approaches above show what some of the vectors can do, and although somewhat disappointing they have established working methods and validated assays. Furthermore, for some vectors, MVA for instance, they have given an indication of what their role might be (stimulating CD4+ T cell responses after DNA priming and boosting well-primed CD4+ and CD8+ T cell responses) or, for adenovirus vectors, how they might be improved to avoid pre-existing immunity. There is little doubt that new immunogens, vectors, delivery systems, and vaccine combinations will be found that stimulate strong CD8+ and CD4+ responses to

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Table 2

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Current and recently completed (unpublished) HIV vaccine clinical trialsa

Delivery

Clade

Immunogen

Phaseb

DNA

A, B, C

e, g, p, n

I

DNA

C

g, p

I

DNA

C

e, g, p, n, t

I

DNA

A, B, C

g, p, n, e

I

DNA

A, B, C

e, g, p, n

I

DNA

A, B, C, D, E

epitopes

I

DNA

B

g

I

DNA

B

g, p

I

DNA

B

e, g, p, t, r, n

IIA

DNA

A, B, C

e, g, p, n

IIA

DNA+120

B

e

I

DNA+MVA

A

g

I×3

DNA+MVA

A

g

IIA × 2

DNA+FPV

B

e, g, p, r, t, v

IIA

DNA+Ad5

B

g

I

Lpep

B

g, p, n

I

Lpep

B

g, p, n

IIA

Lpep+CPV

B

g, p, n

IIA

MVA

A

g

I

MVA

C

e, g, p, n, t

I

MVA+FPV

B

e, g, p, t, r, n

I

CPV

B

e, g, p

I

CPV+120

B, E

e, g, p

III

Ad5

A, B, C

g, p, n

I

Ad5

B

g

I

Ad5+CPV

B

g, p

I

Ad5

B

g, p, n

IIB

AAV

C

g, p, n, t

I

VEE

C

g

I

Protein

D

e

I

Protein

B

t

I

Protein

B

e

I

120

B

e

III

120

E

e

III

VLP

B

e, g

I

VLP

C

e

I

a Abbreviations: DNA, plasmid DNA; MVA, modiﬁed vaccinia virus Ankara; 120, glycoprotein gp120; FPV, fowlpox virus; CPV, canarypox virus; Ad5, adenovirus-5; Lpep, lipopeptide; AAV, adeno-associated virus; VEE, Venezualan equine encephalitis virus; VLP, virus-like particle; e, HIV-1 envelope; g, HIV-1 gag; p, HIV-1 polymerase; n, HIV-1 nef; t, HIV-1 tat; v, HIV-1 vpr. b Phase I trials are in HIV-1-uninfected low-risk volunteers, n < 100; Phase IIA trials are in HIV-1-uninfected low-risk volunteers, n > 100; Phase IIB trials are proof-of-principle (efﬁcacy) trials in HIV-1-uninfected high-risk volunteers, n > 100; Phase III trials are powered efﬁcacy trials in HIV-1-uninfected high-risk volunteers, n > 5000. For more details of vaccines currently in trial, see http://www.iavireport.org/trialsdb/.

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Figure 4 Worldwide distribution of the major HIV-1 subtypes (clades) and HIV-2. The subtypes A–H are subdivisions of the M strain; the O and N strains are rare, but are very close to SIV strains found in chimpanzees.

recombinant antigens. Also, immunization protocols that optimize the size of response and include mucosal immune responses will be worked out. However, the process has to be done through clinical trials that are very highly regulated, and so progress is slow. Standing back to view the mass of current trials, one is excited that so many groups are working on HIV-1 vaccines but disappointed by the degree of repetition and by the extent to which current trials aim to test only a single hypothesis: that CD8 T cells induced by vaccination will protect against HIV-1 as well as they do against challenge viruses such as SHIV89.6P in macaques. However, if HIV-1 behaves more like SIVmac239, investigators in this ﬁeld will face serious difﬁculty and will need much more basic information about what type of T cell response is needed. In addition, virus variability is likely to be a serious problem.

VIRUS VARIABILITY AND VACCINE DESIGN HIV-1 is highly variable, and investigators have argued that this alone could compromise the success of any vaccine that goes into efﬁcacy trials (73). Phylogenetic tree analysis of virus RNA and protein sequences reveals considerable heterogeneity (110). There are three 240

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main types of HIV-1, M, N, and O, that are thought to represent separate introductions into humans from chimpanzees. The main M type is further divided into six principle subtypes, or clades, A, B, C, D, E, and G, that represent early branching of the virus in its evolution in humans (110). The clades differ by ∼20% of amino acid sequence and have distinct geographical distributions, as shown Figure 4. All the clades are found in central equatorial Africa, where the transfer of virus from chimpanzees to humans probably ﬁrst occurred. In most of the other countries, there is a single dominant clade, probably representing a founder effect. Within the clades there is further variability in up to 10% of the amino acid sequence. Despite this huge heterogeneity, some features of the sequences suggest conservation as well. There are clear consensus sequences for each clade that suggest ﬁtness costs as the virus varies so that there is a tendency toward conservation. Much of the intraclade sequence variability in the virus core and regulatory proteins is probably driven by immune escape from HLA class I–restricted CD8+ T cells. Moore et al. (40) have elegantly demonstrated that common HLA class I types in a population are associated with virus sequence variability in individual patients, the same HLA types selecting the same mutations

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repeatedly (40). It is also notable that when escape mutations have been monitored closely, they tend to reiterate known variability in the very large virus sequence databases (111, 112). This tendency to mutate repeatedly in the same way suggests that there are constraints on virus sequence variability, with possible ﬁtness costs for some escape mutations. In at least two cases, a single escape mutation that affects epitope presentation is strongly associated with other sequence changes that are probably compensatory (38, 113). Also, when the virus is transmitted to partners or infants of different HLA type, the sequence may revert to consensus (111). This suggests a balance between immune control, virus escape, and viral ﬁtness. There are very few, possibly no, HIV-1 epitopes from which the virus cannot escape by changing one or more amino acids. Envelope variability is largely driven by neutralizing antibody. Much of the change occurs in the hypervariable loops that have little or no structural role and therefore no ﬁtness cost, although changes in the V3 loop contribute to coreceptor selection, with CCR5 speciﬁcity changing to CXCR4 speciﬁcity (114). The ﬁve broadly neutralizing monoclonal antibodies discussed above are less completely cross-neutralizing on non-B clade viruses, compatible with some structural variation, for instance in the distribution of glycosylation sites (82). Further analysis is needed to determine whether there might be different broadly neutralizing epitopes for the different clades. For the CD8 T cell response, vaccine designers need to know how sensitive T cell clones are to virus variability. There has been a widespread notion in the vaccine ﬁeld that T cells can cross-react quite broadly and therefore that a vaccine might not have to match the prevalent virus subtype (115). The idea of cross-reactive cytotoxic T cells may have come from very early studies on antiinﬂuenza-speciﬁc T cells that cross-reacted across inﬂuenza virus subtypes; however, this cross-reaction occurred because the T cells

respond to conserved internal virus proteins rather than to the variable surface glycoproteins (116). One study that emphasizes this point explored T cell recognition of all 171 single amino acid variants of an HIV gag nonamer epitope and found that only one third were recognized, even though most were bound by the presenting HLA A2 molecule (117). Similar results were found with different T cell clones. These data imply that most sequence variation in an epitope can generate escape mutants. This ﬁnding is compatible with many careful studies of individual escape mutations and the imprinting of sequence variation in the virus by HLA type (40). The implication for vaccine design is clear: Vaccine immunogen sequence must match the sequence of prevalent virus in the population to be vaccinated. At the very minimum, this means matching vaccine to clades. However, it also raises concerns that variability that already exists within HIV-1 subtypes could evade vaccine-induced T cell immunity quite easily. Furthermore, if the vaccine does not prevent infection but allows infection to occur, albeit better controlled than in unvaccinated people, there is a real risk of virus escape if the T cell response is too narrow. This risk was dramatically demonstrated in macaques when a single amino acid change in an immunodominant epitope enabled SIV to escape control in a vaccinated monkey (99) (Figure 3). The problem of virus variability has been raised as an almost insuperable obstacle to a successful HIV vaccine (73). However, there are ways to lessen the chances of virus escape. Having matched the vaccine and virus clades, vaccine developers will need to ensure that the T cell response is directed at several epitopes. The chances of escape are reduced if multiple mutations are needed, particularly if the T cell response can dominate the virus early on with a relatively low level of virus replication. The breadth in T cell response not only refers to number of epitopes seen but also to the clonality of the T cells. The phenomenon of immunodominance could be a serious obstacle www.annualreviews.org • HIV Vaccines

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ART: antiretroviral drug therapy

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to vaccine design and must be understood if it is to be avoided. Immunodominance can focus the majority of the response to a few, or even one, of the epitopes and is usually apparent in T cell responses to HIV (39, 118). Indeed, immunodominance makes the T cell responses predictable according to HLA type of the infected person. These large T cell responses probably exert more selective pressure for virus escape. Possible mechanisms of immunodominance include the selection of epitopes during antigen processing, differing afﬁnity of different epitopes on the presenting HLA molecules, and half-life at the cell surface (119). Similarly, differing precursor frequencies of particular T cell clones, possibly inﬂuenced by cross-reactions to previous infections, can inﬂuence which T cell responses dominate (120). T cell receptor avidity for particular peptide-HLA complexes may also have an inﬂuence. This avidity is more complex than simply one-to-one afﬁnity and is not a matter of highest afﬁnity being best. Surface density of the receptor and coreceptors may be important. Competition between T cell clones may ultimately be a major determinant of T cell clone selection (120). Thus, some vaccine regimes aimed at stimulating high-level T cell responses may inadvertently favor narrow T cell responses. This possibility will have to be addressed by direct experimentation in humans, comparing different vaccination regimens. The use of complex mixtures of sequence variants in vaccines may favor more broadly cross-reactive T cell responses but could also run into trouble with T cell receptor antagonism, in which an altered epitope can inhibit the T cell response to the cognate epitope and therefore lessen rather than enhance the response (121). If variants are given sequentially, the response to the ﬁrst variant may dominate and impair the subsequent response to subsequent variants—a kind of original antigenic sin (122). The latter may arise because weak cross-reactivity to the variant gives a stronger signal to the weakly crossreacting memory T cells than to naive T cells McMichael

with more appropriate T cell receptors. This implies some suppression of the primary response by the memory response, possibly involving regulatory T cells (123). Thus, stimulation of inappropriate T cell responses by the vaccine might impair the subsequent response to the infecting virus, although this seems likely to be a more theoretical than real concern. All these issues need to be addressed. Studies in mice and macaques can only be of limited value because of differing MHC types and different rules for different epitopes and T cell receptors. There is a clear need for clinical studies that are small scale but intense in terms of the assays used and that can directly address these questions. To speed up the discovery process, these experimental medicine studies need to be less rigidly regulated than conventional phase I trials.

DAMAGE LIMITATION: A ROLE FOR THERAPEUTIC VACCINATION? If the most likely beneﬁcial effect of a T cell– inducing vaccine is better control of HIV-1 in those vaccinated, the question has to be asked whether therapeutic vaccination of those already infected might achieve this more efﬁciently (124, 125). The ﬁrst attempts at immunotherapy were disappointing (e.g., 126). Immunization with gp120, gag particles, or whole inactivated HIV, but without the use of effective antiretroviral drug therapy (ART), failed to reduce virus load, despite demonstrable increases in T cell immune responses (127–133). A more logical approach is to use a combination of effective (normally triple drug) ART and vaccination. Patients who start ART reduce virus load substantially, and, probably as a direct consequence, the level of T cell response declines (28, 134). When ART is interrupted, virus rebounds within 3–6 weeks to pretreatment levels (29), but although an increased T cell response follows, this does not result in better immune control (135). Some

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investigators have argued that this rebound may be reduced by increasing the T cell response by vaccination to pretreatment levels (124, 125). The idea has been modeled in macaques infected with SIV. When animals that were chronically infected with SIVmac251 were treated with ART and then immunized with a recombinant attenuated vaccinia virus, NYVAC, expressing multiple SIV genes, temporary virus control was achieved after stopping the drugs (136). A similar result was achieved when animals were given ART and immunized with a NYVAC-SIV vaccine in primary infection (137). Control of virus rebound after stopping ART was also seen in SIV-infected monkeys after an intradermal immunization with DC transfected with DNA encoding SIV genes (138). These results are encouraging, although the effects are more modest than the surprisingly large reductions of virus load described by Lu et al. (139), who immunized untreated SIV-infected macaques with autologous DC pulsed with inactivated SIV. Vaccination of HIV-infected people who are on ART with subsequent drug treatment interruption has given more mixed results. A phase III trial of inactivated whole HIV, Remune, failed to show any effect (140). Similarly, when Remune was compared with a recombinant canarypox vaccine in patients who had started ART in acute infection, no effect was seen after 24 weeks of drug treatment interruption with either vaccination (141). Plasmid DNA vaccines encoding HIV genes have been tested for immunogenicity in ARTuntreated and -treated patients but have only stimulated weak T cell responses (101, 142), and drugs were not withdrawn. Several trials are being conducted of recombinant virus vaccines, canarypox, fowlpox, and MVA, with treatment interruptions. Initial reports at conferences indicate good boosting of T cell responses in patients on ART, and in some studies there have been some effects on virus load, e.g., delay in time to reach the virus load at which ART has to be restarted or a small reduction in virus load (143, 144). In another

trial, no reduction of virus load was seen on stopping ART after immunization with recombinant canarypox (145), but Harrer et al. (146), in an uncontrolled study, found a sustained reduction in viral load after immunization with an MVA-nef vaccine and subsequent interruption of ART. Finally, conﬂicting results have been obtained by immunizing patients, who were not on drug therapy, with HLA-matched or autologous DC pulsed with epitope peptides (147) or whole inactivated virus (148). In the former study, no effect on virus load was seen, whereas in the latter experiment, a substantial reduction in virus was seen in approximately half the patients studied. This surprising result, which is similar to a ﬁnding by the same investigators in macaques (139), implies correction of a major defect in DC function, and it needs urgent independent conﬁrmation. Despite rather equivocal results so far, therapeutic immunization is still worth pursuing. Care needs to be taken to deﬁne the quantity and quality of the T cell responses. Interruption of ART, which in effect gives a virus challenge in vivo, has been considered safe because it does not appear to select drug resistance and because virus control can be rapidly regained on treatment resumption (29). Therefore, this approach provides an opportunity to study correlates of protection, as virus control by the immune response is regained, and to modify and improve such responses. There is real scope for improving the quality of the anti-HIV-1 immune response (e.g., enhancing HIV-1-speciﬁc CD4 T cell activity or refocusing T cell responses on conserved epitopes) before treatment interruption. One could argue that positive effects of vaccination are likely to be seen more rapidly in this scenario than in the painfully slow efﬁcacy trials of prophylactic vaccines that are only just beginning. If a reliable way of gaining lasting immune control could be found, it would offer possibilities for therapy in developing countries at relatively low cost. It is also noteworthy that original antigenic sin could work in www.annualreviews.org • HIV Vaccines

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favor of the patient because if the vaccine did not match the virus perfectly it might nevertheless boost T cells highly speciﬁc for autologous virus—in contrast to prophylactic vaccine for which, in theory, vaccination that sets up responses to the wrong epitope variants could impair natural immune responses to a later infecting virus. Finally, such an approach would offer something for the millions who are already infected with little hope of access to effective ART and for whom prophylactic vaccination offers nothing.

ARE THERE OTHER APPROACHES TO HIV-1 VACCINATION? Three other approaches to protection against SIV appear to work in macaques but have not been fully explained. In the ﬁrst approach, Stott et al. (149) showed that macaques that had been vaccinated with inactivated SIV that had been grown on human cells were fully protected against challenge with SIV that had also been grown on humans cells, but not against SIV that had been grown on macaque cells. Further investigation implicated an immune response, probably antibody, to HLA antigens, which are incorporated into virions as they bud at cell surfaces (150). However, there has been no convincing evidence that allo- (rather than xeno-) antibodies to HLA antigens can protect against HIV-1. This approach might be worth revisiting now that there are better ways of characterizing both antigens and antibody immune responses. Induction of antibodies to self-antigens such as CCR5, although probably not to HLA, might be safe and feasible (151), given that people who are homozygous for the CCR5 ∂32 gene and who lack the protein are healthy and resistant to HIV-1 infection (152). In the second approach, Desrosiers and colleagues (153) showed that infection of macaques with attenuated SIV offered good protection against superinfection with the unattenuated parent strain of SIV. Re244

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searchers usually argue that the protection is due to an immune response, but sustained efforts to identify which components of the immune response are responsible have not given clear answers (73). In particular, the protection cannot be transferred by passive antibody (154), and protection could only be partly abrogated by depletion of CD8 T cells (155) or of all T cells (156). Furthermore, the level of measured immune response does not correlate with efﬁcacy (73). Protection against challenge with nonhomologous virus was less complete, with a median virus load reduction of 100-fold rather than 1000-fold that could reﬂect less efﬁcient immune recognition of variant epitopes (157); this is a key experiment that, although not perfect, may be the best evidence that the protection is mediated by a speciﬁc immune response. However, there could also be involvement of innate immune responses at the sites of virus infection. Innate antiviral function could be chronically activated by the persisting infection with the attenuated virus. It is also possible that a component of the protection is not immune at all. Retroviral interference in vitro or in vivo occurs when the ﬁrst virus depletes susceptible target cells or blockades the receptor so that the second virus cannot infect (66). This interference could explain why protection is seen before development of detectable immune responses (158). It could also account for the escape of virulent wild-type SIV in vivo in two macaques infected with an attenuated SIV with a 4 amino acid deletion in nef, which was repaired in the escaping virus (159). It is hard to explain this escape in the presence of an immune response that was able to protect against deliberate superinfection with a large dose of virulent virus in the same animals. Given the much greater susceptibility to infection of gut-associated memory T cells compared to other peripheral lymphoid tissue (160), it is possible (and testable) that attenuated SIV could infect and destroy the gut-associated T cells that are most susceptible to SIV infection. The difference

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between the virulent and attenuated virus might be more related to the virus’ ability to activate and induce apoptosis in uninfected bystander T cells (14, 161). Thus, retroviral interference needs to be seriously reconsidered as one of the possible mechanisms behind the protection in animals infected with attenuated SIV. Whatever the mechanism of protection against attenuated SIV, this approach is very unlikely to be possible in humans, even though other attenuated virus vaccines have an excellent record in human vaccine successes. The possible use of attenuated HIV-1 would trigger concerns about long-term safety, heightened by instances of AIDS in macaques infected with attenuated SIV (162). The third approach was described by Lifson et al. (163), in which short-term treatment of macaques with ART within 48 h of SIV infection resulted in long-term control of the infection after withdrawal of the drugs and resistance to superinfection. Given the time frame of early virus spread (Figure 1), this type of protection is likely to be immune rather than retroviral interference because virus should not have disseminated at the time of ART introduction and virus suppression. This conclusion is largely conﬁrmed by the ﬁnding that infusion of anti-CD8 antibody resulted in large increases in viremia

of the infecting virus; however, the measured T cell immune responses were low (163), so the exact immune correlates of protection in this model need to be found.

CONCLUSIONS The design of an effective HIV-1 vaccine remains an enormous challenge. Nearly all previous vaccine successes have been built on traditional empirical approaches. So far, these have failed for HIV-1. Immunologists are therefore, for the ﬁrst time, attempting to design a real vaccine. This challenge asks penetrating questions about our understanding of the immune response. Although we have a good primate model of HIV infection, we ultimately have to test the vaccines in humans. Human testing is a very slow process because of the very complex safety and regulatory issues. There is now widespread recognition of the scale of the problem in the scientiﬁc and funding communities (164), and more coordinated approaches involving extensive international collaborations are essential. But this alone will not be enough. New ideas based on deeper understanding of all aspects of adaptive and innate, peripheral and mucosal immune responses are badly needed. The challenge, therefore, is to all of the immunological community.

DISCLOSURE STATEMENT A.J.M is cofounder and shareholder (<2.5%) of Oxxon Therapeutics.

ACKNOWLEDGMENTS The views expressed in this review are my own, but I have been greatly helped and inﬂuenced by extensive discussions with my Oxford colleagues, Tomas Hanke, Lucy Dorrell, Nilu Goonetilleke, and Seph Borrow, and with many other friends in the ﬁeld, notably Sarah Rowland-Jones, Peter Kwong, Bart Haynes, Norman Letvin, Dan Barouch, George Shaw, Beatrice Hahn, Joe Sodroski, Mike Cohen, David Monteﬁore, Wayne Koff, Jill Gilmour, Pat Fast, Brigitte Autran, Bruce Walker, and Ita Askonas.

LITERATURE CITED 1. Willerford DM, Bwayo JJ, Hensel M, Emonyi W, Plummer FA, et al. 1993. Human immunodeﬁciency virus infection among high-risk seronegative prostitutes in Nairobi. J. Infect. Dis. 167:1414–17 www.annualreviews.org • HIV Vaccines

245

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2. Gray RH, Wawer MJ, Brookmeyer R, Sewankambo NK, Serwadda D, et al. 2001. Probability of HIV-1 transmission per coital act in monogamous, heterosexual, HIV1-discordant couples in Rakai, Uganda. Lancet 357:1149–53 3. Vittinghoff E, Douglas J, Judson F, McKirnan D, MacQueen K, Buchbinder SP. 1999. Per-contact risk of human immunodeﬁciency virus transmission between male sexual partners. Am. J. Epidemiol. 150:306–11 4. Modjarrad K, Zulu I, Karita E, Kancheya N, Funkhouser E, Allen S. 2005. Predictors of HIV serostatus among HIV discordant couples in Lusaka, Zambia and female antenatal clinic attendants in Kigali, Rwanda. AIDS Res. Hum. Retrovir. 21:5–12 5. Fideli US, Allen SA, Musonda R, Trask S, Hahn BH, et al. 2001. Virologic and immunologic determinants of heterosexual transmission of human immunodeﬁciency virus type 1 in Africa. AIDS Res. Hum. Retrovir. 17:901–10 6. Li Q, Duan L, Estes JD, Ma ZM, Rourke T, et al. 2005. Peak SIV replication in resting memory CD4+ T cells depletes gut lamina propria CD4+ T cells. Nature 434:1148–52 7. Pope M, Haase AT. 2003. Transmission, acute HIV-1 infection and the quest for strategies to prevent infection. Nat. Med. 9:847–52 8. Fong L, Mengozzi M, Abbey NW, Herndier BG, Engleman EG. 2002. Productive infection of plasmacytoid dendritic cells with human immunodeﬁciency virus type 1 is triggered by CD40 ligation. J. Virol. 76:11033–41 9. Fonteneau JF, Larsson M, Beignon AS, McKenna K, Dasilva I, et al. 2004. Human immunodeﬁciency virus type 1 activates plasmacytoid dendritic cells and concomitantly induces the bystander maturation of myeloid dendritic cells. J. Virol. 78:5223–32 10. Geijtenbeek TB, Kwon DS, Torensma R, van Vliet SJ, van Duijnhoven GC, et al. 2000. DC-SIGN, a dendritic cell-speciﬁc HIV-1-binding protein that enhances trans-infection of T cells. Cell 100:587–97 11. Kwon DS, Gregorio G, Bitton N, Hendrickson WA, Littman DR. 2002. DC-SIGNmediated internalization of HIV is required for trans-enhancement of T cell infection. Immunity 16:135–44 12. Pope M, Betjes MG, Romani N, Hirmand H, Cameron PU, et al. 1994. Conjugates of dendritic cells and memory T lymphocytes from skin facilitate productive infection with HIV-1. Cell 78:389–98 13. Douek DC, Brenchley JM, Betts MR, Ambrozak DR, Hill BJ, et al. 2002. HIV preferentially infects HIV-speciﬁc CD4+ T cells. Nature 417:95–98 14. Mattapallil JJ, Douek DC, Hill B, Nishimura Y, Martin M, Roederer M. 2005. Massive infection and loss of memory CD4+ T cells in multiple tissues during acute SIV infection. Nature 434:1093–97 15. Phillips AN. 1996. Reduction of HIV concentration during acute infection: independence from a speciﬁc immune response. Science 271:497–99 16. Borrow P, Lewicki H, Hahn BH, Shaw GM, Oldstone MB. 1994. Virus-speciﬁc CD8+ cytotoxic T-lymphocyte activity associated with control of viremia in primary human immunodeﬁciency virus type 1 infection. J. Virol. 68:6103–10 17. Safrit JT, Andrews CA, Zhu T, Ho DD, Koup RA. 1994. Characterization of human immunodeﬁciency virus type 1-speciﬁc cytotoxic T lymphocyte clones isolated during acute seroconversion: recognition of autologous virus sequences within a conserved immunodominant epitope. J. Exp. Med. 179:463–72 18. Ioannidis JP, McQueen PG, Goedert JJ, Kaslow RA. 1998. Use of neural networks to model complex immunogenetic associations of disease: human leukocyte antigen impact on the progression of human immunodeﬁciency virus infection. Am. J. Epidemiol. 147:464–71

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19. Bhardwaj N. 2001. Processing and presentation of antigens by dendritic cells: implications for vaccines. Trends Mol. Med. 7:388–94 20. Pulendran B, Palucka K, Banchereau J. 2001. Sensing pathogens and tuning immune responses. Science 293:253–56 21. Alter G, Malenfant JM, Delabre RM, Burgett NC, Yu XG, et al. 2004. Increased natural killer cell activity in viremic HIV-1 infection. J. Immunol. 173:5305–11 22. Wilson JD, Ogg GS, Allen RL, Davis C, Shaunak S, et al. 2000. Direct visualization of HIV-1-speciﬁc cytotoxic T lymphocytes during primary infection. AIDS 14:225–33 23. Schmitz JE, Kuroda MJ, Santra S, Sasseville VG, Simon MA, et al. 1999. Control of viremia in simian immunodeﬁciency virus infection by CD8+ lymphocytes. Science 283:857–60 24. Evans DT, O’Connor DH, Jing P, Dzuris JL, Sidney J, et al. 1999. Virus-speciﬁc cytotoxic T-lymphocyte responses select for amino-acid variation in simian immunodeﬁciency virus Env and Nef. Nat. Med. 5:1270–76 25. Borrow P, Lewicki H, Wei X, Horwitz MS, Peffer N, et al. 1997. Antiviral pressure exerted by HIV-1-speciﬁc cytotoxic T lymphocytes (CTLs) during primary infection demonstrated by rapid selection of CTL escape virus. Nat. Med. 3:205–11 26. Jones NA, Wei X, Flower DR, Wong M, Michor F, et al. 2004. Determinants of human immunodeﬁciency virus type 1 escape from the primary CD8+ cytotoxic T lymphocyte response. J. Exp. Med. 200:1243–56 27. Price DA, Goulder PJ, Klenerman P, Sewell AK, Easterbrook PJ, et al. 1997. Positive selection of HIV-1 cytotoxic T lymphocyte escape variants during primary infection. Proc. Natl. Acad. Sci. USA 94:1890–95 28. Pitcher CJ, Quittner C, Peterson DM, Connors M, Koup RA, et al. 1999. HIV-1-speciﬁc CD4+ T cells are detectable in most individuals with active HIV-1 infection, but decline with prolonged viral suppression. Nat. Med. 5:518–25 29. Rosenberg ES, Altfeld M, Poon SH, Phillips MN, Wilkes BM, et al. 2000. Immune control of HIV-1 after early treatment of acute infection. Nature 407:523–26 30. Amyes E, Hatton C, Montamat-Sicotte D, Gudgeon N, Rickinson AB, et al. 2003. Characterization of the CD4+ T cell response to Epstein-Barr virus during primary and persistent infection. J. Exp. Med. 198:903–11 31. McMichael AJ, Rowland-Jones SL. 2001. Cellular immune responses to HIV. Nature 410:980–87 32. Shedlock DJ, Shen H. 2003. Requirement for CD4 T cell help in generating functional CD8 T cell memory. Science 300:337–39 33. Sun JC, Bevan MJ. 2003. Defective CD8 T cell memory following acute infection without CD4 T cell help. Science 300:339–42 34. Sun JC, Williams MA, Bevan MJ. 2004. CD4+ T cells are required for the maintenance, not programming, of memory CD8+ T cells after acute infection. Nat. Immunol. 5:927– 33 35. Zajac AJ, Blattman JN, Murali-Krishna K, Sourdive DJ, Suresh M, et al. 1998. Viral immune evasion due to persistence of activated T cells without effector function. J. Exp. Med. 188:2205–13 36. Jin X, Bauer DE, Tuttleton SE, Lewin S, Gettie A, et al. 1999. Dramatic rise in plasma viremia after CD8+ T cell depletion in simian immunodeﬁciency virus-infected macaques. J. Exp. Med. 189:991–98 37. Phillips RE, Rowland-Jones S, Nixon DF, Gotch FM, Edwards JP, et al. 1991. Human immunodeﬁciency virus genetic variation that can escape cytotoxic T cell recognition. Nature 354:453–59 www.annualreviews.org • HIV Vaccines

247

ARI

15 February 2006

18:20

38. Kelleher AD, Long C, Holmes EC, Allen RL, Wilson J, et al. 2001. Clustered mutations in HIV-1 gag are consistently required for escape from HLA-B27-restricted cytotoxic T lymphocyte responses. J. Exp. Med. 193:375–86 39. Goulder PJ, Phillips RE, Colbert RA, McAdam S, Ogg G, et al. 1997. Late escape from an immunodominant cytotoxic T-lymphocyte response associated with progression to AIDS. Nat. Med. 3:212–17 40. Moore CB, John M, James IR, Christiansen FT, Witt CS, Mallal SA. 2002. Evidence of HIV-1 adaptation to HLA-restricted immune responses at a population level. Science 296:1439–43 41. Goulder PJ, Altfeld MA, Rosenberg ES, Nguyen T, Tang Y, et al. 2001. Substantial differences in speciﬁcity of HIV-speciﬁc cytotoxic T cells in acute and chronic HIV infection. J. Exp. Med. 193:181–94 42. Appay V, Dunbar PR, Callan M, Klenerman P, Gillespie GM, et al. 2002. Memory CD8+ T cells vary in differentiation phenotype in different persistent virus infections. Nat. Med. 8:379–85 43. Champagne P, Ogg GS, King AS, Knabenhans C, Ellefsen K, et al. 2001. Skewed maturation of memory HIV-speciﬁc CD8 T lymphocytes. Nature 410:106–11 44. Appay V, Nixon DF, Donahoe SM, Gillespie GM, Dong T, et al. 2000. HIV-speciﬁc CD8+ T cells produce antiviral cytokines but are impaired in cytolytic function. J. Exp. Med. 192:63–75 45. Younes SA, Yassine-Diab B, Dumont AR, Boulassel MR, Grossman Z, et al. 2003. HIV1 viremia prevents the establishment of interleukin 2-producing HIV-speciﬁc memory CD4+ T cells endowed with proliferative capacity. J. Exp. Med. 198:1909–22 46. Derdeyn CA, Decker JM, Bibollet-Ruche F, Mokili JL, Muldoon M, et al. 2004. Envelope-constrained neutralization-sensitive HIV-1 after heterosexual transmission. Science 303:2019–22 47. Wei X, Decker JM, Wang S, Hui H, Kappes JC, et al. 2003. Antibody neutralization and escape by HIV-1. Nature 422:307–12 48. Richman DD, Wrin T, Little SJ, Petropoulos CJ. 2003. Rapid evolution of the neutralizing antibody response to HIV type 1 infection. Proc. Natl. Acad. Sci. USA 100:4144 – 49 49. Friedrich TC, Dodds EJ, Yant LJ, Vojnov L, Rudersdorf R, et al. 2004. Reversion of CTL escape-variant immunodeﬁciency viruses in vivo. Nat. Med. 10:275–81 50. Goulder PJ, Watkins DI. 2004. HIV and SIV CTL escape: implications for vaccine design. Nat. Rev. Immunol. 4:630–40 51. Alabi AS, Jaffar S, Ariyoshi K, Blanchard T, van der Loeff MS, et al. 2003. Plasma viral load, CD4 cell percentage, HLA and survival of HIV-1, HIV-2, and dually infected Gambian patients. AIDS 17:1513–20 52. Van Baalen CA, Schutten M, Huisman RC, Boers PH, Gruters RA, Osterhaus AD. 1998. Kinetics of antiviral activity by human immunodeﬁciency virus type 1-speciﬁc cytotoxic T lymphocytes (CTL) and rapid selection of CTL escape virus in vitro. J. Virol. 72:6851–57 53. Girard M, Mahoney J, Wei Q, van der Ryst E, Muchmore E, et al. 1998. Genital infection of female chimpanzees with human immunodeﬁciency virus type 1. AIDS Res. Hum. Retrovir. 14:1357–67 54. Fowke KR, Nagelkerke NJ, Kimani J, Simonsen JN, Anzala AO, et al. 1996. Resistance to HIV-1 infection among persistently seronegative prostitutes in Nairobi, Kenya. Lancet 348:1347–51

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-08

248

McMichael

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-08

ARI

15 February 2006

18:20

55. Langlade-Demoyen P, Ngo-Giang-Huong N, Ferchal F, Oksenhendler E. 1994. Human immunodeﬁciency virus (HIV) nef-speciﬁc cytotoxic T lymphocytes in noninfected heterosexual contact of HIV-infected patients. J. Clin. Invest. 93:1293–97 56. Rowland-Jones SL, McMichael A. 1995. Immune responses in HIV-exposed seronegatives: Have they repelled the virus? Curr. Opin. Immunol. 7:448–55 57. Rowland-Jones SL, Dong T, Fowke KR, Kimani J, Krausa P, et al. 1998. Cytotoxic T cell responses to multiple conserved HIV epitopes in HIV-resistant prostitutes in Nairobi. J. Clin. Invest. 102:1758–65 58. Goh WC, Markee J, Akridge RE, Meldorf M, Musey L, et al. 1999. Protection against human immunodeﬁciency virus type 1 infection in persons with repeated exposure: evidence for T cell immunity in the absence of inherited CCR5 coreceptor defects. J. Infect. Dis. 179:548–57 59. Clerici M, Levin JM, Kessler HA, Harris A, Berzofsky JA, et al. 1994. HIV-speciﬁc T-helper activity in seronegative health care workers exposed to contaminated blood. JAMA 271:42–46 60. Hladik F, Desbien A, Lang J, Wang L, Ding Y, et al. 2003. Most highly exposed seronegative men lack HIV-1-speciﬁc, IFN-γ-secreting T cells. J. Immunol. 171:2671–83 61. Kaul R, Plummer FA, Kimani J, Dong T, Kiama P, et al. 2000. HIV-1-speciﬁc mucosal CD8+ lymphocyte responses in the cervix of HIV-1-resistant prostitutes in Nairobi. J. Immunol. 164:1602–11 62. Mazzoli S, Trabattoni D, Lo Caputo S, Piconi S, Ble C, et al. 1997. HIV-speciﬁc mucosal and cellular immunity in HIV-seronegative partners of HIV-seropositive individuals. Nat. Med. 3:1250–57 63. Dorrell L, Hessell AJ, Wang M, Whittle H, Sabally S, et al. 2000. Absence of speciﬁc mucosal antibody responses in HIV-exposed uninfected sex workers from the Gambia. AIDS 14:1117–22 64. Kaul R, Rowland-Jones SL, Kimani J, Dong T, Yang HB, et al. 2001. Late seroconversion in HIV-resistant Nairobi prostitutes despite pre-existing HIV-speciﬁc CD8+ responses. J. Clin. Invest. 107:341–49 65. Zhu T, Corey L, Hwangbo Y, Lee JM, Learn GH, et al. 2003. Persistence of extraordinarily low levels of genetically homogeneous human immunodeﬁciency virus type 1 in exposed seronegative individuals. J. Virol. 77:6108–16 66. Mitchell T, Risser R. 1992. Interference established in mice by infection with Friend murine leukemia virus. J. Virol. 66:5696–702 67. Parren PW, Marx PA, Hessell AJ, Luckay A, Harouse J, et al. 2001. Antibody protects macaques against vaginal challenge with a pathogenic R5 simian/human immunodeﬁciency virus at serum levels giving complete neutralization in vitro. J. Virol. 75:8340–47 68. Gauduin MC, Parren PW, Weir R, Barbas CF, Burton DR, Koup RA. 1997. Passive immunization with a human monoclonal antibody protects hu-PBL-SCID mice against challenge by primary isolates of HIV-1. Nat. Med. 3:1389–93 69. Mascola JR, Stiegler G, VanCott TC, Katinger H, Carpenter CB, et al. 2000. Protection of macaques against vaginal transmission of a pathogenic HIV-1/SIV chimeric virus by passive infusion of neutralizing antibodies. Nat. Med. 6:207–10 70. Veazey RS, Shattock RJ, Pope M, Kirijan JC, Jones J, et al. 2003. Prevention of virus transmission to macaque monkeys by a vaginally applied monoclonal antibody to HIV-1 gp120. Nat. Med. 9:343–46 71. Moore JP, Cao Y, Qing L, Sattentau QJ, Pyati J, et al. 1995. Primary isolates of human immunodeﬁciency virus type 1 are relatively resistant to neutralization by monoclonal www.annualreviews.org • HIV Vaccines

249

ANRV270-IY24-08

ARI

15 February 2006

72.

73. 74.

75.

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

76. 77. 78. 79.

80.

81.

82.

83.

83a.

84.

84a.

85.

86.

250

18:20

antibodies to gp120, and their neutralization is not predicted by studies with monomeric gp120. J. Virol. 69:101–9 Pitisuttithum P, Berman PW, Phonrat B, Suntharasamai P, Raktham S, et al. 2004. Phase I/II study of a candidate vaccine designed against the B and E subtypes of HIV-1. J. Acquir. Immune Deﬁc. Syndr. 37:1160–65 Desrosiers RC. 2004. Prospects for an AIDS vaccine. Nat. Med. 10:221–23 Kwong PD, Wyatt R, Robinson J, Sweet RW, Sodroski J, Hendrickson WA. 1998. Structure of an HIV gp120 envelope glycoprotein in complex with the CD4 receptor and a neutralizing human antibody. Nature 393:648–59 Wyatt R, Kwong PD, Desjardins E, Sweet RW, Robinson J, et al. 1998. The antigenic structure of the HIV gp120 envelope glycoprotein. Nature 393:705–11 Chen B, Vogan EM, Gong H, Skehel JJ, Wiley DC, Harrison SC. 2005. Structure of an unliganded simian immunodeﬁciency virus gp120 core. Nature 433:834–41 Kwong PD. 2005. Human immunodeﬁciency virus: refolding the envelope. Nature 433:815–16 Reitter JN, Means RE, Desrosiers RC. 1998. A role for carbohydrates in immune evasion in AIDS. Nat. Med. 4:679–84 Cardoso RM, Zwick MB, Stanﬁeld RL, Kunert R, Binley JM, et al. 2005. Broadly neutralizing anti-HIV antibody 4E10 recognizes a helical conformation of a highly conserved fusion-associated motif in gp41. Immunity 22:163–73 Calarese DA, Scanlan CN, Zwick MB, Deechongkit S, Mimura Y, et al. 2003. Antibody domain exchange is an immunological solution to carbohydrate cluster recognition. Science 300:2065–71 Choe H, Li W, Wright PL, Vasilieva N, Venturi M, et al. 2003. Tyrosine sulfation of human antibodies contributes to recognition of the CCR5 binding region of HIV-1 gp120. Cell 114:161–70 Binley JM, Wrin T, Korber B, Zwick MB, Wang M, et al. 2004. Comprehensive crossclade neutralization analysis of a panel of anti-human immunodeﬁciency virus type 1 monoclonal antibodies. J. Virol. 78:13232–52 Moulard M, Phogat SK, Shu Y, Labrijn AF, Xiao X, et al. 2002. Broadly cross-reactive HIV-1-neutralizing human monoclonal Fab selected for binding to gp120-CD4-CCR5 complexes. Proc. Natl. Acad. Sci. USA 99:6913–18 Kwong PD, Doyle ML, Casper DJ, Cicala C, Leavitt SA, et al. 2002. HIV-1 evades antibody-mediated neutralization through conformational masking of receptor-binding sites. Nature 420:678–82 Scanlan CN, Pantophlet R, Wormald MR, Saphire EO, Stanﬁeld R, et al. 2002. The broadly neutralizing anti-human immunodeﬁciency virus type 1 antibody 2G12 recognizes a cluster of α1 ≥ 2 mannose residues on the outer face of gp120. J. Virol. 76:7306–21 Ofek G, Tang M, Sambor A, Katinger H, Mascola JR, et al. 2004. Structure and mechanistic analysis of the anti-human immunodeﬁciency virus type 1 antibody 2F5 in complex with its gp41 epitope. J. Virol. 78:10724–37 Haynes BF, Fleming J, St. Clair WE, Katinger H, Stiegler G, et al. 2005. Cardiolipin polyspeciﬁc autoreactivity in two broadly neutralizing HIV-1 antibodies. Science 308:1906–8 Vine AM, Heaps AG, Kaftantzi L, Mosley A, Asquith B, et al. 2004. The role of CTLs in persistent viral infection: cytolytic gene expression in CD8+ lymphocytes distinguishes between individuals with a high or low proviral load of human T cell lymphotropic virus type 1. J. Immunol. 173:5121–29

McMichael

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-08

ARI

15 February 2006

18:20

87. Guidotti LG, Borrow P, Brown A, McClary H, Koch R, Chisari FV. 1999. Noncytopathic clearance of lymphocytic choriomeningitis virus from the hepatocyte. J. Exp. Med. 189:1555–64 88. Altfeld M, Allen TM, Yu XG, Johnston MN, Agrawal D, et al. 2002. HIV-1 superinfection despite broad CD8+ T-cell responses containing replication of the primary virus. Nature 420:434–39 89. Mackewicz CE, Blackbourn DJ, Levy JA. 1995. CD8+ T cells suppress human immunodeﬁciency virus replication by inhibiting viral transcription. Proc. Natl. Acad. Sci. USA 92:2308–12 90. Amara RR, Villinger F, Altman JD, Lydy SL, O’Neil SP, et al. 2001. Control of a mucosal challenge and prevention of AIDS by a multiprotein DNA/MVA vaccine. Science 292:69– 74 91. Barouch DH, Santra S, Schmitz JE, Kuroda MJ, Fu TM, et al. 2000. Control of viremia and prevention of clinical AIDS in rhesus monkeys by cytokine-augmented DNA vaccination. Science 290:486–92 92. Shiver JW, Fu TM, Chen L, Casimiro DR, Davies ME, et al. 2002. Replicationincompetent adenoviral vaccine vector elicits effective anti-immunodeﬁciency-virus immunity. Nature 415:331–35 93. Rose NF, Marx PA, Luckay A, Nixon DF, Moretto WJ, et al. 2001. An effective AIDS vaccine based on live attenuated vesicular stomatitis virus recombinants. Cell 106:539–49 94. Chen X, Scala G, Quinto I, Liu W, Chun TW, et al. 2001. Protection of rhesus macaques against disease progression from pathogenic SHIV-89.6PD by vaccination with phagedisplayed HIV-1 epitopes. Nat. Med. 7:1225–31 95. Horton H, Vogel TU, Carter DK, Vielhuber K, Fuller DH, et al. 2002. Immunization of rhesus macaques with a DNA prime/modiﬁed vaccinia virus Ankara boost regimen induces broad simian immunodeﬁciency virus (SIV)-speciﬁc T-cell responses and reduces initial viral replication but does not prevent disease progression following challenge with pathogenic SIVmac239. J. Virol. 76:7187–202 96. Allen TM, Mortara L, Mothe BR, Liebl M, Jing P, et al. 2002. Tat-vaccinated macaques do not control simian immunodeﬁciency virus SIVmac239 replication. J. Virol. 76:4108– 12 97. Pal R, Venzon D, Letvin NL, Santra S, Monteﬁori DC, et al. 2002. ALVAC-SIV-gag-polenv-based vaccination and macaque major histocompatibility complex class I (A∗ 01) delay simian immunodeﬁciency virus SIVmac -induced immunodeﬁciency. J. Virol. 76:292–302 98. Vogel TU, Reynolds MR, Fuller DH, Vielhuber K, Shipley T, et al. 2003. Multispeciﬁc vaccine-induced mucosal cytotoxic T lymphocytes reduce acute-phase viral replication but fail in long-term control of simian immunodeﬁciency virus SIVmac239. J. Virol. 77:13348–60 99. Barouch DH, Kunstman J, Kuroda MJ, Schmitz JE, Santra S, et al. 2002. Eventual AIDS vaccine failure in a rhesus monkey by viral escape from cytotoxic T lymphocytes. Nature 415:335–39 100. Norbury CC, Basta S, Donohue KB, Tscharke DC, Princiotta MF, et al. 2004. CD8+ T cell cross-priming via transfer of proteasome substrates. Science 304:1318–21 101. MacGregor RR, Boyer JD, Ugen KE, Lacy KE, Gluckman SJ, et al. 1998. First human trial of a DNA-based vaccine for treatment of human immunodeﬁciency virus type 1 infection: safety and host response. J. Infect. Dis. 178:92–100 102. Maecker HT, Rinfret A, D’Souza P, Darden J, Roig E, et al. 2005. Standardization of cytokine ﬂow cytometry assays. BMC Immunol. 6:13 www.annualreviews.org • HIV Vaccines

251

ARI

15 February 2006

18:20

103. MacGregor RR, Ginsberg R, Ugen KE, Baine Y, Kang CU, et al. 2002. T-cell responses induced in normal volunteers immunized with a DNA-based vaccine containing HIV-1 env and rev. AIDS 16:2137–43 104. Vuola JM, Keating S, Webster DP, Berthoud T, Dunachie S, et al. 2005. Differential immunogenicity of various heterologous prime-boost vaccine regimens using DNA and viral vectors in healthy volunteers. J. Immunol. 174:449–55 105. Edupuganti S, Weber D, Poole C. 2004. Cytotoxic T-lymphocyte responses to canarypox vector-based HIV vaccines in HIV-seronegative individuals: a meta-analysis of published studies. HIV Clin. Trials 5:259–68 106. McShane H, Pathan AA, Sander CR, Keating SM, Gilbert SC, et al. 2004. Recombinant modiﬁed vaccinia virus Ankara expressing antigen 85A boosts BCG-primed and naturally acquired antimycobacterial immunity in humans. Nat. Med. 10:1240–44 107. Harro C, Edupuganti S, Goepfert P, Priddya F, Lally M, et al. 2005. Safety and immunogenicity of Adenovirus Type 5 (Ad5) HIV-1 gag vaccines. 12th Conf. Retrovir. Oppor. Infect. Abstr. 504. http://www.retroconference.org/2005/ 108. Sumida SM, Truitt DM, Lemckert AA, Vogels R, Custers JH, et al. 2005. Neutralizing antibodies to adenovirus serotype 5 vaccine vectors are directed primarily against the adenovirus hexon protein. J. Immunol. 174:7179–85 109. Trimble C, Lin CT, Hung CF, Pai S, Juang J, et al. 2003. Comparison of the CD8+ T cell responses and antitumor effects generated by DNA vaccine administered through gene gun, biojector, and syringe. Vaccine 21:4036–42 110. Korber B, Muldoon M, Theiler J, Gao F, Gupta R, et al. 2000. Timing the ancestor of the HIV-1 pandemic strains. Science 288:1789–96 111. Leslie AJ, Pfafferott KJ, Chetty P, Draenert R, Addo MM, et al. 2004. HIV evolution: CTL escape mutation and reversion after transmission. Nat. Med. 10:282–89 112. Leslie A, Kavanagh D, Honeyborne I, Pfafferott K, Edwards C, et al. 2005. Transmission and accumulation of CTL escape variants drive negative associations between HIV polymorphisms and HLA. J. Exp. Med. 201:891–902 113. Peyerl FW, Bazick HS, Newberg MH, Barouch DH, Sodroski J, Letvin NL. 2004. Fitness costs limit viral escape from cytotoxic T lymphocytes at a structurally constrained epitope. J. Virol. 78:13901–10 114. Cocchi F, DeVico AL, Garzino-Demo A, Cara A, Gallo RC, Lusso P. 1996. The V3 domain of the HIV-1 gp120 envelope glycoprotein is critical for chemokine-mediated blockade of infection. Nat. Med. 2:1244–47 115. Graham BS. 2002. Clinical trials of HIV vaccines. Annu. Rev. Med. 53:207–21 116. Townsend AR, Rothbard J, Gotch FM, Bahadur G, Wraith D, McMichael AJ. 1986. The epitopes of inﬂuenza nucleoprotein recognized by cytotoxic T lymphocytes can be deﬁned with short synthetic peptides. Cell 44:959–68 117. Lee JK, Stewart-Jones G, Dong T, Harlos K, Di Gleria K, et al. 2004. T cell crossreactivity and conformational changes during TCR engagement. J. Exp. Med. 200:1455– 66 118. Yu XG, Addo MM, Rosenberg ES, Rodriguez WR, Lee PK, et al. 2002. Consistent patterns in the development and immunodominance of human immunodeﬁciency virus type 1 (HIV-1)-speciﬁc CD8+ T-cell responses following acute HIV-1 infection. J. Virol. 76:8690–701 119. Yewdell JW, Del Val M. 2004. Immunodominance in TCD8+ responses to viruses: cell biology, cellular immunology, and mathematical models. Immunity 21:149–53 120. Kedl RM, Kappler JW, Marrack P. 2003. Epitope dominance, competition and T cell afﬁnity maturation. Curr. Opin. Immunol. 15:120–27

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-08

252

McMichael

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-08

ARI

15 February 2006

18:20

121. Klenerman P, Rowland-Jones S, McAdam S, Edwards J, Daenke S, et al. 1994. Cytotoxic T-cell activity antagonized by naturally occurring HIV-1 Gag variants. Nature 369:403–7 122. Mongkolsapaya J, Dejnirattisai W, Xu XN, Vasanawathana S, Tangthawornchaikul N, et al. 2003. Original antigenic sin and apoptosis in the pathogenesis of dengue hemorrhagic fever. Nat. Med. 9:921–27 123. Haeryfar SM, DiPaolo RJ, Tscharke DC, Bennink JR, Yewdell JW. 2005. Regulatory T cells suppress CD8+ T cell responses induced by direct priming and cross-priming and moderate immunodominance disparities. J. Immunol. 174:3344–51 124. Autran B, Carcelain G, Combadiere B, Debre P. 2004. Therapeutic vaccines for chronic infections. Science 305:205–8 125. Dorrell L. 2005. Therapeutic immunisation strategies for the control of HIV-1. Expert Rev. Vaccines 4:513–20 126. Churdboonchart V, Sakondhavat C, Kulpradist S, Ayudthya BIN, Chandeying V, et al. 2000. A double-blind, adjuvant-controlled trial of human immunodeﬁciency virus type 1 (HIV-1) immunogen (Remune) monotherapy in asymptomatic, HIV-1-infected Thai subjects with CD4-cell counts of >300. Clin. Diagn. Lab. Immunol. 7:728–33 127. Birx DL, Loomis-Price LD, Aronson N, Brundage J, Davis C, et al. 2000. Efﬁcacy testing of recombinant human immunodeﬁciency virus (HIV) gp160 as a therapeutic vaccine in early-stage HIV-1-infected volunteers. rgp160 Phase II Vaccine Investigators. J. Infect. Dis. 181:881–89 128. Kundu SK, Katzenstein D, Moses LE, Merigan TC. 1992. Enhancement of human immunodeﬁciency virus (HIV)-speciﬁc CD4+ and CD8+ cytotoxic T-lymphocyte activities in HIV-infected asymptomatic patients given recombinant gp160 vaccine. Proc. Natl. Acad. Sci. USA 89:11204–8 129. Goebel FD, Mannhalter JW, Belshe RB, Eibl MM, Grob PJ, et al. 1999. Recombinant gp160 as a therapeutic vaccine for HIV-infection: results of a large randomized, controlled trial. European Multinational IMMUNO AIDS Vaccine Study Group. AIDS 13:1461–68 130. Veenstra J, Williams IG, Colebunders R, Dorrell L, Tchamouroff SE, et al. 1996. Immunization with recombinant p17/p24:Ty virus-like particles in human immunodeﬁciency virus-infected persons. J. Infect. Dis. 174:862–66 131. Lindenburg CE, Stolte I, Langendam MW, Miedema F, Williams IG, et al. 2002. Longterm follow-up: no effect of therapeutic vaccination with HIV-1 p17/p24:Ty virus-like particles on HIV-1 disease progression. Vaccine 20:2343–47 132. Moss RB, Giermakowska WK, Savary JR, Theofan G, Daigle AE, et al. 1998. A primer on HIV type 1-speciﬁc immune function and REMUNE. AIDS Res. Hum. Retrovir. 14 (Suppl. 2):S167–75 133. Kelleher AD, Roggensack M, Jaramillo AB, Smith DE, Walker A, et al. 1998. Safety and immunogenicity of a candidate therapeutic vaccine, p24 virus-like particle, combined with zidovudine, in asymptomatic subjects. Community HIV Research Network Investigators. AIDS 12:175–82 134. Ogg GS, Jin X, Bonhoeffer S, Moss P, Nowak MA, et al. 1999. Decay kinetics of human immunodeﬁciency virus-speciﬁc effector cytotoxic T lymphocytes after combination antiretroviral therapy. J. Virol. 73:797–800 135. Oxenius A, Price DA, Gunthard HF, Dawson SJ, Fagard C, et al. 2002. Stimulation of HIV-speciﬁc cellular immunity by structured treatment interruption fails to enhance viral control in chronic HIV infection. Proc. Natl. Acad. Sci. USA 99:13747–52 136. Tryniszewska E, Nacsa J, Lewis MG, Silvera P, Monteﬁori D, et al. 2002. Vaccination of macaques with long-standing SIVmac251 infection lowers the viral set point after cessation of antiretroviral therapy. J. Immunol. 169:5347–57 www.annualreviews.org • HIV Vaccines

253

ARI

15 February 2006

18:20

137. Hel Z, Venzon D, Poudyal M, Tsai WP, Giuliani L, et al. 2000. Viremia control following antiretroviral treatment and therapeutic immunization during primary SIV251 infection of macaques. Nat. Med. 6:1140–46 138. Lisziewicz J, Trocio J, Xu J, Whitman L, Ryder A, et al. 2005. Control of viral rebound through therapeutic immunization with DermaVir. AIDS 19:35–43 139. Lu W, Wu X, Lu Y, Guo W, Andrieu JM. 2003. Therapeutic dendritic-cell vaccine for simian AIDS. Nat. Med. 9:27–32 140. Kahn JO, Cherng DW, Mayer K, Murray H, Lagakos S. 2000. Evaluation of HIV1 immunogen, an immunologic modiﬁer, administered to patients infected with HIV having 300 to 549 × 10(6)/L CD4 cell counts: a randomized controlled trial. JAMA 284:2193–202 141. Kinloch S, Perrin L, Hoen B, Lampe F, Phillips A, Goh L, et al. 2004. Evaluation of 2 therapeutic vaccination regimens in HAART-treated primary HIV infection subjects following analytical treatment interruption: QUEST PROB3005, a randomized, placebocontrolled study. 11th Conf. Retrovir. Oppor. Infect. Abstr. 168 142. Dorrell L, Yang H, Iversen AK, Conlon C, Suttill A, et al. 2005. Therapeutic immunization of highly active antiretroviral therapy-treated HIV-1 infected patients: safety and immunogenicity of an HIV-1 gag/polyepitope DNA vaccine. AIDS 19:1321–23 143. Levy Y, Gahery-Segard H, Durier C, Lascaux AS, Goujard C, et al. 2005. Immunological and virological efﬁcacy of a therapeutic immunization combined with interleukin-2 in chronically HIV-1 infected patients. AIDS 19:279–86 144. Kilby JM, Wang R, Mildvan D, Fischl M, Santana-Bagur J, et al. 2005. A phase II randomized partially blinded trial of antiretroviral therapy, HIV speciﬁc immunizations, and IL-2 cycles to promote efﬁcient control of viral replication (ACTG A5024). 12th Conf. Retrovir. Oppor. Infect. Abstr. 525 145. Markowitz M, Jin X, Hurley A, Simon V, Ramratnam B, et al. 2002. Discontinuation of antiretroviral therapy commenced early during the course of human immunodeﬁciency virus type 1 infection, with or without adjunctive vaccination. J. Infect. Dis. 186:634– 43 146. Harrer E, Bauerle M, Ferstl B, Chaplin P, Petzold B, et al. 2005. Therapeutic vaccination of HIV-1-infected patients on HAART with a recombinant HIV-1 nef-expressing MVA: safety, immunogenicity and inﬂuence on viral load during treatment interruption. Antivir. Ther. 10:285–300 147. Kundu SK, Engleman E, Benike C, Shapero MH, Dupuis M, et al. 1998. A pilot clinical trial of HIV antigen-pulsed allogeneic and autologous dendritic cell therapy in HIVinfected patients. AIDS Res. Hum. Retrovir. 14:551–60 148. Lu W, Arraes LC, Ferreira WT, Andrieu JM. 2004. Therapeutic dendritic-cell vaccine for chronic HIV-1 infection. Nat. Med. 10:1359–65 149. Stott J, Almond N. 1995. Assessing animal models of AIDS. Nat. Med. 1:295–97 150. Chan WL, Rodgers A, Grief C, Almond N, Ellis S, et al. 1995. Immunization with class I human histocompatibility leukocyte antigen can protect macaques against challenge infection with SIVmac-32H. AIDS 9:223–28 151. Bogers WM, Bergmeier LA, Ma J, Oostermeijer H, Wang Y, et al. 2004. A novel HIVCCR5 receptor vaccine strategy in the control of mucosal SIV/HIV infection. AIDS 18:25–36 152. Liu R, Paxton WA, Choe S, Ceradini D, Martin SR, et al. 1996. Homozygous defect in HIV-1 coreceptor accounts for resistance of some multiply-exposed individuals to HIV-1 infection. Cell 86:367–77

Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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153. Daniel MD, Kirchhoff F, Czajak SC, Sehgal PK, Desrosiers RC. 1992. Protective effects of a live attenuated SIV vaccine with a deletion in the nef gene. Science 258:1938–41 154. Almond N, Rose J, Sangster R, Silvera P, Stebbings R, et al. 1997. Mechanisms of protection induced by attenuated simian immunodeﬁciency virus. I. Protection cannot be transferred with immune serum. J. Gen. Virol. 78(Pt. 8):1919–22 155. Schmitz JE, Johnson RP, McClure HM, Manson KH, Wyand MS, et al. 2005. Effect of CD8+ lymphocyte depletion on virus containment after simian immunodeﬁciency virus SIVmac251 challenge of live attenuated SIVmac2393-vaccinated rhesus macaques. J. Virol. 79:8131–41 156. Stebbings R, Stott J, Almond N, Hull R, Lines J, et al. 1998. Mechanisms of protection induced by attenuated simian immunodeﬁciency virus. II. Lymphocyte depletion does not abrogate protection. AIDS Res. Hum. Retrovir. 14:1187–98 157. Wyand MS, Manson K, Monteﬁori DC, Lifson JD, Johnson RP, Desrosiers RC. 1999. Protection by live, attenuated simian immunodeﬁciency virus against heterologous challenge. J. Virol. 73:8356–63 158. Stebbings R, Berry N, Stott J, Hull R, Walker B, et al. 2004. Vaccination with live attenuated simian immunodeﬁciency virus for 21 days protects against superinfection. Virology 330:249–60 159. Whatmore AM, Cook N, Hall GA, Sharpe S, Rud EW, Cranage MP. 1995. Repair and evolution of nef in vivo modulates simian immunodeﬁciency virus virulence. J. Virol. 69:5117–23 160. Brenchley JM, Schacker TW, Ruff LE, Price DA, Taylor JH, et al. 2004. CD4+ T cell depletion during all stages of HIV disease occurs predominantly in the gastrointestinal tract. J. Exp. Med. 200:749–59 161. Xu XN, Screaton GR, Gotch FM, Dong T, Tan R, et al. 1997. Evasion of cytotoxic T lymphocyte (CTL) responses by nef-dependent induction of Fas ligand (CD95L) expression on simian immunodeﬁciency virus-infected cells. J. Exp. Med. 186:7–16 162. Baba TW, Liska V, Khimani AH, Ray NB, Dailey PJ, et al. 1999. Live attenuated, multiply deleted simian immunodeﬁciency virus causes AIDS in infant and adult macaques. Nat. Med. 5:194–203 163. Lifson JD, Rossio JL, Piatak M Jr, Parks T, Li L, et al. 2001. Role of CD8+ lymphocytes in control of simian immunodeﬁciency virus infection and resistance to rechallenge after transient early antiretroviral treatment. J. Virol. 75:10187–99 164. Klausner RD, Fauci AS, Corey L, Nabel GJ, Gayle H, et al. 2003. Medicine. The need for a global HIV vaccine enterprise. Science 300:2036–39 165. Saphire EO, Parren PW, Pantophlet R, Zwick MB, Morris GM, et al. 2001. Crystal structure of a neutralizing human IGG against HIV-1: a template for vaccine design. Science 293:1155–59 166. Labrijn AF, Poignard P, Raja A, Zwick MB, Delgado K, et al. 2003. Access of antibody molecules to the conserved coreceptor binding site on glycoprotein gp120 is sterically restricted on primary human immunodeﬁciency virus type 1. J. Virol. 77:10557–65 167. Zwick MB, Komori HK, Stanﬁeld RL, Church S, Wang M, et al. 2004. The long third complementarity-determining region of the heavy chain is important in the activity of the broadly neutralizing anti-human immunodeﬁciency virus type 1 antibody 2F5. J. Virol. 78:3155–61

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:227-255. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo Cytokines and Lymphoid Development Unit, INSERM Unit 668 Institut Pasteur, 75724 Paris, France; email: [email protected]

Annu. Rev. Immunol. 2006. 24:257–86 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090700 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0257$20.00

Key Words commitment, lymphocyte, hematopoietic precursor, transcription factor

Abstract NK cells sit at the crossroads of innate and adaptive immunity and help coordinate tumor immunosurveillance and the immune response against pathogens. Balancing signals to NK cell precursors is crucial for their early development, when transcription factors compete to specify the different lymphocyte subsets. Despite an elaborate schema for NK cell development and differentiation, several major issues remain to be addressed, such as identifying the sites for NK cell maturation and deﬁning the peripheral NK cell niche.

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INTRODUCTION

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NKT cell: natural killer T cell Effector functions: the ability of NK cells to spontaneously kill target cell lines in vitro (natural cytotoxicity), to produce cytokines that prime the immune system and mediate antiviral and cytotoxic effects, and to produce chemokines that attract other hematopoietic cells

In the 1970s, several groups described a spontaneous cytotoxic antitumor activity in the spleens of unmanipulated mice and rats and in the peripheral blood of normal human subjects (1–5). Although initially received by the scientiﬁc community with a healthy dose of skepticism, the initial observations were rapidly conﬁrmed, and the term natural killer (NK) cell (4) durably entered the immunological lexicon. NK cells were distinguished from B and T cells (because they lack B and T cells’ characteristic cell surface antigens) and were initially referred to as null lymphocytes (reviewed in 6). The distinction of NK cells from adaptive lymphocytes was further underscored by the normal development and function of NK cells in mice and humans that were unable to rearrange their antigen receptor genes (7, 8). NK cells therefore belong to the innate arm of the immune system, whose cellular actors also include granulocytes, macrophages, and mast cells. NK cells represent one of the ﬁrst lines of immune defense; this ﬁts well with their putative role in tumor immunosurveillance and with their known roles in combating infection, in graft rejection, and in pregnancy (9–13). This review focuses on murine NK cells, although parallels to human NK cell biology are also made.

NK Cell Surface Markers Several antigens were initially described as NK speciﬁc, including the NK1.1 antigen (NKR-P1C, a C-type lectin) in C57BL/6 mice, the integrin α2 (CD49b) recognized by the DX5 antibody in most mice, the CD56 antigen in humans, and the asialoganglioN-tetraosylceramide (asialo-GM1) in most species. We now know that these cell surface antigens are also expressed by T cells, mast cells, and granulocytes. Moreover, murine NK cells also bear signatures of other hematopoietic cell types and express low levels of the dendritic cell (DC) marker CD11c 258

Di Santo

(14, 15), the B cell marker CD45R (B220) (16, 17), and the B and T cell CD2 antigen (16, 18). Some NK cell markers appear developmentally regulated: The integrin αM (CD11b or Mac-1, expressed by monocytes, macrophages, and DC) and the leukosialin CD43 are more highly expressed by splenic NK cells than bone marrow NK cells (19). Table 1 provides a nonexhaustive list of NK cell receptors expressed by mouse and human NK cells. A comprehensive review of all NK cell receptors is beyond the scope of this review, and interested readers are referred to recent reviews of the subject (20–22). A truly NK-speciﬁc antigen has not been identiﬁed in mice, although natural cytotoxicity receptors (NCRs) appear NK-restricted in humans (23). Murine NK cells should be analyzed using a combination of several antibodies detecting pan-NK cell determinants. Murine NK cells can be unambiguously identiﬁed as CD3− NK1.1+ CD122+ cells because (a) all NK cells express CD122, which is required for IL-15 responsiveness and is essential for NK cell generation and peripheral survival (reviewed in 24); (b) NK1.1 expression is attained early during NK cell maturation (18, 19) and appears stable throughout the lifespan of NK cells in vivo; and (c) CD3 allows exclusion of the only other hematopoietic cells that can potentially coexpress NK1.1 and CD122 [including natural killer T cells (NKT cells)]. In mouse strains that are NK1.1− , NK cells can be detected as CD3− DX5+ CD122+ cells, although some NK1.1+ NK cells that are DX5− will be missed (18, 25, 26). The inclusion of CD122 in combination with DX5 is important because many bone marrow and splenic cells have the DX5+ CD122− phenotype and are not bona ﬁde NK cells (C.A.J. Vosshenrich & J.P. Di Santo, unpublished observations).

NK Cell Functions NK cells are characterized by several important effector functions, including their capacity to spontaneously lyse susceptible

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Table 1

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NK cell receptors in humans (H) and mice (M)

Receptor

Species

Class

Motif/Adaptor

Ligand

CD2 (LFA-2)

H, M

IgSF

Proline-rich domain

CD48, CD58 (LFA-3)

CD11a (LFA-1)

H, M

IgSF

Src family kinases, PI3K

CD54 (ICAM-1), CD102 (ICAM-2)

CD11b (Mac-1)

H, M

IgSF

?

CD54 (ICAM-1)

CD43 (sialoadhesin)

H, M

IgSF

?

?

CD44

H, M

IgSF

?

Hyaluronic acid

CD49b (DX5)

M

IgSF

?

?

CD56 (N-CAM)

H

IgSF

?

?

Lag3

H

IgSF

?

HLA class II

CD16 (FcγRIII)

H, M

IgSF

ITAM/FcγR

Immune complexes

CD25 (IL-2Rα)

H

CytoR

?

IL-2

CD27

H, M

TNFRSF

TRAF

CD70

CD28

H, M

IgSF

YXXM/PI3K

CD80, CD86

CD69

H, M

C-lectin

?

?

CD94/NKG2C, E

H, M

C-lectin

ITAM/DAP12

HLA-E (H), Qa-1b (M)

CD122 (IL-2Rβ)

H, M

CytoR

JAK1, 3/STAT5a, 5b

IL-2, IL-15

CD161

H, M

C-lectin

?

Clr-g (NKR-P1F)

CD226 (DNAM-1)

H

IgSF

?

CD112 (Nectin-2), CD155

CD244 (2B4)

H, M

SLAM

TXYXXV-I/SAP, Fyn

CD48

NKG2D

H, M

C-lectin

YINM/DAP10, PI3K

MICA, B, ULBs (H), Rae1s(H, M), H60 (M)

KIR2S, KIR3S

H

IgSF

ITAM/DAP12

HLA class I

Ly49D, H, P

M

C-lectin

ITAM/DAP12

H-2 class I, MCMV m157 (Ly49H)

NCR (NKp30, 44, 46)

H, M (NKp46)

IgSF

ITAM/FcγR, CD3ζ (H), DAP12

Viral hemagglutinins ( ?)

ILT-1 (Ig-like transcript 1)

H

IgSF

ITAM/FcγR, DAP12

?

IFN-α/βR

H, M

CytoR

JAK1, Tyk2/STAT1, 4

Type I interferons

gp49A

M

C-lectin

ITAM

?

CD85 (ILT-2)

H

IgSF

ITIM/SHP-1

HLA-A, -B, -G

CD94/NKG2A

H, M

C-lectin

ITIM/SHP-1, -2

HLA-E (H), Qa-1b (M)

CD161

M

C-lectin

ITIM/SHP-1, -2

Clr-b (NKR-P1D)

CD244 (2B4)

H, M

SLAM

TXYXXV-I/SAP, Fyn

CD48

KIR2DL, KIR3DL

H

IgSF

ITIM/SHP-1, -2

HLA class I

Ly49 A-C, E-G, I-O

M

C-lectin

ITIM/SHP-1, -2

H-2 class I

KLRG1

M

C-lectin

ITIM/SHP-1, -2

?

TGF-βR

H, M

CytoR

Smad2

TGF-β family

IL-10R

?

CytoR

JAK2, Tyk2/STAT3

IL-10

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Adhesion receptor

Activating receptor

Inhibitory Receptor

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Hematopoietic stem cell (HSC): deﬁned functionally as the one cell capable of self-renewing and generating all cell types in the blood Missing-self hypothesis: Klas K¨arre predicted the existence of inhibitory receptors on NK cells that would recognize self MHC molecules. The lack of the relevant MHC on target cells (missing-self) would consequently activate NK cells and result in target cell elimination KIR: killer cell immunoglobulinlike receptor SHP: SH2-containing protein-tyrosine phosphatase ITIM: immunoreceptor tyrosine-based inhibitory motifs NK cell repertoire: the collection of cell surface–expressed receptors on a population of NK cells. Individual NK cells can express combinations of activating and inhibitory NK cell receptors

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targets. Natural killing involves exocytosis of perforin- and granzyme-containing cytoplasmic granules via a metabolically active process (reviewed in 27). However, NK cells are equipped with a variety of destructive arms and can also eliminate target cells via FasLand TRAIL-mediated pathways, sometimes in a developmentally related fashion (25, 28). An equally important function of NK cells involves their capacity to promptly produce cytokines [notably IFN-γ, TNF-α, and in some cases GM-CSF or IL-10 (29)] and chemokines [including MIP-1α and -β and RANTES (reviewed in 30, 31)]. Together, these functional activities place NK cells in a position to actively eliminate susceptible targets through multiple, nonredundant mechanisms and to recruit and amplify the inﬂammatory response. NK cell cytokine and chemokine production, as well as their lytic capacity, is stimulated by soluble factors (IL-12, IL-18, IFN-α/β) and is acquired during NK cell maturation. As is discussed in detail below, the mechanisms that control the acquisition of NK cell effector functions are being unraveled and require the expression of several transcription factors (TFs) in developing NK cells and interactions with soluble and membrane-bound factors present in the microenvironment. An outstanding question in NK cell biology involves the diversiﬁcation of NK cells. Do functionally distinct subsets of NK cells exist ? If so, how do they develop and what is their biological relevance? Human NK cells comprise several subsets that are phenotypically and functionally distinct (reviewed in 29). The CD56hi CD16− NK subset appears poised for cytokine secretion and, with a CCR7+ CD62L+ phenotype, can selectively trafﬁc to the lymph nodes (LNs) (32). In contrast, the CD56lo CD16hi subset has more potent cytotoxic activity and may be recruited into inﬂamed tissues (31). Until now, clear evidence for functional diversiﬁcation of NK cells has not been obtained in rodents or in other species. A developmental shift in cytokine production proﬁles has been Di Santo

observed for murine NKT cells in the thymus (33) and proposed for human NK cells that have been derived from hematopoietic stem cell (HSC) in vitro (34). The possibility that NK cells could modify their functional attributes depending on their environmental context could represent another form of NK cell functional plasticity.

NK Cell Receptors Several classes of receptors are employed in the regulation of NK cell effector functions. K¨arre’s initial observations that NK cells could lyse major histocompatibility complex (MHC)-negative targets (35) led to the formulation of the missing-self hypothesis (36) that predicted the existence of NK cell negative regulatory receptors that would interact with MHC ligands and thereby spare target cell destruction by NK cells. Inhibitory receptors (IR) for MHC were subsequently identiﬁed, cloned, and characterized (reviewed in 37, 38), thereby largely fulﬁlling K¨arre’s hypothesis. Multiple classes and alleles of IR for MHC exist in vertebrates [including the most Ly49 receptors in mice, the killer cell immunoglobulin-like receptor (KIR) L family in human, and the CD94/NKG2A complex in both species (reviewed in 37, 39, 40)]. Upon interaction with MHC ligands, these IR recruit SHP-1 and SHP-2 tyrosine phosphatases to immunoreceptor tyrosine-based inhibitory motifs (ITIM) in the cytoplasmic domain of these receptors, thereby blunting activation signals (reviewed in 20, 37, 39). IR for MHC are expressed as NK cell repertoires, with each being expressed by a subset of NK cells and most NK cells expressing at least one IR (reviewed in 39, 40), although there are exceptions (41). NK cells become functionally competent following interactions of IR with self-MHC molecules (41a). Non-MHCbinding IR also exist (Table 1) that broaden the deﬁnition of self (reviewed in 41b). Establishing a diverse IR repertoire on individual NK cells allows the global NK cell pool to carefully monitor self (and foreign) MHC

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molecules and to react when pathological processes perturb MHC expression. IR do not control NK cell reactivity in isolation, but rather work in concert with diverse activating receptors (AR) (reviewed in 22). In recent years, a plethora of NK cell AR have been identiﬁed; these include a subset of Ly49 receptors in mice, the KIR S family in humans, and the CD94/NKG2C complex and the NKG2D receptor in both species (Table 1; reviewed in 22, 42–44). Signal transduction pathways used by AR couple through DAP10, DAP12, and SAP/Fyn adaptors (reviewed in 20–22, 45). Interestingly, diverse functional activities of NK cells appear hardwired through distinct AR-coupled adaptors (46–49). The concept that NK cell reactivity results from a delicate balance between activating and inhibitory signals is now well accepted (reviewed in 20, 22, 37). Moreover, NK cell triggering is not stereotyped, but can be due to imbalance of activating and inhibitory pathways (including not only MHC receptors, but also cytokines and costimulatory molecules, leading to missing, altered, stressed, or non-self situations; see Reference 50, for example). The mechanisms that control NK cell functional activity have provided new paradigms for understanding other types of self-reactive hematopoietic cells, including NKT cells, γδ T cells, mast cells, and B-1 B cells (51, 52).

NK Cells in Innate and Adaptive Immunity It was initially proposed that NK cells participate in tumor immunosurveillance, and there is evidence that NK cells (in concert with NKT cells) are essential actors that prevent tumor initiation and shape tumor evolution through their capacity to kill and secrete cytokines (reviewed in 53). Interestingly, although the lytic capacity of hematopoietic cells against NK-sensitive targets appears maintained in evolution, most signature features of modern NK cells (activating/ inhibitory MHC receptors and granule-

mediated lysis) are not (54). Recognition systems and effector molecules used to achieve target cell lysis vary depending on the species studied. Thus, NK cells, like their adaptive lymphocyte counterparts, are continually evolving. NK cells are also essential for protection against several types of viral infection, in particular herpesvirus in humans (55) and cytomegalovirus (CMV) in mice (56, 57). The role for NK cells in the antiviral response includes both lytic activity (destruction of virally infected cells) and cytokine secretion (IFN-γ as antiviral and chemokines as proinﬂammatory mediators), which may be dispensed in an organ-speciﬁc fashion (reviewed in 58). Nevertheless, NK cells alone are no match for a mutating virus (mutants emerge that can escape NK cell recognition), which will eventually overwhelm an immune system that lacks an adaptive response (59). Cytokine secretion by activated NK cells has been recently shown to reinforce Th1 differentiation of T cells in secondary lymphoid organs (14, 32, 60). These results demonstrate that NK cells can provide a link between innate and adaptive immunity and can thereby contribute to a concerted and effective immune response. Nevertheless, the cell:cell interactions that mediate the pro-Th1 effect of NK cells are undeﬁned. Knowing the precise physical localization of NK cells, DC, and T cells and their migration patterns before and after immunization could provide clues to the kinetics and directionality of the NK cell effect in the LN.

A MODEL FOR MURINE NK CELL DEVELOPMENT Technological advances have played a decisive role in allowing us to propose and re-formulate models for the NK cell differentiation process. Some of the most important developments have included (a) liquidand stroma-based culture systems for the generation of differentiated hematopoietic cells in vitro; (b) multiparameter cell surface immunophenotyping and puriﬁcation of www.annualreviews.org • NK Cell Development

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HSC

NKP

STAGE

A

iNK

B

mNK

C

D

E

F

CD122 NKG2D

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NK1.1 CD94 TRAIL CD51(α α v) Ly49 CD49b (DX5, α 2) CD11b (Mac-1) CD43

Cytotoxicity IFN-γγ

Figure 1 Phenotypic markers of developing NK cells in the mouse. NK precursors (NKP, stage A) are characterized by CD122 and NKG2D expression, but they lack other NK cell markers. Immature NK cells (iNK) upregulate NK1.1 and CD94 and transiently express TRAIL and CD51 (stages B and C). Ly49 receptor repertoires and DX5 are relatively late markers of NK cell differentiation expressed by mature NK cells (mNK, stages D and E). CD11b and CD43 expression increase as NK cells differentiate, although effector functions do not depend on upregulation of these markers.

hematopoietic precursors; (c) gene-modiﬁed mouse models for assessing the roles of a given gene for hematopoietic development in vivo; and (d ) sensitive in vitro and in vivo assays of NK cell function. The pioneer research groups placed NK cells on the map of hematopoiesis by showing that they were transplantable cells from the bone marrow (61, 62) and required growth factors, cytokines, and stromal cell interactions to mature to phenotypically recognizable NK cells 262

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with lytic capacity (17, 63–67). With the generation of additional antibodies against NK cell receptors, the map of NK cell development has been reﬁned and redeﬁned (Figure 1), reﬂecting a constant evolution from the previous models (17, 19, 24, 68). The scheme represented in Figure 1 is a composite map of immature and mature NK cells that have been identiﬁed and characterized in different organs (bone marrow, liver, spleen) at different ages in the mouse. It should

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be emphasized that clear precursor-product relationships have been established between adjacent stages in some, but not all, cases (18, 19, 25, 69). NK cell development can be operationally divided into different stages (Figure 1), although this is obviously arbitrary, as the process of generating mature NK cells from HSC is a continuous process. Nevertheless, the major events in the life of the NK cell considered in this review include (a) commitment of HSC to the NK cell lineage (stage A), (b) phenotypic and functional NK cell maturation (stages B–E), and (c) peripheral NK cell homeostasis (stage F). For each of these steps, I summarize what is known about the differentiation process and highlight the recent key ﬁndings that have shed light on the governing molecular and cellular mechanisms.

NK CELL COMMITMENT—TAKING THE FIRST STEP The ﬁrst step down the road to becoming a mature NK cell involves making a commitment to becoming a natural killer. This point of no return means that other developmental options within the hematopoietic system are no longer available. Hematopoietic precursors having this proﬁle are deﬁned as NK cell precursors (NKP) and result from a sequential loss of pluripotency as HSC differentiate to more committed hematopoietic precursors. Several hematopoietic developmental intermediates have been identiﬁed in the mouse that retain lymphoid (including NK) potential, but have essentially lost the capacity to produce myeloid or erythroid lineages. These lymphoid progenitors include early lymphoid progenitors [ELP; deﬁned as lineage (Lin)negative = CD3− CD19− Ter119− Gr-1− and expressing c-kit and fms-like tyrosine kinase 3 (Flt3) (70)] and common lymphoid progenitors [CLP; deﬁned as Lin− ckit+ IL-7Rα+ (71)]. ELP and CLP are characterized by their ability to give rise to B, T, and NK

cells in culture or upon transfer to recipient mice. Although not solely committed to the NK cell lineage, the analysis of these progenitors has provided important clues about the transcriptional program that is active during lymphoid speciﬁcation (72). Human CLP have a CD34+ CD10+ phenotype [73; see also Blom & Spits (73a) in this volume]. Although these different hematopoietic intermediates have NK potential in vitro and in vivo (70, 71, 73), it is not clear whether developing NK cells must transit through these intermediates in order to mature. For example, bone marrow NKP and peripheral NK cell numbers are normal in c-kit-deﬁcient Vickid mice that completely lack CLP (74, 75). A bipotent T/NK progenitor (TNKP) has been clearly identiﬁed in the murine fetal liver and in the fetal thymus in mice and humans (66, 76, 77). TNKP in the thymus has the Lin− c-kit+ NK1.1+ phenotype in mice and the CD34+ CD7+ CD1a− phenotype in humans (reviewed in 78, 79). In the fetal thymus, TNKP cohabitate with more restricted NKP and T cell progenitors and are probably their immediate precursors. With the development of novel stromal cell–based culture systems expressing Notch ligands (80), the laboratories of Zuniger-Pﬂucker and Petrie have reassessed the T and NK potential of early murine thymocyte subsets (81, 82). The earliest thymocyte precursors (CD44+ CD25− ) harbor both T and NK potential. Interestingly, NK potential also exists (at low frequency) at the CD44+ CD25+ stage (82), when early thymocyte progenitors presumably have received ample Notch signals and already have clear molecular markers of T cell commitment (83). Although these observations could simply represent heterogeneity in the CD44+ CD25+ subset, an alternative interpretation is that NK potential remains possible even following Notch signaling of intrathymic precursors. Notch1 signaling is essential for T cell development in vivo (84), and mice with an inducible deletion in Notch1 lack the earliest T cell committed www.annualreviews.org • NK Cell Development

Homeostasis: a self-regulating process by which biological systems tend to maintain stability while adjusting to conditions that are optimal for survival. When applied to lymphocyte subsets, homeostasis refers to the mechanisms that regulate cell numbers in the peripheral pool Commitment: lineage commitment is a progressive process that results in the loss of the ability of pluripotent precursors to differentiate into multiple lineages NK cell precursors (NKP): deﬁned functionally as cells capable of differentiating into NK cells but not other hematopoietic lineages Developmental intermediates: phenotypically deﬁned cell types known to be part of a developmental pathway; in the hematopoietic system, CLP, ELP, and NKP are more committed lymphoid progenitors derived from HSC that may be intermediates in NK cell development TNKP: bipotent T/NK precursor

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precursors (A. Wilson, personal communication). Nevertheless, thymic NK cells are present in normal numbers under these conditions, suggesting that thymic NK cells can develop in the absence of TNKP. It seems reasonable to conclude that the thymic bipotent TNKP precursor is mainly involved with T cell development, and its NK potential may be elicited under certain conditions when T cell development fails or is inefﬁcient. Whether NK cells that develop outside the thymus transit via TNKP in the bone marrow, liver, or LN is unknown.

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Natural Killer Cell Precursors (NKP) NKP have been identiﬁed and characterized in the fetal thymus and the bone marrow of adult mice with the following phenotype: Lin− CD122+ NK1.1− DX5− (18, 85). CD122 acquisition on Lin− hematopoietic precursors appears to correlate with their commitment to the NK lineage since murine NKP has no B, T, myeloid, or erythroid cell potential (18). CD122 expression is critical for allowing NKP to respond to IL-15 stimulation, and NKP likely represent the cellular substrate on which IL-15 acts to promote NK cell development in the bone marrow (17, 86, 87). Interestingly, the generation of NKP from HSC does not require IL-15 [or any common γ chain (γc )-dependent cytokine (87)], thereby deﬁning this ﬁrst stage of NK cell development as IL-15 independent. Upon stimulation by IL-15, NKP rapidly proliferate and differentiate into phenotypically mature NK cells capable of cytotoxicity and inducible cytokine synthesis (18). In mice, NKP are also found in the thymus, spleen, and LN (18, 85; C.A.J. Vosshenrich & J.P. Di Santo, unpublished observations), where they represent a relatively minor lymphocyte subset. Phenotypically characterized NKP are not a homogeneous population, yet they are enriched in NK potential [one in eight NKP give rise to NK cells in vitro (18)]. Freshly isolated NKP are not cytotoxic for YAC1 targets. Nevertheless, 10% 264

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of NKP express NKG2D at the cell surface (87), suggesting that true NKP may have the Lin− CD122+ NKG2D+ NK1.1− DX5− phenotype. Whether NKG2D+ NKP have lytic potential remains to be tested. NKP in humans have not been rigorously characterized, but they are assumed to play an analogous role. Attempts have been made to identify human NKP after stimulation of hematopoietic precursors with IL-2 or IL-15 in liquid culture alone or in combination with stromal cells (64, 65, 88, 89). Still, this approach reveals not only NKP but also immature and mature NK cells. Human NKP should, like their murine counterparts, express CD122 but not other lineagespeciﬁc markers. Nevertheless, CD122 expression by human lymphoid progenitors (CD34+ CD38+ CD10+ ) has been difﬁcult to demonstrate, despite the fact that these precursors have NK cell potential and are highly responsive to IL-15 (73, 89). A recent report from the Caligiuri laboratory identiﬁed what may represent one type of human NKP (88). A novel CD34dim CD45RA+ integrin α4 β7 hi hematopoietic precursor was shown to differentiate on stromal cells in the presence of IL-2 or IL-15 into mature CD56hi NK cells with lytic capacity and cytokine production. These precursors were present in the bone marrow and peripheral blood and were highly enriched in LNs. LN homing is driven by CCR7 ligands and requires CD62L expression to interact with high endothelial venules. The α4 β7 hi NK precursors were CCR7+ CD62L+ , leading the authors to propose that these cells migrate from the bone marrow via the blood to reach the LN, where they are local precursors of LN-resident CD56hi NK cells (88). The α4 β7 hi NK precursors bear similarities to murine and human lymphoid tissue inducer cells that also have DC and NK cell potential (90, 91). It will be interesting to know whether the α4 β7 hi precursors identiﬁed by Freud and colleagues (88) are functional lymphoid inducer cells with DC potential.

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Generating NKP: Soluble and Membrane Factors The signals that regulate the HSC to NKP transition are poorly understood but involve cell:cell interactions that license NKP to respond to soluble factors elaborated in the microenvironment. The latter quality depends on NKP expression of several distinct TFs. NKP generation likely involves interactions between HSC and stromal cells in vivo because highly puriﬁed human or murine HSC generate few NK cells in liquid culture (even with high levels of IL-15), whereas NK cell development is greatly enhanced when HSC are cultured on relevant stromal cell lines (63– 65, 89, 92, 93). In this way, the process of NK commitment bears similarities to early B cell commitment, for which the earliest stages have also been described as stromal cell dependent (94). The molecular signals delivered by stromal cells that induce NK commitment are unknown but must include those that control CD122 expression. Cytokine signals play a general role in lymphoid commitment, including those delivered via Flt3, c-kit, or γc -dependent receptors. These cytokines could inﬂuence NK commitment because NKP express these receptors (87), and culture of mouse or human HSC with Flt3L or stem cell factor was reported to induce CD122 responsiveness (89, 93). Nevertheless, absolute numbers of NKP are unaffected in c-kit mutants (75) and in γc -deﬁcient mice (87) and are only slightly reduced in the absence of Flt3 (C.A.J. Vosshenrich & J.P. Di Santo, unpublished observations). Individually, these pathways are therefore not essential for the ﬁrst step toward the NK lineage in vivo. This process may have some level of redundancy that could be revealed by studying the appropriate double-mutant mice. Interactions between lymphotoxin (LT) α1 β2 -expressing hematopoietic cells and LTβ receptor (LTβR)-expressing stromal cells play an important role in the generation of NK and NKT cells and in the formation of secondary lymphoid organ architecture (re-

viewed in 95). Mice deﬁcient in LTα or LTβR have slightly reduced absolute numbers of peripheral NK cells (96–98), which has been correlated with a decreased capacity of mutant bone marrow cells to generate NK cells upon culture in IL-15 (99). Whether this defect in NK cell production is accompanied by a decrease in NKP in LTα or LTβRdeﬁcient mice has not been analyzed. An intrinsic abnormality of the stromal cell compartment (which fails to support normal NK cell generation after transfer of HSC from wild-type mice) has been demonstrated (99). LTβR signals in thymic stromal cells result in a NIK (NF-κB-inducing kinase)-mediated activation of RelB, which is required for normal NKT cell development (100). A similar mechanism may be required for normal stromal cell function, a deﬁciency of which underlies the defective NK cell development in the absence of LTα/LTβR. Vitamin D3 upregulated protein 1 (VDUP-1) is a stress-response gene that was initially identiﬁed in vitamin D3–treated HL-60 leukemic cells (101). Recently, VDUP-1-deﬁcient mice were characterized and demonstrated a selective defect in NK and NKT cells (102). Bone marrow, spleen, and LN NK cells were substantially reduced in the absence of VDUP-1, which could be traced to a defect in CD122 expression. NKP could not be detected in the bone marrow of VDUP-1-mutant mice, although some mature NK cells were found in the spleen. Interestingly, vitamin D3 increased the yield of NK cells from IL-15-supplemented stromal cell/HSC cultures, which was correlated with increased CD122 expression. These results suggest a model in which regulation of VDUP-1 could be a determinant of NK cell commitment, through control of CD122 expression and IL-15 sensitivity.

Generating NKP: Transcription Factors TFs play determinant roles in hematopoietic lineage speciﬁcation, and distinct TFs have www.annualreviews.org • NK Cell Development

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been implicated in early lymphocyte development, including the Ets family members PU.1 and Ets-1 and members of the Ikaros zinc-ﬁnger family, including Ikaros, Helios, and Aiolos (reviewed in 103, 104). Deﬁciency in PU.1 affects all lymphocyte lineages [as well as certain myeloid cells (105)]. The lymphoid defect in PU.1-deﬁcient mice can be traced to deﬁciencies in expression of the IL7Rα chain and of the Flt3 receptor (106, 107), which are crucial for B and T cell development (reviewed in 108) but less so for NK cells (109). Transfer of PU.1-deﬁcient HSC generated markedly reduced numbers of NKP (perhaps secondary to reduced numbers of lymphoid progenitors) and a correspondingly diminished peripheral NK cell pool, although the phenotype and cytotoxic activity of residual PU.1-deﬁcient NK cells was largely conserved (110). These results suggest that PU.1 acts early in the NK cell lineage but may be dispensable once NK cells have fully matured, a notion supported by analysis of PU.1-GFPreporter mice (111). Ets-1 is critical for NK cell development (112). In its absence, NK cells are largely absent in the bone marrow, spleen, and LNs, with subsequent defects in NK cell cytotoxicity and cytokine production. Because IL-15 cannot rescue the NK cell defect in Ets-1-deﬁcient bone marrow cultures (112), NKP are likely severely reduced or absent in this setting. Concerning Ikaros, Helios, and Aiolos, so far only a deﬁciency in Ikaros appears to have a negative effect on NK cell development, which may be related to a diminished expression of Flt3 and CD122 (113). Whether Ikaros is required for the generation of NKP or at later stages of NK cell differentiation has not been reported. The inhibitors of DNA binding (Id proteins) are a family of four helix-loop-helix (HLH) proteins that can heterodimerize with the basic HLH E-box TF, including E2A (comprising E12 and E47), E2-2, and HEB, and thereby inhibit their transcriptional activity (reviewed in 114). Relative E-box and Id activities in developing lymphocyte precursors appear to determine cell fate decisions:

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E2A-mutant mice lack B cells (115), and overexpression of Id2 or Id3 can inhibit B and T cell development from HSC (116), whereas Id2-mutant mice lack NK cells (117). The mechanism by which E-box/Id TF ratios control NK cell development is unclear. Thymic NKP appear to be severely reduced in Id2deﬁcient mice (118), suggesting that an excess of E-box transcriptional activity inhibits NK cell generation at least at the level of the NKP. Because E-box TF can interact with histoneacetyltransferase (HAT) complexes (119), the choice to become a B cell (or a T cell) may involve an active chromatin remodeling event with transcriptional activation through E-box interactions with HAT. In this case, NK cells would be generated when E-box TF levels are insufﬁcient to drive early lymphocyte precursors to the B or T cell lineage (Figure 2). This hypothesis is consistent with the lack of appreciable NK cell phenotype in mice deﬁcient in E2A (120) and with the reduced expression of E-box transcripts in thymic NK cells (121). The process of generating NKP from HSC appears tightly controlled and results from a careful balance of extracellular signals derived from stromal cells that cooperate with optimal levels of critical TFs. The identiﬁcation and characterization of murine and putative human NKP provide a means to understand more fully this key step in the process of NK cell development.

FROM NKP TO MATURE NK CELL—ARE WE THERE YET? Once committed to the NK cell lineage, NKP must acquire the phenotypic and functional qualities that characterize mature peripheral NK cells. The already perplexing process of NK cell maturation has come to seem more complex in recent years as investigators have identiﬁed several new markers that subdivide differentiating NK cells (Figure 1) and characterized several mutant mice that manifest partial defects in NK cell differentiation (Table 2). How do we deﬁne an immature NK cell (iNK)? The broadest deﬁnition

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E2A

E2A

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B

pro-T

T

Notch, HEB

ELP

Ikaros

HSC

CLP

pro-B

PU.1 Id2

Id2

NKP

NKP

Id2

NKP

NK Figure 2 A balance of E-box and Id proteins determines NK cell fate. HSC commit to the lymphocyte lineage and generate early lymphoid progenitors (ELP) and common lymphoid progenitors (CLP) under the inﬂuence of the TFs Ikaros, PU.1, and E2A. B and T cell development is associated with increased expression of E-box TFs (E2A, HEB), whereas NK cell development requires Id2, which counterbalances E-box TF activity. Notch activation subsequently promotes T cell development at the expense of B cells.

includes any NK cell that lacks the complete phenotypic and functional attributes of mature peripheral NK cells (having uniform expression of CD122, NK1.1, DX5, NKG2D, and CD16, with variable expression of Ly49 receptors and CD94/NKG2A/C and high expression of CD11b and CD43; stage F). Using these criteria, iNK cells have been identiﬁed in the liver that express NK1.1 but not DX5, Ly49 receptors, or CD11b (25). Conspicuously, liver iNK cells express TRAIL on their cell surface and can use this effector molecule to lyse susceptible targets. These results in the mouse recall earlier studies on human NK cells differentiated in vitro, in which CD161+ CD56− iNK cells were characterized that could kill targets in a TRAIL-dependent, perforin-independent fashion (67). TRAIL+ iNK cells could further develop into TRAIL−

progeny that express Ly49s and DX5 and produce cytokines. Whether TRAIL+ iNK cells in the liver develop in situ or derive from bone marrow NKP is not known. One possibility is that the liver represents an important site for iNK cells to ﬁnish their maturation, a process that could depend on transcription factors that specify hepatic-homing potential (109). Kim and colleagues (19) identiﬁed several iNK cells in the bone marrow and liver. Some of these iNK cells lack Ly49 receptors and DX5 and co-express CD51 (corresponding to Figure 1, stages B and C). Others have a more mature NK cell phenotype (DX5+ Ly49+ ) but are CD11blo CD43lo (Figure 1, stages D and E). Although these cells show lower cytotoxicity and cytokine production compared with their CD11bhi CD43hi counterparts, they nevertheless do exert NK effector function, so www.annualreviews.org • NK Cell Development

iNK cell: immature NK cell

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Table 2

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Genetic mutations affecting the maturation of NK cells

Genes

Phenotype

Repertoire

Stage

Cytotoxicity

Cytokines

Reference

IL-15

CD11blo , CD43lo

Normal

E

Reduced

Normal

87

LTα

?

Dec Ly49A, C/1, D, G2

?

Reduced

?

97

LTβR

?

Normal

?

?

?

98

VDUP-1

?

Absent

?

Reduced

?

102

Syk/ZAP70

Normal

Normal

F

Normal (except ITAM)

Normal (except ITAM)

16

DAP12

Normal

Normal

F

Normal (except Ly49D)

?

138

DAP10

Normal

Normal

F

Reduced (NKG2D)

?

136

SAP/Fyn

Normal

Normal

F

Normal (except 2B4)

Normal (except 2B4)

135

Tyro 3 family

CD11blo , CD43lo

Dec Ly49D

C

Reduced

Reduced

143

PLCγ2

Normal

Red Ly49C/I, G2

E

Absent

Normal

48, 141b

Vav family

Normal

Normal

F

Reduced

Reduced

47

Soluble Factors

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Signaling

Transcription Factors PU.1

Normal

Dec Ly49A, D

A

Normal

Normal

110

Ets-1

?

?

A

Reduced

Reduced

112

Id2

?

?

A

Reduced

Reduced

117

Gata-3

CD11blo , CD43lo

Dec Ly49C/I, D,

C

Normal

Reduced

109

IRF-2

CD11blo , CD43lo

Dec Ly49D, C/I, G2

D

Normal

Reduced

129

T-bet

CD11blo , CD43lo

Normal

E

Normal

Normal

145

MEF, MITF, CEBP-γ

Normal

Normal

F

Reduced

Normal

24, 68

they could simply represent a subset of mature NK cells with a different functional capacity. Care should be taken when applying the label immature to NK cells based solely on lower CD11b and CD43 expression levels. One way to address the functional consequences of NK cell heterogeneity is to isolate and study the cells that have nonuniform expression. For NK cells, this includes cells bearing the CD11b, CD43, CD45R, CD69, CD90, and c-kit markers. Although this can be achieved (with some difﬁculty), an alternative solution involves culture systems that expand and maintain iNK cells. Two recent reports suggest that this may be possible for human and mouse NK cell precursors (122, 268

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123). Both rely on the use of IL-7 to maintain cell viability and generate poorly lytic NK cells that robustly respond to IL-15 exposure with additional proliferation, differentiation, and cytotoxic potential. Using these new culture systems, questions relating to the roles of TFs, soluble and membrane factors, and precursor-product relationships between different subsets of developing NK cells may be answered.

NK Maturation: Soluble and Membrane Factors IL-15 is the essential fuel for NK cell development and for maintenance of NK cells in

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the periphery (124–126). IL-15 is presented by the IL-15Rα subunit (127, and see below) and is produced by hematopoietic and nonhematopoietic stromal cells in an IFN regulatory factor (IRF)-1-dependent fashion (128). The analysis of the few residual NK cells in IL-15-deﬁcient mice revealed that some degree of NK maturation could proceed in the absence of this cytokine (87, 97), although the published results were not always in complete agreement. Vosshenrich et al. (87) found that NK cells in Rag2-deﬁcient x IL-15-deﬁcient mice express Ly49A, D, G2, and CD94 at near normal levels and have lytic- and cytokine-producing potential, despite a CD11blo CD43lo phenotype, a result conﬁrmed independently (129). In contrast, the Kumar laboratory (97, 130) found abnormalities in Ly49 receptor expression in IL-15deﬁcient mice. Exogenous IL-15 administration can correct the defect in NK cell numbers (86) and the Ly49 receptor repertoire in IL-15-deﬁcient mice (97), implying that IL-15 might control Ly49 receptor acquisition. Still, it is not clear at what stage IL-15 is acting under these conditions (NKP, iNK, etc.) or whether IL-15 is acting directly or indirectly to inﬂuence the overall numbers of Ly49+ NK cells. The mechanism by which IL-15 affects NK cells (via survival, expansion, or differentiation) remains enigmatic and may be different for NK cells and their precursors. A report from the Raulet laboratory (131) demonstrated that mature NK cells could expand normally in irradiated IL-15-deﬁcient hosts and suggested that IL-15 was not a major proliferative factor for NK cells. Whether this holds true for iNK cells is not known. The residual iNK cells in IL-15-deﬁcient mice have normal Bcl-2 levels and mitochondrial membrane potential (87) and therefore show no overt signs of ongoing apoptosis; this contrasts with mature NK cells that lose Bcl-2 expression when transferred to an IL15-deprived environment (126, 132). Thus, IL-15 responses of immature and mature NK cells are clearly different.

Beyond the role for LTα/LTβR in the generation of NKP (described above), conﬂicting results exist concerning the role for LTα/LTβR in the formation of the NK cell Ly49 receptor repertoire (97, 98). Lian and colleagues (97) studied LTα-deﬁcient mice and found alterations in Ly49 expression (two- to threefold decreases in percentages of Ly49A-, C-, D-, and G-expressing NK cells). Stevenaert and colleagues (98) studied LTβR-deﬁcient mice and found no evidence for perturbations in the Ly49 repertoire. A trivial explanation for these differences involves technical approaches (the use of DX5 versus NK1.1 to study NK cells, and the absence of co-staining with CD122). Stevenaert raises the alternative explanation that another LTβR ligand—LIGHT—may play a negative regulatory role in this process. Further studies will be required to decipher the relative roles of LIGHT versus LTα signals not only in generating the NK cell Ly49 receptor repertoire, but also in understanding how this pathway promotes the full phenotypic and functional maturation of NK cells (133). Soluble factors also drive the maturation of human NK cells. The relationship between the two well-deﬁned human NK cell subsets (CD56hi CD16− , CD56dim CD16+ ) remains enigmatic. A recent report from Ferlazzo and colleagues (134) demonstrated that IL-2 can promote the acquisition of CD16 and KIR on CD56hi CD16− NK cells. Because the latter are selectively enriched in the LN, one possibility is that T cell–derived IL-2 is involved in the conversion of CD56hi CD16− NK cells to the CD56dim CD16+ phenotype in vivo (88, 134). This would represent a clear example of adaptive immunity inﬂuencing innate immune development.

IRF: IFN regulatory factor

NK Maturation: Signaling Pathways Most activating receptors on NK cells (Table 1) couple to one of three primary signaling pathways: DAP12/Syk-ZAP70, DAP10/PI-3K, or SAP/Fyn. Phospholipase Cγ (PLCγ)-1 and PLCγ2 and Vav family www.annualreviews.org • NK Cell Development

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members are subsequently activated to transmit these membrane events to NK cell effector functions (reviewed in 20, 21, 45). Recent reports provide evidence for selective targeting of NK cell cytotoxicity through some of these biochemical pathways (Table 2). NK cells develop normally in the absence of Syk-ZAP70, DAP10, or SAP/Fyn (16, 135, 136), demonstrating that individually these signaling pathways are redundant for normal NK cell generation. This result is in marked contrast with B and T cell development, which require signaling through immunoreceptor tyrosine-based activating motif (ITAM)-containing antigen receptors (16). In contrast, cytotoxicity and cytokine production are completely inhibited when NK cells are triggered via CD16 or Ly49D in Syk-ZAP70-deﬁcient mice, via NKG2D in DAP10-deﬁcient mice, or via 2B4 in SAP/Fyn-deﬁcient mice (16, 135, 136). Further analyses of NK cells in SykZAP70-, DAP10-, or SAP/Fyn-mutant mice have provided new insights into the receptors responsible for natural cytotoxicity. SykZAP70-deﬁcient NK cells kill YAC1 targets normally, demonstrating that ITAM-bearing receptors are not required for lysis (16); YAC1 killing can be inhibited by blocking NKG2D (137), and YAC1 killing is compromised in the absence of DAP10 (136), suggesting that NKG2D/DAP10 signaling (138) is the major receptor involved in YAC1 recognition and lysis. Interestingly, Syk-ZAP70-deﬁcient NK cells could lyse targets that do not express NKG2D ligands (16), thereby suggesting additional pathways that are involved in natural cytotoxicity. The cytosolic protein adaptor SAP is necessary to couple activating signals from several NK cell receptors (2B4, NTB-A; reviewed in 21). Recent reports from the Latour and Kumar laboratories demonstrate that 2B4:CD48 interactions that signal through a SAP/Fyn cascade operate independently of NKG2D activation (135, 139). Whether 2B4/SAP/Fyn activation provides an ITAM- and NKG2D-independent path-

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way for NK cell–mediated cytotoxicity remains to be tested. PLCγ is a central regulator of intracellular calcium mobilization and protein kinase C activation that is engaged following triggering of different lymphocyte receptors (reviewed in 140). Previous studies have demonstrated a nonredundant role for PLCγ2 in NK cytotoxicity (141); whether this molecule played other roles in NK cell development was not fully appreciated. Work from the Colonna and Colucci laboratories shows that PLCγ2 controls not only NK cell cytotoxicity but also inﬂuences the formation of the Ly49 receptor repertoire (48, 141a). How PLCγ2 controls cytotoxicity is not known, but evidence points to a defect in the process of granule polarization and secretion. These NK abnormalities have biological consequences in vivo, as PLCγ2-deﬁcient mice fail to control experimental CMV infection and to reject tumors. PLCγ2 therefore plays an essential role in establishing a diverse NK cell pool and in maintaining distinct NK cell functions in mature NK cells. Work from the Leibson laboratory indicates that ITAM-containing receptors in human NK cells can use either PLCγ1 or PLCγ2, whereas NKG2D only activates PLCγ2, demonstrating that certain receptor pathways may be hardwired (49, 142). This observation provides a potential means to selectively target NKG2D function in cytotoxic cells. Similar observations were made in mice deﬁcient in the three different members of the Vav family of guanine nucleotide exchange factors (47). In this case Vav1 was selectively coupled to NKG2D, whereas Vav2 and 3 were required for ITAM-mediated signaling. Interestingly, mice lacking all three Vav family members developed normal numbers of phenotypically mature NK cells (47). A recent NK meeting report suggests that receptors of the Axl protein tyrosine kinase (PTK) family are involved in NK cell maturation in the bone marrow (143). A differential transcript analysis identiﬁed Axl in functionally distinct subsets of NK cells. Axl is a

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member of the Tyro3 family of PTKs (which includes Tyro3, Mer, and Axl) that control diverse biological processes, including spermatogenesis, inﬂammation, and possibly malignant transformation (reviewed in 144). Ligands for Axl include Gas6 and Protein S, which are expressed by stromal cells. Interestingly, NK cells require Axl PTKs for their functional maturation and for their acquisition of Ly49 receptor repertoire (143). These results identify a new signaling axis that is crucial for proper NK cell differentiation in mice.

NK Maturation: Transcription Factors Several TFs (see Table 2) guide the process of NK cell maturation, and their absence results in incomplete or aberrant NK cell differentiation (reviewed in 24, 68). Closer examination of the phenotype and function of NK cells in TF mutant mice reveals that certain TFs appear to exert their roles during the iNK stage in the bone marrow (Gata-3, IRF-2, T-bet), whereas others inﬂuence the effector functions of mature NK cells [myeloid ELF1-like factor (MEF), the microphthalmia-associated TF (MITF), and the CCAAT/enhancer-binding protein-γ (CEBP-γ); Figure 3]. There are some striking similarities in the phenotype of bone marrow NK cells from some of these different TF mutant mice, suggesting that certain aspects of the NK cell differentiation process may be linked by TF networks (68). Gata-3 and the T-box TF T-bet have opposing roles in T helper cell differentiation, yet these TFs are co-expressed in developing NK cells at the population (145) and singlecell level (J.P. Di Santo, unpublished observations). The comparison of Gata-3-deﬁcient and T-bet-deﬁcient NK cells suggests a complex interaction between these two TFs during NK cell development. Previous studies on NK cells from chimeras generated with Gata-3-deﬁcient HSC demonstrated that this TF is required for proper expression of CD11b, CD43, and

TRANSCRIPTION FACTORS

NKP

iNK

mNK

Ets-1

Gata-3

CEBP-γ

Id2

IRF-2

MEF

Ikaros

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MITF

PU.1 Figure 3 TFs that condition NK cell development, maturation, and effector functions. Several TFs have been identiﬁed that play an important role in the generation of NKP (Ets-1, Id2, Ikaros, PU.1), the further maturation of iNK (Gata-3, IRF-2, T-bet), or the functional differentiation of mNK (CEBP-γ, MEF, MITF). The interplay between these TFs remains to be fully elucidated.

Ly49 receptors on developing NK cells (109). Although abnormal in phenotype, Gata3-deﬁcient NK cells were fully cytotoxic, demonstrating that the acquisition of some effector functions could be achieved in the absence of a mature NK cell phenotype. Gata3-deﬁcient NK cells were poor producers of IFN-γ, and this defect was associated with reduced levels of T-bet and Hlx (109). Thus, Gata-3 promotes several different aspects of the NK cell differentiation program (reviewed in 68), and its expression is linked to T-bet expression. In comparison, T-bet-deﬁcient NK cells showed a similar phenotypic abnormality (CD11blo CD43lo ) and were normally cytotoxic. Interestingly, NK cells that develop in the absence of T-bet were unable to sustain IFN-γ production after stimulation (145). The residual IFN-γ production may result from the compensatory expression of another T-box TF, Eomesodermin, in T-bet-deﬁcient NK cells (145). It would be interesting to know whether Eomesodermin expression is modiﬁed in Gata-3-deﬁcient NK cells. A recent study reported a more complete analysis of NK cell differentiation in mice deﬁcient in IRF-2 (129). Previous work had shown that IRF-2 is required in a www.annualreviews.org • NK Cell Development

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cell-intrinsic fashion for normal development of NK cells (146). Taki and colleagues (129) found that IRF-2-deﬁcient NK cells are poor IFN-γ producers, which recalls previous reports of the defect in Th1 development seen in these mice (146). They further demonstrated that IRF-2 plays an important role in NK cell development in the bone marrow and is required for normal CD11b and CD43 expression. NK cells in the bone marrow of IRF-2-deﬁcient mice have an immature DX5+ Ly49+ CD11blo CD43lo phenotype (Figure 1, stage D), although they appear normally cytotoxic against sensitive targets (129). In contrast, splenic NK cells are strongly reduced in number, have an even less mature DX5− phenotype, and lack Ly49 receptor expression altogether (Figure 1, stage C). Fully mature NK cells are basically absent in the bone marrow and spleen of IRF-2deﬁcient mice. This result suggests that IRF-2 plays a major role in maintaining developing NK cells in the bone marrow after stage D. The presence of the few immature stage C NK cells in the spleen of IRF-2-deﬁcient mice suggests that NK cells can export the bone marrow before their differentiation is complete. TFs likely act in a sequential fashion to promote NK cell maturation such that different TFs specify the transcriptional proﬁles that could be associated with unique functional outcomes (Figure 3). For example, early stages of NK cell development in the bone marrow and liver are characterized by TRAIL expression (Figure 1, stage B), which does not persist as NK cells mature and take up residence in the spleen. One may speculate that certain TFs encode TRAIL expression at this stage, and transcriptional proﬁling of developing NK may provide an answer. Nevertheless, the interplay between Gata-3, IRF-2, and T-bet appears complex. Some hierarchical organization can be proposed with a sequential (Gata-3 to IRF-2 to T-bet) TF activation cascade. Gata-3 may control CD11b and CD43 expression through IRF-2 and/or T-bet, since NK cells from mice

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lacking these TFs share the same phenotype (129, 145). Moreover, Gata-3 expression in IRF-2-deﬁcient NK cells and IRF-2 expression in T-bet-deﬁcient NK cells are normal (129, 145). The effector functions in phenotypically mature NK cells are insured by a group of TFs that include the MEF, MITF, and CEBP-γ (Figure 3). Eomesodermin may also belong in this group, although the phenotype of Eomesodermin-deﬁcient NK cells has not yet been reported. NK cells deﬁcient in MEF, MITF, or CEBP-γ develop normally but have reduced cytotoxicity and cytokine production. The molecular mechanisms include decreased perforin or granzyme expression, providing an example of how TFs can promote speciﬁc functional outcomes in fully differentiated NK cells. One conclusion from the analysis of TF mutants is that phenotypically immature NK cells can attain functional cytolytic competence. This may reﬂect a degree of functional heterogeneity within the NK cell pool: NK cells may be capable of eliciting independent or linked effector functions. T-bet and Gata-3 co-expression in NK cells could be an example of a mechanism to maximize options for programming diverse effector functions. NK cells lacking classical functions (cytotoxicity, IFN-γ secretion) are not necessarily anergic but may nonetheless contribute to immune responses in other ways (for example, as regulatory NK cells secreting chemotactic factors).

HOMEOSTASIS OF THE PERIPHERAL NK CELL POOL Mature NK cells are found in the blood, bone marrow, spleen, liver, LNs, lung, and omentum and in the uterus during gestation. The steady-state levels of these peripheral NK cells are tightly controlled through a process referred to as homeostasis. NK cell homeostasis is controlled by cell-intrinsic mechanisms (via TFs) and by extrinsic signals; the latter include the cytokine IL-15 and the transforming growth factor (TGF)-β (14, 124,

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4 NKT mNK Figure 4 The peripheral NK cell niche. Newly generated (or recirculating) NK cells enter peripheral tissues (1) and require IL-15 signals for survival (2). Upon stimulation (during infection or stress), NK cells may proliferate (3) or further differentiate (4). The peripheral NK cell niche may be the site of cellular competition for IL-15 between IL-15-dependent cells, such as CD8mem and NKT cells (5). NK cells may leave the peripheral niche to recirculate to other sites (6).

126, 131). Once again, a balance must be achieved between survival, proliferation, and cell death to maintain an optimal pool of NK cells that can participate in immune defense (Figure 4). IL-15 plays a major role in peripheral NK cell homeostasis. Transfer studies have shown that IL-15 maintains the survival and promotes the proliferation of mature NK cells in vivo (124, 126, 131). This effect is mediated through maintenance of the antiapoptotic factor bcl-2, which can protect mature NK cells

from death after cytokine withdrawal (132). As alluded to above, IRF-2 and T-bet are also involved in protecting mature NK cells from apoptosis. Deﬁciencies in either of these two TFs result in a reduced ability to maintain the peripheral NK cell pool (129, 145). The mechanisms that account for the cell death observed in IRF-2- or T-bet-deﬁcient NK cells are unknown, but the enhanced apoptosis does not appear to be due to an inability to respond to IL-15 (129, 145). One possibility is that an activation-induced signal is involved www.annualreviews.org • NK Cell Development

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because IRF-2-deﬁcient NK cells express elevated levels of CD69 (129). Substantial progress has been made in deciphering the cellular context in which IL-15 promotes NK cell homeostasis. Studies from the Waldmann laboratory provided the ﬁrst evidence for a novel mode of IL-15 transpresentation, involving a cell surface IL-15Rα– IL-15 complex that could activate responding cells via a complex comprising CD122 (IL2Rβ) and γc (127). In vivo HSC transfer experiments have clearly validated this model by demonstrating that both hematopoietic and parenchymal (nonhematopoietic) cells can transpresent IL-15, and that IL-15 and IL-15Rα need to be expressed by the same cell to do so (147–149). Once in the periphery, NK cells reside in IL-15-dependent niches, where they may simply survive, proliferate, or further differentiate (69, 126, 131, 150). Competition between NK cells and other IL-15dependent lymphocytes has been documented after adoptive transfer (151), but whether these cells actually compete in vivo under physiological conditions is not known. Collectively, these results provide a ﬁrst glimpse into the NK cell niche that can be further dissected by visualizing the cells that produce the IL-15Rα–IL-15 complex. Negative regulation also appears to operate under steady-state conditions to maintain the homeostasis of the NK cell pool. TGF-β is a potent antiproliferative factor that can suppress the proliferation of different lymphocyte subsets in vivo (reviewed in 152). In this way, tumor-derived TGF-β may contribute to tumor escape from immunosurveillance. The absence of TGF-β results in a multiorgan inﬂammatory syndrome with prominent lymphocytic inﬁltration (153). A recent report from the Flavell laboratory showed that TGF-β also regulates NK cell homeostasis in the mouse (14). A dominantnegative truncated form of the TGF-βRII controlled by the CD11c promoter was expressed in NK cells and resulted in the expansion and activation of the splenic NK cell pool. NK cell maturation was normal, but the

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peripheral NK cell pool was established more rapidly. Because cycling peripheral NK cells were not increased, NK cell survival was likely augmented in the absence of TGF-β signaling. An alternative explanation is that bone marrow NK cell development was quantitatively enhanced. Thus, TGF-β signaling acts as a negative regulator of NK cell homeostasis. It is not clear at which stage of NK cell development TGF-β exerts its effects or the nature of the cell type that provides TGF-β. Human NK cells treated with TGF-β modulate their expression of NKG2D and NCR, providing a potential mechanism to account for the suppressive effects of TGF-β on NK cell cytotoxicity (154). It is tempting to speculate that regulatory T cell subsets might also be involved because these cells can use a TGF-β-dependent effector mechanism to suppress T cell activation and thereby maintain tolerance (155).

ORGAN-SPECIFIC FUNCTIONS FOR NK CELLS? The fact that NK cells are derived from HSC is unquestioned. In contrast, the deﬁnitive site(s) for NK cell development can only be inferred from where we ﬁnd immature and mature NK cells. NK cells present in different organs may subserve different functions. A tissue hierarchy is suggested when one examines the presence of NK cells at different developmental stages in various organs (Figure 5).

NK Cell Development: Beyond the Bone Marrow The bone marrow is considered the primary site of NK cell generation because it harbors the cellular substrates and soluble factors required for NK cells to mature. Bone marrow ablation (after treatment with bone-seeking isotopes or following estrogen-inducing osteopetrosis) affects peripheral NK cell numbers and their functional capacities (17, 156, 157). Although often cited as evidence that the

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Thymus

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bone marrow is required for complete NK cell maturation, these treatments may affect the capacity of NK precursors to respond to maturation signals that they might see elsewhere. Whether the bone marrow alone is sufﬁcient to assure complete NK cell development is still unknown. The bone marrow may only be involved in the initial steps of NK cell differentiation, with other sites required for ﬁnal maturation. Could the bone marrow represent the generative site for NKP that ﬁnish their maturation elsewhere, perhaps in the liver, LN, or spleen? The identiﬁcation of a LN-resident NKP in humans (88) that apparently trafﬁcs from the bone marrow via the blood to the LN would be consistent with this possibility. The identiﬁcation of NKP and immature TRAIL+ NK cells in the liver and spleen (19, 25, 109; C.A.J. Vosshenrich & J.P. Di Santo, unpublished observations) provide additional evidence. Sev-

Lymph nodes

Generation and recirculation of NK cells. ELP, NKP, and iNK are found in the bone marrow. Whether these developmental intermediates leave the bone marrow to complete their differentiation elsewhere is unknown, although NKP and iNK can be found in the liver and spleen. Whether NK cells generated in the thymus have speciﬁc homing potential is not known.

eral alternatives to the bone marrow for completing some aspects of NK cell differentiation exist and should be considered. The bone marrow environment may also provide a niche for mature NK cells. It is accepted that long-lived, terminally differentiated plasma cells reside in the bone marrow (158), and recent evidence demonstrates that memory CD8+ T cells ﬁnd long-term refuge in the bone marrow as well (159). Could a long-lived subset of NK cells exist that follows a similar pattern?

NK Cells in the Thymus Small numbers of NK cells are found in the thymus that are phenotypically mature, are cytotoxic, and can produce cytokines (78, 160). The thymus can export mature NK cells as evidenced by transplants of Ragdeﬁcient thymuses in alymphoid recipients www.annualreviews.org • NK Cell Development

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(Rag2-deﬁcient x γc -deﬁcient mice) that show a small but detectable population of donorderived NK cells in the spleen ( J.P. Di Santo, unpublished observations). An open question remains as to whether the mature NK cells found in the thymus develop and differentiate in situ in a fashion similar to that observed in the bone marrow. NK cells present in Ragdeﬁcient thymuses resemble their wild-type counterparts, suggesting that neither more mature thymocytes nor a fully developed thymic medullary stromal compartment is required for thymic NK cell homeostasis. What could be the function of NK cells within the thymus? Speciﬁc roles could include modulating thymopoiesis, maintaining thymic architecture, or contributing to defense against thymocyte transformation. Previous reports demonstrate that NK cells could speciﬁcally lyse early thymocytes (161) such that thymic NK cells could be involved in immunosurveillance of rapidly dividing thymic precursors.

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NK Cell Recirculation Few data are available on the recirculation of NK cells between various organs that harbor them: Adoptive transfer experiments have shown that splenic NK cells can repopulate the spleen, liver, and bone marrow of recipient mice (69, 124, 126). Whether this tissue repartition is uniform (all NK cells migrate equally well to all tissues) or selective (subsets

of NK cells migrate to speciﬁc tissues) has not been extensively studied. NK cells are rare or absent in most tissues, including the digestive tract, muscle, brain, and skin. NK cells are not present in uterine tissue in the steady-state in the mouse (but are in humans), and are massively increased during pregnancy in both species (reviewed in 162). There is no evidence that NK cells develop at these sites. Nevertheless, NK cells are rapidly recruited to tissues after infection or inﬂammation. Chemokines play a major role in this process, and NK cells express a panoply of chemokine receptors (including CCR2, CCR5, CX3CR1, and CXCR3) that allow them to participate in diverse types of inﬂammatory reactions (reviewed in 30, 31). Whether chemokine receptors also dictate NK cell export from the bone marrow or thymus or allow speciﬁc homing to the liver, spleen, or LN is probable, although no published data exist in the mouse. Little is known about the precise physical localization of mature NK or their precursor cells in the bone marrow, spleen, or LN under normal or pathological conditions. Liver NK cells patrol the sinusoids and likely act as sensors for hepatic parenchymal inﬂammation and stress. The topological organization of developing NK cells within generative organs (bone marrow, thymus) under steadystate conditions and following stress of infection or inﬂammation could provide important clues to the mechanisms that control NK cell development.

SUMMARY POINTS 1. NK precursors and immature NK cells have been identiﬁed in mice and humans. The developmental intermediates in NK cell differentiation mature through the concerted action of transcription factors that specify their phenotype and effector functions. The wealth of new information on developing NK cells has been integrated into a coherent model for NK cell development from hematopoietic precursors. Nevertheless, the precise sites where NK cell maturation take place are not yet fully deﬁned.

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2. Clear evidence for functional NK cell subsets exists in humans. Whether this “division of labor” applies to NK cells from other species is unknown, but could provide a means to allow NK cells to respond under diverse conditions. The presence of NK cells in the thymus and immature NK cells in the liver suggests speciﬁc functions for NK cells in these tissues. 3. New assays for NK cell development should help investigators to understand the process. Novel stromal cell–based systems and humanized mouse models will undoubtedly provide the impetus for re-examining the models of human NK cell differentiation and may provide answers to the long-standing questions regarding the origins of different NK cell subsets and their biological roles in normal and pathophysiological processes.

ACKNOWLEDGMENTS J.P.D. is supported by grants from the Institut Pasteur, the Institut National de la Sant´e et de la Recherche M´edicale (INSERM), and the Ligue Nationale contre le Cancer. I thank past and present members of my laboratory for their enthusiastic collaboration, and M. Colonna, F. Colucci, C.A.J. Vosshenrich, and A. Wilson for sharing unpublished results.

LITERATURE CITED 1. Herberman RB, Nunn ME, Holden HT, Lavrin DH. 1975. Natural cytotoxic reactivity of mouse lymphoid cells against syngeneic and allogeneic tumors. II. Characterization of effector cells. Int. J. Cancer 16:230–39 2. Herberman RB, Nunn ME, Lavrin DH. 1975. Natural cytotoxic reactivity of mouse lymphoid cells against syngeneic acid allogeneic tumors. I. Distribution of reactivity and speciﬁcity. Int. J. Cancer 16:216–29 3. Kiessling R, Klein E, Pross H, Wigzell H. 1975. “Natural” killer cells in the mouse. II. Cytotoxic cells with speciﬁcity for mouse Moloney leukemia cells. Characteristics of the killer cell. Eur. J. Immunol. 5:117–21 4. Kiessling R, Klein E, Wigzell H. 1975. “Natural” killer cells in the mouse. I. Cytotoxic cells with speciﬁcity for mouse Moloney leukemia cells. Speciﬁcity and distribution according to genotype. Eur. J. Immunol. 5:112–17 5. Rosenberg EB, McCoy JL, Green SS, Donnelly FC, Siwarski DF, et al. 1974. Destruction of human lymphoid tissue-culture cell lines by human peripheral lymphocytes in 51Cr-release cellular cytotoxicity assays. J. Natl. Cancer Inst. 52:345–52 6. Herberman RB, Ortaldo JR. 1981. Natural killer cells: their roles in defenses against disease. Science 214:24–30 7. de Villartay JP, Fischer A, Durandy A. 2003. The mechanisms of immune diversiﬁcation and their disorders. Nat. Rev. Immunol. 3:962–72 8. Shinkai Y, Rathbun G, Lam KP, Oltz EM, Stewart V, et al. 1992. RAG-2-deﬁcient mice lack mature lymphocytes owing to inability to initiate V(D)J rearrangement. Cell 68:855–67 9. Trinchieri G. 1989. Biology of natural killer cells. Adv. Immunol. 47:187–376 10. Yu YY, Kumar V, Bennett M. 1992. Murine natural killer cells and marrow graft rejection. Annu. Rev. Immunol. 10:189–213 www.annualreviews.org • NK Cell Development

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11. Moffett-King A. 2002. Natural killer cells and pregnancy. Nat. Rev. Immunol. 2:656–63 12. Orange JS. 2002. Human natural killer cell deﬁciencies and susceptibility to infection. Microbes Infect. 4:1545–58 13. French AR, Yokoyama WM. 2003. Natural killer cells and viral infections. Curr. Opin. Immunol. 15:45–51 14. Laouar Y, Sutterwala FS, Gorelik L, Flavell RA. 2005. Transforming growth factor-β controls T helper type 1 cell development through regulation of natural killer cell interferon-γ. Nat. Immunol. 6:600–7 15. Pelayo R, Hirose J, Huang J, Garrett KP, Delogu A, et al. 2005. Derivation of 2 categories of plasmacytoid dendritic cells in murine bone marrow. Blood 105:4407–15 16. Colucci F, Schweighoffer E, Tomasello E, Turner M, Ortaldo JR, et al. 2002. Natural cytotoxicity uncoupled from the Syk and ZAP-70 intracellular kinases. Nat. Immunol. 3:288–94 17. Puzanov IJ, Bennett M, Kumar V. 1996. IL-15 can substitute for the marrow microenvironment in the differentiation of natural killer cells. J. Immunol. 157:4282–85 18. Rosmaraki EE, Douagi I, Roth C, Colucci F, Cumano A, Di Santo JP. 2001. Identiﬁcation of committed NK cell progenitors in adult murine bone marrow. Eur. J. Immunol. 31:1900–9 19. Kim S, Iizuka K, Kang HS, Dokun A, French AR, et al. 2002. In vivo developmental stages in murine natural killer cell maturation. Nat. Immunol. 3:523–28 20. Colucci F, Di Santo JP, Leibson PJ. 2002. Natural killer cell activation in mice and men: different triggers for similar weapons? Nat. Immunol. 3:807–13 21. Latour S, Veillette A. 2004. The SAP family of adaptors in immune regulation. Semin. Immunol. 16:409–19 22. Lanier LL. 2005. NK cell recognition. Annu. Rev. Immunol. 23:225–74 23. Moretta L, Moretta A. 2004. Unravelling natural killer cell function: triggering and inhibitory human NK receptors. EMBO J. 23:255–59 24. Colucci F, Caligiuri MA, Di Santo JP. 2003. What does it take to make a natural killer? Nat. Rev. Immunol. 3:413–25 25. Takeda K, Cretney E, Hayakawa Y, Ota T, Akiba H, et al. 2005. TRAIL identiﬁes immature natural killer cells in newborn mice and adult mouse liver. Blood 105:2082– 89 26. Arase H, Saito T, Phillips JH, Lanier LL. 2001. Cutting edge: the mouse NK cellassociated antigen recognized by DX5 monoclonal antibody is CD49b (α2 integrin, very late antigen-2). J. Immunol. 167:1141–44 27. Smyth MJ, Cretney E, Kelly JM, Westwood JA, Street SE, et al. 2005. Activation of NK cell cytotoxicity. Mol. Immunol. 42:501–10 28. Screpanti V, Wallin RP, Ljunggren HG, Grandien A. 2001. A central role for death receptor-mediated apoptosis in the rejection of tumors by NK cells. J. Immunol. 167:2068–73 29. Cooper MA, Fehniger TA, Caligiuri MA. 2001. The biology of human natural killer-cell subsets. Trends Immunol. 22:633–40 30. Robertson MJ. 2002. Role of chemokines in the biology of natural killer cells. J. Leukoc. Biol. 71:173–83 31. Maghazachi AA. 2003. G protein-coupled receptors in natural killer cells. J. Leukoc. Biol. 74:16–24 32. Fehniger TA, Cooper MA, Nuovo GJ, Cella M, Facchetti F, et al. 2003. CD56bright natural killer cells are present in human lymph nodes and are activated by T cell-derived IL-2: a potential new link between adaptive and innate immunity. Blood 101:3052–57

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

278

Di Santo

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

ARI

15 February 2006

17:51

33. Benlagha K, Kyin T, Beavis A, Teyton L, Bendelac A. 2002. A thymic precursor to the NK T cell lineage. Science 296:553–55 34. Loza MJ, Perussia B. 2001. Final steps of natural killer cell maturation: a model for type 1-type 2 differentiation? Nat. Immunol. 2:917–24 35. Karre K, Ljunggren HG, Piontek G, Kiessling R. 1986. Selective rejection of H2-deﬁcient lymphoma variants suggests alternative immune defence strategy. Nature 319:675–78 36. Ljunggren HG, Karre K. 1990. In search of the ‘missing self’: MHC molecules and NK cell recognition. Immunol. Today 11:237–44 37. Long EO. 1999. Regulation of immune responses through inhibitory receptors. Annu. Rev. Immunol. 17:875–904 38. Yokoyama WM. 1993. Recognition structures on natural killer cells. Curr. Opin. Immunol. 5:67–73 39. Lanier LL. 1998. NK cell receptors. Annu. Rev. Immunol. 16:359–93 40. Raulet DH, Vance RE, McMahon CW. 2001. Regulation of the natural killer cell receptor repertoire. Annu. Rev. Immunol. 19:291–330 41. Fernandez NC, Treiner E, Vance RE, Jamieson AM, Lemieux S, Raulet DH. 2005. A subset of natural killer cells achieves self-tolerance without expressing inhibitory receptors speciﬁc for self-MHC molecules. Blood 105:4416–23 41a. Kim S, Poursine-Laurent J, Truscott SM, Lybarger L, Song YJ, et al. 2005. Licensing of natural killer cells by host major histocompatibility complex class I molecules. Nature 436:709–13 41b. Kumar V, McNerney ME. 2005. A new self:MHC-class-I-independent natural-killercell self-tolerance. Nat. Rev. Immunol. 5:363–74 42. Moretta A, Bottino C, Vitale M, Pende D, Cantoni C, et al. 2001. Activating receptors and coreceptors involved in human natural killer cell-mediated cytolysis. Annu. Rev. Immunol. 19:197–223 43. Raulet DH. 2003. Roles of the NKG2D immunoreceptor and its ligands. Nat. Rev. Immunol. 3:781–90 44. Yokoyama WM, Plougastel BF. 2003. Immune functions encoded by the natural killer gene complex. Nat. Rev. Immunol. 3:304–16 45. Vivier E, Nunes JA, Vely F. 2004. Natural killer cell signaling pathways. Science 306:1517–19 46. Colucci F, Rosmaraki E, Bregenholt S, Samson SI, Di Bartolo V, et al. 2001. Functional dichotomy in natural killer cell signaling: Vav-1-dependent and -independent mechanisms. J. Exp. Med. 193:1–13 47. Cella M, Fujikawa K, Tassi I, Kim S, Latinis K, et al. 2004. Differential requirements for Vav proteins in DAP10- and ITAM-mediated NK cell cytotoxicity. J. Exp. Med. 200:817–23 48. Tassi I, Presti R, Kim S, Yokoyama WM, Gilﬁllan S, Colonna M. 2005. PLCγ2 is a critical signaling mediator for murine NK cell activating receptors. J. Immunol. 175:749– 54 49. Upshaw JL, Schoon RA, Dick CJ, Billadeau DD, Leibson PJ. 2005. The isoforms of phospholipase C-γ are differentially used by distinct human NK activating receptors. J. Immunol. 175:213–18 50. Carayannopoulos LN, Yokoyama WM. 2004. Recognition of infected cells by natural killer cells. Curr. Opin. Immunol. 16:26–33 www.annualreviews.org • NK Cell Development

279

ARI

15 February 2006

17:51

51. Colonna M, Nakajima H, Navarro F, Lopez-Botet M. 1999. A novel family of Ig-like receptors for HLA class I molecules that modulate function of lymphoid and myeloid cells. J. Leukoc. Biol. 66:375–81 52. Bendelac A, Bonneville M, Kearney JF. 2001. Autoreactivity by design: innate B and T lymphocytes. Nat. Rev. Immunol. 1:177–86 53. Smyth MJ, Hayakawa Y, Takeda K, Yagita H. 2002. New aspects of natural-killer-cell surveillance and therapy of cancer. Nat. Rev. Cancer 2:850–61 54. Du Pasquier L, Zucchetti I, De Santis R. 2004. Immunoglobulin superfamily receptors in protochordates: before RAG time. Immunol. Rev. 198:233–48 55. Biron CA, Byron KS, Sullivan JL. 1989. Severe herpesvirus infections in an adolescent without natural killer cells. N. Engl. J. Med. 320:1731–35 56. Brown MG, Dokun AO, Heusel JW, Smith HR, Beckman DL, et al. 2001. Vital involvement of a natural killer cell activation receptor in resistance to viral infection. Science 292:934–37 57. Lee SH, Girard S, Macina D, Busa M, Zafer A, et al. 2001. Susceptibility to mouse cytomegalovirus is associated with deletion of an activating natural killer cell receptor of the C-type lectin superfamily. Nat. Genet. 28:42–45 58. Biron CA, Nguyen KB, Pien GC, Cousens LP, Salazar-Mather TP. 1999. Natural killer cells in antiviral defense: function and regulation by innate cytokines. Annu. Rev. Immunol. 17:189–220 59. French AR, Pingel JT, Wagner M, Bubic I, Yang L, et al. 2004. Escape of mutant double-stranded DNA virus from innate immune control. Immunity 20:747–56 60. Martin-Fontecha A, Thomsen LL, Brett S, Gerard C, Lipp M, et al. 2004. Induced recruitment of NK cells to lymph nodes provides IFN-γ for TH 1 priming. Nat. Immunol. 5:1260–55 61. Haller O, Kiessling R, Orn A, Wigzell H. 1977. Generation of natural killer cells: an autonomous function of the bone marrow. J. Exp. Med. 145:1411–16 62. Hackett J Jr, Bosma GC, Bosma MJ, Bennett M, Kumar V. 1986. Transplantable progenitors of natural killer cells are distinct from those of T and B lymphocytes. Proc. Natl. Acad. Sci. USA 83:3427–31 63. Shibuya A, Nagayoshi K, Nakamura K, Nakauchi H. 1995. Lymphokine requirement for the generation of natural killer cells from CD34+ hematopoietic progenitor cells. Blood 85:3538–46 64. Miller JS, Alley KA, McGlave P. 1994. Differentiation of natural killer (NK) cells from human primitive marrow progenitors in a stroma-based long-term culture system: identiﬁcation of a CD34+ 7+ NK progenitor. Blood 83:2594–601 65. Mrozek E, Anderson P, Caligiuri MA. 1996. Role of interleukin-15 in the development of human CD56+ natural killer cells from CD34+ hematopoietic progenitor cells. Blood 87:2632–40 66. Jaleco AC, Blom B, Res P, Weijer K, Lanier LL, et al. 1997. Fetal liver contains committed NK progenitors, but is not a site for development of CD34+ cells into T cells. J. Immunol. 159:694–702 67. Zamai L, Ahmad M, Bennett IM, Azzoni L, Alnemri ES, Perussia B. 1998. Natural killer (NK) cell-mediated cytotoxicity: differential use of TRAIL and Fas ligand by immature and mature primary human NK cells. J. Exp. Med. 188:2375–80 68. Vosshenrich CA, Samson-Villeger SI, Di Santo JP. 2005. Distinguishing features of developing natural killer cells. Curr. Opin. Immunol. 17:151–58 69. Prlic M, Blazar BR, Farrar MA, Jameson SC. 2003. In vivo survival and homeostatic proliferation of natural killer cells. J. Exp. Med. 197:967–76

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

280

Di Santo

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

ARI

15 February 2006

17:51

70. Igarashi H, Gregory SC, Yokota T, Sakaguchi N, Kincade PW. 2002. Transcription from the RAG1 locus marks the earliest lymphocyte progenitors in bone marrow. Immunity 17:117–30 71. Kondo M, Weissman IL, Akashi K. 1997. Identiﬁcation of clonogenic common lymphoid progenitors in mouse bone marrow. Cell 91:661–72 72. Terskikh AV, Miyamoto T, Chang C, Diatchenko L, Weissman IL. 2003. Gene expression analysis of puriﬁed hematopoietic stem cells and committed progenitors. Blood 102:94–101 73. Galy A, Travis M, Cen D, Chen B. 1995. Human T, B, natural killer, and dendritic cells arise from a common bone marrow progenitor cell subset. Immunity 3:459–73 73a. Blom B, Spits H. 2006. Development of human lymphoid cells. Annu. Rev. Immunol. 24:287–320 74. Waskow C, Paul S, Haller C, Gassmann M, Rodewald H. 2002. Viable c-KitW/W mutants reveal pivotal role for c-kit in the maintenance of lymphopoiesis. Immunity 17:277–88 75. Waskow C, Rodewald HR. 2002. Lymphocyte development in neonatal and adult cKit-deﬁcient (c-KitW/W ) mice. Adv. Exp. Med. Biol. 512:1–10 76. Michie AM, Carlyle JR, Schmitt TM, Ljutic B, Cho SK, et al. 2000. Clonal characterization of a bipotent T cell and NK cell progenitor in the mouse fetal thymus. J. Immunol. 164:1730–33 77. Douagi I, Colucci F, Di Santo JP, Cumano A. 2002. Identiﬁcation of the earliest prethymic bipotent T/NK progenitor in murine fetal liver. Blood 99:463–71 78. Carlyle JR, Michie AM, Cho SK, Zuniga-Pﬂucker JC. 1998. Natural killer cell development and function precede αβ T cell differentiation in mouse fetal thymic ontogeny. J. Immunol. 160:744–53 79. Spits H, Lanier LL, Phillips JH. 1995. Development of human T and natural killer cells. Blood 85:2654–70 80. Schmitt TM, Zuniga-Pﬂucker JC. 2002. Induction of T cell development from hematopoietic progenitor cells by delta-like-1 in vitro. Immunity 17:749–56 81. Porritt HE, Rumfelt LL, Tabrizifard S, Schmitt TM, Zuniga-Pﬂucker JC, Petrie HT. 2004. Heterogeneity among DN1 prothymocytes reveals multiple progenitors with different capacities to generate T cell and non-T cell lineages. Immunity 20:735–45 82. Schmitt TM, Ciofani M, Petrie HT, Zuniga-Pﬂucker JC. 2004. Maintenance of T cell speciﬁcation and differentiation requires recurrent Notch receptor-ligand interactions. J. Exp. Med. 200:469–79 83. Taghon TN, David ES, Zuniga-Pﬂucker JC, Rothenberg EV. 2005. Delayed, asynchronous, and reversible T-lineage speciﬁcation induced by Notch/Delta signaling. Genes Dev. 19:965–78 84. Radtke F, Wilson A, Stark G, Bauer M, van Meerwijk J, et al. 1999. Deﬁcient T cell fate speciﬁcation in mice with an induced inactivation of Notch1. Immunity 10:547–58 85. Ikawa T, Fujimoto S, Kawamoto H, Katsura Y, Yokota Y. 2001. Commitment to natural killer cells requires the helix-loop-helix inhibitor Id2. Proc. Natl. Acad. Sci. USA 98:5164– 69 86. Kennedy MK, Glaccum M, Brown SN, Butz EA, Viney JL, et al. 2000. Reversible defects in natural killer and memory CD8 T cell lineages in interleukin 15-deﬁcient mice. J. Exp. Med. 191:771–80 87. Vosshenrich CA, Ranson T, Samson SI, Corcuff E, Colucci F, et al. 2005. Roles for common cytokine receptor γ-chain-dependent cytokines in the generation, differentiation, and maturation of NK cell precursors and peripheral NK cells in vivo. J. Immunol. 174:1213–21 www.annualreviews.org • NK Cell Development

281

ARI

15 February 2006

17:51

88. Freud AG, Becknell B, Roychowdhury S, Mao HC, Ferketich AK, et al. 2005. A human CD34+ subset resides in lymph nodes and differentiates into CD56bright natural killer cells. Immunity 22:295–304 89. Yu H, Fehniger TA, Fuchshuber P, Thiel KS, Vivier E, et al. 1998. Flt3 ligand promotes the generation of a distinct CD34+ human natural killer cell progenitor that responds to interleukin-15. Blood 92:3647–57 90. Mebius RE, Rennert P, Weissman IL. 1997. Developing lymph nodes collect CD4+ CD3− LTβ+ cells that can differentiate to APC, NK cells, and follicular cells but not T or B cells. Immunity 7:493–504 91. Yoshida H, Kawamoto H, Santee SM, Hashi H, Honda K, et al. 2001. Expression of α4 β7 integrin deﬁnes a distinct pathway of lymphoid progenitors committed to T cells, fetal intestinal lymphotoxin producer, NK, and dendritic cells. J. Immunol. 167:2511–21 92. Williams NS, Klem J, Puzanov IJ, Sivakumar PV, Bennett M, Kumar V. 1999. Differentiation of NK1.1+ , Ly49+ NK cells from ﬂt3+ multipotent marrow progenitor cells. J. Immunol. 163:2648–56 93. Williams NS, Moore TA, Schatzle JD, Puzanov IJ, Sivakumar PV, et al. 1997. Generation of lytic natural killer 1.1+ , Ly-49− cells from multipotential murine bone marrow progenitors in a stroma-free culture: deﬁnition of cytokine requirements and developmental intermediates. J. Exp. Med. 186:1609–14 94. Melchers F, Haasner D, Streb M, Rolink A. 1992. B-lymphocyte lineage-committed, IL-7 and stroma cell-reactive progenitors and precursors, and their differentiation to B cells. Adv. Exp. Med. Biol. 323:111–17 95. Ware CF. 2005. Network communications: lymphotoxins, LIGHT, and TNF. Annu. Rev. Immunol. 23:787–819 96. Iizuka K, Chaplin DD, Wang Y, Wu Q, Pegg LE, et al. 1999. Requirement for membrane lymphotoxin in natural killer cell development. Proc. Natl. Acad. Sci. USA 96:6336–40 97. Lian RH, Chin RK, Nemeth HE, Libby SL, Fu YX, Kumar V. 2004. A role for lymphotoxin in the acquisition of Ly49 receptors during NK cell development. Eur. J. Immunol. 34:2699–707 98. Stevenaert F, Van Beneden K, De Colvenaer V, Franki AS, Debacker V, et al. 2005. Ly49 and CD94/NKG2 receptor acquisition by NK cells does not require lymphotoxin-β receptor expression. Blood 106:956–62 99. Wu Q, Sun Y, Wang J, Lin X, Wang Y, et al. 2001. Signal via lymphotoxin-βR on bone marrow stromal cells is required for an early checkpoint of NK cell development. J. Immunol. 166:1684–89 100. Sivakumar V, Hammond KJ, Howells N, Pfeffer K, Weih F. 2003. Differential requirement for Rel/nuclear factor κB family members in natural killer T cell development. J. Exp. Med. 197:1613–21 101. Chen KS, DeLuca HF. 1994. Isolation and characterization of a novel cDNA from HL60 cells treated with 1,25-dihydroxyvitamin D-3. Biochim. Biophys. Acta 1219:26–32 102. Lee KN, Kang HS, Jeon JH, Kim EM, Yoon SR, et al. 2005. VDUP1 is required for the development of natural killer cells. Immunity 22:195–208 103. Rothenberg EV, Taghon T. 2005. Molecular genetics of T cell development. Annu. Rev. Immunol. 23:601–49 104. Singh H, Medina KL, Pongubala JM. 2005. Contingent gene regulatory networks and B cell fate speciﬁcation. Proc. Natl. Acad. Sci. USA 102:4949–53 105. Scott EW, Simon MC, Anastasi J, Singh H. 1994. Requirement of transcription factor PU.1 in the development of multiple hematopoietic lineages. Science 265:1573–77

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

282

Di Santo

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

ARI

15 February 2006

17:51

106. DeKoter RP, Lee HJ, Singh H. 2002. PU.1 regulates expression of the interleukin-7 receptor in lymphoid progenitors. Immunity 16:297–309 107. Medina KL, Pongubala JM, Reddy KL, Lancki DW, Dekoter R, et al. 2004. Assembling a gene regulatory network for speciﬁcation of the B cell fate. Dev. Cell 7:607–17 108. Maeurer MJ, Lotze MT. 1998. Interleukin-7 (IL-7) knockout mice. Implications for lymphopoiesis and organ-speciﬁc immunity. Int. Rev. Immunol. 16:309–22 109. Samson SI, Richard O, Tavian M, Ranson T, Vosshenrich CA, et al. 2003. GATA-3 promotes maturation, IFN-γ production, and liver-speciﬁc homing of NK cells. Immunity 19:701–11 110. Colucci F, Samson SI, DeKoter RP, Lantz O, Singh H, Di Santo JP. 2001. Differential requirement for the transcription factor PU.1 in the generation of natural killer cells versus B and T cells. Blood 97:2625–32 111. Nutt SL, Metcalf D, D’Amico A, Polli M, Wu L. 2005. Dynamic regulation of PU.1 expression in multipotent hematopoietic progenitors. J. Exp. Med. 201:221–31 112. Barton K, Muthusamy N, Fischer C, Ting CN, Walunas TL, et al. 1998. The Ets-1 transcription factor is required for the development of natural killer cells in mice. Immunity 9:555–63 113. Boggs SS, Trevisan M, Patrene K, Geogopoulos K. 1998. Lack of natural killer cell precursors in fetal liver of Ikaros knockout mutant mice. Nat. Immunol. 16:137–45 114. Engel I, Murre C. 2001. The function of E- and Id proteins in lymphocyte development. Nat. Rev. Immunol. 1:193–99 115. Bain G, Maandag EC, Izon DJ, Amsen D, Kruisbeek AM, et al. 1994. E2A proteins are required for proper B cell development and initiation of immunoglobulin gene rearrangements. Cell 79:885–92 116. Heemskerk MH, Blom B, Nolan G, Stegmann AP, Bakker AQ, et al. 1997. Inhibition of T cell and promotion of natural killer cell development by the dominant negative helix loop helix factor Id3. J. Exp. Med. 186:1597–602 117. Yokota Y, Mansouri A, Mori S, Sugawara S, Adachi S, et al. 1999. Development of peripheral lymphoid organs and natural killer cells depends on the helix-loop-helix inhibitor Id2. Nature 397:702–6 118. Ikawa T, Kawamoto H, Fujimoto S, Katsura Y. 1999. Commitment of common T/natural killer (NK) progenitors to unipotent T and NK progenitors in the murine fetal thymus revealed by a single progenitor assay. J. Exp. Med. 190:1617–26 119. Massari ME, Grant PA, Pray-Grant MG, Berger SL, Workman JL, Murre C. 1999. A conserved motif present in a class of helix-loop-helix proteins activates transcription by direct recruitment of the SAGA complex. Mol. Cell 4:63–73 120. Ikawa T, Kawamoto H, Wright LY, Murre C. 2004. Long-term cultured E2A-deﬁcient hematopoietic progenitor cells are pluripotent. Immunity 20:349–60 121. Anderson MK, Hernandez-Hoyos G, Diamond RA, Rothenberg EV. 1999. Precise developmental regulation of Ets family transcription factors during speciﬁcation and commitment to the T cell lineage. Development 126:3131–48 122. Barao I, Hudig D, Ascensao JL. 2003. IL-15-mediated induction of LFA-1 is a late step required for cytotoxic differentiation of human NK cells from CD34+ Lin- bone marrow cells. J. Immunol. 171:683–90 123. DeHart SL, Heikens MJ, Tsai S. 2005. Jagged2 promotes the development of natural killer cells and the establishment of functional natural killer cell lines. Blood 105:3521–27 124. Cooper MA, Bush JE, Fehniger TA, VanDeusen JB, Waite RE, et al. 2002. In vivo evidence for a dependence on interleukin 15 for survival of natural killer cells. Blood 100:3633–38 www.annualreviews.org • NK Cell Development

283

ARI

15 February 2006

17:51

125. Kennedy MK, Glaccum M, Brown SN, Butz EA, Viney JL, et al. 2000. Reversible defects in natural killer and memory CD8 T cell lineages in interleukin 15-deﬁcient mice. J. Exp. Med. 191:771–80 126. Ranson T, Vosshenrich CA, Corcuff E, Richard O, Muller W, Di Santo JP. 2003. IL-15 is an essential mediator of peripheral NK cell homeostasis. Blood 101:4887–93 127. Dubois S, Mariner J, Waldmann TA, Tagaya Y. 2002. IL-15Rα recycles and presents IL-15 in trans to neighboring cells. Immunity 17:537–47 128. Ogasawara K, Hida S, Azimi N, Tagaya Y, Sato T, et al. 1998. Requirement for IRF-1 in the microenvironment supporting development of natural killer cells. Nature 391:700–3. Erratum. 1998. Nature 392(6678):843 129. Taki S, Nakajima S, Ichikawa E, Saito T, Hida S. 2005. IFN regulatory factor-2 deﬁciency revealed a novel checkpoint critical for the generation of peripheral NK cells. J. Immunol. 174:6005–12 130. Kawamura T, Koka R, Ma A, Kumar V. 2003. Differential roles for IL-15R α-chain in NK cell development and Ly-49 induction. J. Immunol. 171:5085–90 131. Jamieson AM, Isnard P, Dorfman JR, Coles MC, Raulet DH. 2004. Turnover and proliferation of NK cells in steady state and lymphopenic conditions. J. Immunol. 172:864–70 132. Carson WE, Fehniger TA, Haldar S, Eckhert K, Lindemann MJ, et al. 1997. A potential role for interleukin-15 in the regulation of human natural killer cell survival. J. Clin. Invest. 99:937–43 133. Iizuka K, Chaplin DD, Wang Y, Wu Q, Pegg LE, et al. 1999. Requirement for membrane lymphotoxin in natural killer cell development. Proc. Natl. Acad. Sci. USA 96:6336–40 134. Ferlazzo G, Thomas D, Lin SL, Goodman K, Morandi B, et al. 2004. The abundant NK cells in human secondary lymphoid tissues require activation to express killer cell Ig-like receptors and become cytolytic. J. Immunol. 172:1455–62 135. Bloch-Queyrat C, Fondaneche MC, Chen R, Yin L, Relouzat F, et al. 2005. Regulation of natural cytotoxicity by the adaptor SAP and the Src-related kinase Fyn. J. Exp. Med. 202:181–92 136. Gilﬁllan S, Ho EL, Cella M, Yokoyama WM, Colonna M. 2002. NKG2D recruits two distinct adapters to trigger NK cell activation and costimulation. Nat. Immunol. 3:1150–55 137. Ogasawara K, Hamerman JA, Hsin H, Chikuma S, Bour-Jordan H, et al. 2003. Impairment of NK cell function by NKG2D modulation in NOD mice. Immunity 18:41–51 138. Wu J, Song Y, Bakker AB, Bauer S, Spies T, et al. 1999. An activating immunoreceptor complex formed by NKG2D and DAP10. Science 285:730–32 139. Lee KM, Forman JP, McNerney ME, Stepp S, Kuppireddi S, et al. 2005. Requirement of homotypic NK cell interactions through 2B4(CD244)/CD48 in the generation of NK effector functions. Blood. In press 140. Wilde JI, Watson SP. 2001. Regulation of phospholipase C γ isoforms in haematopoietic cells: why one, not the other? Cell. Signal. 13:691–701 141. Wang D, Feng J, Wen R, Marine JC, Sangster MY, et al. 2000. Phospholipase Cγ2 is essential in the functions of B cell and several Fc receptors. Immunity 13:25–35 141a. Caraux A, Kim N, Bell SE, Zompi S, Ranson T, et al. 2005. Phospholipase C-γ2 is essential for NK cell cytotoxicity and innate immunity to malignant and virally infected cells. Blood. In press 142. Ting AT, Karnitz LM, Schoon RA, Abraham RT, Leibson PJ. 1992. Fcγ receptor activation induces the tyrosine phosphorylation of both phospholipase C (PLC)-γ 1 and PLC-γ 2 in natural killer cells. J. Exp. Med. 176:1751–55

Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-09

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Di Santo

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143. Caraux A, Lu Q, Fernandez N, Di Santo JP, Raulet DH, Lemke G, et al. 2004. Receptor Tyrosine kinases of the Tyro 3 family play a critical role in natural killer cell differentiation. Presented at 8th Meet. Soc. Nat. Immun., The Netherlands 144. Crosier KE, Crosier PS. 1997. New insights into the control of cell growth; the role of the AxI family. Pathology 29:131–35 145. Townsend MJ, Weinmann AS, Matsuda JL, Salomon R, Farnham PJ, et al. 2004. Tbet regulates the terminal maturation and homeostasis of NK and Vα14i NKT cells. Immunity 20:477–94 146. Lohoff M, Duncan GS, Ferrick D, Mittrucker HW, Bischof S, et al. 2000. Deﬁciency in the transcription factor interferon regulatory factor (IRF)-2 leads to severely compromised development of natural killer and T helper type 1 cells. J. Exp. Med. 192:325–36 147. Burkett PR, Koka R, Chien M, Chai S, Boone DL, Ma A. 2004. Coordinate expression and trans presentation of interleukin (IL)-15Rα and IL-15 supports natural killer cell and memory CD8+ T cell homeostasis. J. Exp. Med. 200:825–34 148. Koka R, Burkett PR, Chien M, Chai S, Chan F, et al. 2003. Interleukin (IL)-15Rαdeﬁcient natural killer cells survive in normal but not IL-15Rα-deﬁcient mice. J. Exp. Med. 197:977–84 149. Schluns KS, Nowak EC, Cabrera-Hernandez A, Puddington L, Lefrancois L, Aguila HL. 2004. Distinct cell types control lymphoid subset development by means of IL-15 and IL-15 receptor α expression. Proc. Natl. Acad. Sci. USA 101:5616–21 150. Brady J, Hayakawa Y, Smyth MJ, Nutt SL. 2004. IL-21 induces the functional maturation of murine NK cells. J. Immunol. 172:2048–58 151. Ranson T, Vosshenrich CA, Corcuff E, Richard O, Laloux V, et al. 2003. IL-15 availability conditions homeostasis of peripheral natural killer T cells. Proc. Natl. Acad. Sci. USA 100:2663–68 152. Schmidt-Weber CB, Blaser K. 2004. Regulation and role of transforming growth factor-β in immune tolerance induction and inﬂammation. Curr. Opin. Immunol. 16:709– 16 153. Kulkarni AB, Huh CG, Becker D, Geiser A, Lyght M, et al. 1993. Transforming growth factor β 1 null mutation in mice causes excessive inﬂammatory response and early death. Proc. Natl. Acad. Sci. USA 90:770–74 154. Castriconi R, Cantoni C, Della Chiesa M, Vitale M, Marcenaro E, et al. 2003. Transforming growth factor β 1 inhibits expression of NKp30 and NKG2D receptors: consequences for the NK-mediated killing of dendritic cells. Proc. Natl. Acad. Sci. USA 100:4120–25 155. von Boehmer H. 2005. Mechanisms of suppression by suppressor T cells. Nat. Immunol. 6:338–44 156. Haller O, Wigzell H. 1977. Suppression of natural killer cell activity with radioactive strontium: effector cells are marrow dependent. J. Immunol. 118:1503–6 157. Seaman WE, Gindhart TD, Greenspan JS, Blackman MA, Talal N. 1979. Natural killer cells, bone, and the bone marrow: studies in estrogen-treated mice and in congenitally osteopetrotic (mi/mi) mice. J. Immunol. 122:2541–47 158. Manz RA, Thiel A, Radbruch A. 1997. Lifetime of plasma cells in the bone marrow. Nature 388:133–34 159. Mazo IB, Honczarenko M, Leung H, Cavanagh LL, Bonasio R, et al. 2005. Bone marrow is a major reservoir and site of recruitment for central memory CD8+ T cells. Immunity 22:259–70 www.annualreviews.org • NK Cell Development

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160. Garni-Wagner BA, Witte PL, Tutt MM, Kuziel WA, Tucker PW, et al. 1990. Natural killer cells in the thymus. Studies in mice with severe combined immune deﬁciency. J. Immunol. 144:796–803 161. Hansson M, Karre K, Kiessling R, Roder J, Andersson B, Hayry P. 1979. Natural NK-cell targets in the mouse thymus: characteristics of the sensitive cell population. J. Immunol. 123:765–71 162. Moffet-King A. 2002. Natural killer cells and pregnancy. Nat. Rev. Immunol. 2:656–63

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:257-286. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Development of Human Lymphoid Cells Bianca Blom and Hergen Spits Department of Cell Biology and Histology, Academic Medical Center, University of Amsterdam, Amsterdam 1105 AZ, The Netherlands; email: [email protected], [email protected]

Annu. Rev. Immunol. 2006. 24:287–320 First published online as a Review in Advance on December 12, 2005 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090612 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0287$20.00

Key Words human lymphocytes, T cells, B cells, NK cells, DC

Abstract The lymphocytes, T, B, and NK cells, and a proportion of dendritic cells (DCs) have a common developmental origin. Lymphocytes develop from hematopoietic stem cells via common lymphocyte and various lineage-restricted precursors. This review discusses the current knowledge of human lymphocyte development and the phenotypes and functions of the rare intermediate populations that together form the pathways of development into T, B, and NK cells and DCs. Clearly, development of hematopoietic cells is supported by cytokines. The studies of patients with genetic deﬁciencies in cytokine receptors that are discussed here have illuminated the importance of cytokines in lymphoid development. Lineage decisions are under control of transcription factors, and studies performed in the past decade have provided insight into transcriptional control of human lymphoid development, the results of which are summarized and discussed in this review.

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INTRODUCTION CLP: common lymphoid precursor

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All hematopoietic cells are derived from pluripotent hematopoietic stem cells (HSCs). These cells differentiate into mature hematopoietic cells through various intermediate cell types that are deﬁned by expression of cell surface antigens. Traditionally, investigators have assumed that the ﬁrst step in hematopoietic development is differentiation of HSCs into myeloid and lymphocyte precursors. Myeloid precursors differentiate into erythroid, megakaryocytic, and granulocytic/monocytic (GM) lineages, whereas lymphoid precursors develop into natural killer (NK), T, and B cells. A wealth of data support this concept. However, a recent study has put this model into question with the identiﬁcation of a precursor that possesses a combined GM and lymphoid precursor potential but is unable to develop into an erythrocyte or megakaryocyte (1). Most researchers agree that the lymphocyte T, B, and NK cells originate from a common precursor, generally referred to as a common lymphoid precursor (CLP), although some researchers maintain that T and B cells are not derived from a common precursor without myeloid precursor activity but instead that there are T/macrophage and B/macrophage precursors (2). The developmental status of dendritic cells (DCs) is more complicated. The DC lineages consist of several often not clearly deﬁned populations that seem to have a myeloid, lymphoid, or mixed lymphoid/myeloid origin. Most of our knowledge about lymphoid development stems from studies with mice. The obvious reason for the reliance on mice studies is the possibility of performing in vivo experiments to test precursor activities of certain cell populations. In addition, genetically modiﬁed mice have been instrumental not only in illuminating developmental pathways but also in elucidating mechanisms behind certain developmental cellular transitions. By contrast, most information on human hematopoietic development is derived

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from in vitro studies, although certain genetic abnormalities have greatly contributed to our understanding of some underlying mechanisms of human hematopoietic cell development. Studies on human hematopoiesis seem to be generally consistent with principles outlined in experimental models, but the cell surface phenotypes of human transitional cell populations are often different from those in the mouse. In this review, we discuss the current knowledge regarding human lymphoid cell development, in particular of T, B, and NK cells and DCs. We do not exhaustively review current information from mouse models, as excellent reviews have appeared recently (2a–d, 94, 160), but this information is used as reference to compare human and mouse lymphocyte development.

HUMAN LYMPHOID PRECURSORS The idea that hematopoiesis progresses by gradually limiting the developmental potential of precursors predicted that precursor cells exist with limited lymphocyte-restricted precursor potential. Indeed, precursor cells that are restricted to lymphoid lineages have been identiﬁed in both mice and humans. That all hematopoietic precursors in humans are present within a population of cells that express CD34 is well established (reviewed in 3), and this marker is useful in elucidating pathways in the development of particular hematopoietic lineages. In searching CD34+ human bone marrow cells for the presence of cells that express lymphocyte-restricted antigens, several research groups have found CD34+ cells expressing the lymphocyte markers terminal deoxynucleotidyl transferase (TdT) and CD10, and they speculate that these cells represent precursors of T and B cells (4, 5). Galy and coworkers (6) obtained experimental support for this idea by demonstrating that CD34+ CD10+ CD45RA+ cells obtained from fetal and adult bone marrow develop into CD33+ and CD1a+ CD33+ DCs and CD19+

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B cells, although these cells were unable to develop into CD14+ monocytes. Importantly, investigators found that 14% of single-sorted fetal bone marrow CD34+ CD10+ cells produced B cells, NK cells, and DCs. In addition, they showed that a limited number of CD34+ CD10+ cells injected into fragments of fetal thymus were able to develop into T cells, but technical limitations prevented testing T cell development on a clonal level (6). Mouse bone marrow CLP, as described for the ﬁrst time by Kondo et al. (7), express the IL-7Rα chain. Indeed, two groups reported that the IL-7Rα chain is expressed on a large proportion of human bone marrow CD34+ CD10+ cells, which were CD45RA and CD43 positive but negative for CD24 (8, 9). These cells expressed transcripts characteristic for the B cell lineage, such as Pax-5 and Igβ, and the T cell–associated transcripts GATA3 and pTα, and were able to differentiate into B cells and NK cells (8, 9). These characteristics, and the similarity of these cells with those deﬁned by Galy et al. (6), suggested that the CD10+ IL-7Rα + cells represent CLP. However, the number of NK cells recovered after four weeks culture of these cells with Sys-1 cells, a murine stromal cell line in the presence of stem cell factor (SCF), Flt3-L, IL-7, and IL-2, was much lower than that of B cells (9), and the T cell precursor activity of the CD34+ CD10+ IL-7Rα + bone marrow precursors was not tested, leaving open the possibility that these IL-7Rα + precursor cells were biased to the B cell lineage. This notion is supported by identiﬁcation of a very similar cell type in cord blood, which was shown to be B cell biased (10) (see below). Many studies have used CD7 as a marker for lymphoid progenitors because this marker is not expressed or is only weakly expressed on myeloid cells (11). An early study demonstrated that CD34+ CD7+ bone marrow cells contain NK cell precursors, but no information was provided about T cell, B cell, and DC developmental potential (12). More recently, a careful phenotypic and functional analysis of CD34+ bone marrow

cells revealed the presence of CD34+ lin− , CD7+ CD10− (2%), CD7− CD10+ (10%), and CD7+ CD10+ (0.3%) cells (13). Both the bone marrow CD7+ and CD10+ populations had NK and B cell precursor activities, but the B cell potential of CD34+ lin− CD10+ was much higher than that of the CD7+ cells. Because of the scarcity of the CD7+ CD10+ populations, the functional activity of those cells was not tested (13). Polymerase chain reaction (PCR) analysis of B cell– and NK cell–speciﬁc transcripts of these populations strongly suggested that the CD34+ lin− , CD7+ CD10− and CD7− CD10+ , are NK and B cell biased, respectively (13). The chemokine receptor CXCR4 is expressed on cells with restricted lymphoid precursor activities, and the CD34+ CXCR4+ cells can be further subdivided into IL-7Rα + and IL-7Rα − cells. Although the developmental potential of CD34+ CXCR4+ IL-7Rα − bone marrow cells has yet to be tested (14), these cells may be precursors of recently identiﬁed lymphoid precursors in neonatal cord blood, which were found to be IL7Rα − (10, 15). Hao et al. (15) identiﬁed CD34+ CD38lo CD45RA+ CD7+ CD10+ IL7Rα − cells in cord blood able to differentiate into NK cells, B cells, and CD1a+ DCs, and although the T cell potential of these cells was not tested, Hao et al. suggested that these were lymphoid precursors. Two other groups conﬁrmed that CD34+ CD7+ CD45RA+ cord blood cells have lymphoid potential but found that CD10 and CD7 tended not to be coexpressed on CD34+ CD45RA+ cells (10, 13). Interestingly, the CD34+ CD45RA+ CD7− CD10+ population expressed IL-7Rα, whereas IL7Rα was absent on CD34+ CD45RA+ CD7+ cells (10). Both CD34+ CD7+ IL-7Rα − and CD34+ CD10+ IL-7Rα + populations had lymphoid potential, but the CD34+ CD45RA+ CD7+ cells preferentially differentiated into T and NK cells, whereas the CD34+ CD45RA+ CD10+ population was biased to develop into B cells (10, 13). Although differentiation assays revealed

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pDC: plasmacytoid dendritic cell TSP: thymus seeding progenitor

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ETP: early thymic progenitor

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that both CD34+ CD45RA+ CD7+ and CD34+ CD45RA+ CD10+ populations are biased to lymphoid lineages and completely lack the ability to develop into erythrocytes and megakaryocytes, they possess some GM precursor potential (10). It is not clear whether all cells within the CD34+ CD45RA+ CD7+ population are multipotent or whether this population contains both GM and lymphoid precursors. More recently, Chicha et al. (16) reported that either CD34+ CD38+ CD10+ or CD34+ CD38+ CD7+ cord blood precursors can develop into B cells and plasmacytoid DCs (pDCs), but that they fail to develop into GM cells. This study did not show whether the CD34+ CD7+ cord blood cell precursors coexpress CD10 or not, and these authors did not use anti-CD45RA in separating their precursors (16), making it difﬁcult to compare their results with those of others (10, 15). The phenotype of lymphoid-restricted progenitors from cord blood and fetal and adult bone marrow is not yet ﬁrmly established. An important reason for this lies in the use by different groups of dissimilar antibodies against the same differentiation antigen or the same antibodies tagged with different ﬂuorochromes. For example, anti-CD7FITC (ﬂuorescein isothiocyanate) used by Haddad et al. (10) is much less sensitive than anti-CD7-PE (phyco erythrin) or Tricolor, and the use of CD7-FITC misses cells that do express CD7 but at lower levels (11). Another reason the phenotype has not been ﬁrmly established is the controversy concerning expression of CD38, which was absent on lymphoid precursors in one study (17) but expressed in two others (10, 16); this difference could be due to the use of anti-CD38 antibodies with different afﬁnities. With these caveats in mind, there is consensus that CD7 and CD10 deﬁne CD34+ CD45RA+ precursors biased to, but probably not fully committed to, lymphoid lineages (Figure 1). In addition, within the CD34+ CD45RA+ population both in bone marrow and cord blood, there are CD7+ CD10+ , Blom

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CD7+ CD10− , and CD7− CD10+ cells, although the existence and function of the very rare CD7+ CD10+ cells have yet to be conﬁrmed. It is tempting to speculate that CD34+ lin− CD45RA+ CD7+ CD10+ are the precursors of the CD7+ CD10− IL-7Rα − T/NK cell–biased and CD7− CD10+ IL7Rα + B cell–biased populations (10, 13), but this has to be proven by using various antibody preparations combined with functional, preferentially clonal assays.

DEVELOPMENT OF T CELLS Human Thymus Seeding Progenitor (TSP) and Early Thymic Progenitor (ETP) The transient developmental potential of thymic precursors demands that a continuous source of thymus seeding progenitors (TSP) can enter the thymus. Because the thymus remains active probably through an advanced age, TSP should be present in adult blood as well. As discussed in a previous section, CD34+ CD45RA+ CD7+ cells that have T, B, and NK and some GM precursor activities have been found in cord blood (10, 15). These cells resemble CD34+ CD38lo ETP with T cell, NK cell, and DC precursor activities (18) (Figure 2). An analysis of the T cell receptor (TCR) rearrangement status conﬁrms that the CD34+ CD38lo cells form the most immature population in the thymus (19), and it is tempting to speculate that CD34+ CD38lo thymocytes, which mostly coexpress CD10, are the direct progeny of the CD34+ CD45RA+ CD7+ CD10+ CD38lo cord blood cells (15). The presence of multipotential precursors in the human thymus indicates that T cell commitment takes place within this organ in line with data in the mouse (20). Recently, however, investigators argued that the murine thymus is seeded not only by multipotent precursors but also by precursor cells that are lineage restricted (21, 22). If that is also the case in humans, such cells should be present in cord

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HSC

CD34+CD45RA+ (CD7+CD10+)

MEP

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CLP

CMP

CD34+CD45RA–

CD34+CD45RA+ CD10 + IL-7Rα α+

CD7+

T

NK

Figure 1

pDC

GMP CD34+CD45RA+CD123+

B

cDC

blood. One study failed to ﬁnd evidence for the presence of TCR rearrangements in cord blood CD34+ cells (23), but another study reported the presence of complete TCRδ and partial TCRβ (Dβ-Jβ) rearrangements in CD34+ CD7+ cord blood precursors (24). This issue should therefore be readdressed by single-cell PCR analysis of the recently identiﬁed rare CD34+ CD45RA+ CD7+ cord blood population (10, 15). T cell–restricted precursors were convincingly identiﬁed in human bone marrow (25), but whether these cells can migrate to the thymus is unclear. CD34+ CD19+ B cell precursors are also present in the human thymus (26). Recently, it was reported that although CD34+ lin− cord blood cells could develop into B cells in a fetal thymic organ culture (FTOC) in the presence of Notch inhibitors (26a), CD34+ CD1a− thymocytes failed to do so, suggesting that the

pDC

cDC

mo

Gr

Model for development of CLP and GMP. CLP develop into pDCs and cDCs, but whether they derive from precursors that are CD7+ , CD10+ , or CD7+ CD10+ is not completely clear (Reference 16) (MEP, megakaryocyte/erythrocyte progenitor).

latter cells lack B cell precursor potential. Thus, the thymic B lineage cells (26) may be derived from CD34+ CD10+ IL-7Rα + cord blood progenitors that express a B cell transcript signature and are developmentally biased to the B cell lineage (10). In summary, at least a proportion of the precursors that seed the human thymus are multipotential. Whether or not some of the precursors that migrate into the thymus are lineage restricted before entrance is unclear.

Cellular Stages in the Development of ETP into CD4+ CD8+ Double-Positive Immature T Cells Over the past few decades, the various transitional stages of T cell development in the human thymus have been characterized

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Figure 2 Model for the earliest stages of development in the thymus. Evidence from in vitro experiments suggests that NK cells can develop not only from T/NK precursors but also from NK/DC myeloid precursors. On the basis of expression of pTα, CD2, CD5, and CD7, thymic pDCs may develop from ETP via a lymphoid pathway, but it is possible that the CD34+ CD44hi myeloid precursors can also develop into pDCs in the human thymus.

CD4 ISP: CD4 immature single-positive cell

292

CD34hi CD33lo CD38lo CD44+ IL-7Rα+

CD34– CD38hi CD44hi CD33– IL3Rαhi BDCA2+

CD34hi CD38+ CD1a– IL-7Rα+

pDC

pre-T/NK

ETP

NK

pre- cDC

CD34lo CD3+ CD44hi CD33+ GM-CSFR+

regarding phenotype and status of the TCR gene rearrangements (reviewed in 27, 28) (Figure 3). As discussed in the previous section, ETP are enclosed within the CD34+ CD1a− population. The downstream CD34+ CD1a+ population is committed to the T cell lineage because they are unable to develop into non-T cells (27, 29–31). A recent study has shown that the subsequent CD4 immature single-positive (CD4 ISP) (32) population can be divided into two subgroups: a CD4+ CD1a+ (CD4 ISPlo ) and a CD4hi CD1ahi (CD4 ISPhi ) population (S. Ligthart, Y. Yasuda, H. Spits, and B. Blom, manuscript submitted). Downstream of the CD4 ISP subset are CD4+ CD8α + β− (early double-positive) and CD4+ CD8α + β+ populations (29, 32), the latter cells being Blom

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CD34+ CD38+ CD1a+ IL-7Rα+ pre-T

CD34– CD38hi CD56+

cDC

THYMUS

the precursors of double-positive TCRαβ+ cells. During the early stages in T cell development, the TCR loci undergo rearrangement in a sequence TCRδ > γ > β > α (19, 33, 34); however, there is disagreement with respect to the cell type in which rearrangement of particular loci occurs, probably owing to differences in sensitivity of the methods used to analyze TCR rearrangements. For example, by using a PCR-based and GeneScanning analysis, some productive TCRβ V-DJ rearrangements were found in the CD34+ CD1a+ cells, although the sensitivity of this assay raises the possibility that a small contamination was responsible for this signal (19). A less-sensitive southern blot analysis detected the ﬁrst TCRβ V-DJ rearrangements in the CD4 ISP cells (33).

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icTCRβ+ TSP

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THYMUS

CD34

+

+

+

+

-

-

-

CD1a

-

-

+

+

+

+

+

icTCRβ

-

-

-

-

+

+/-

+

icCD3ε

?

+/-

+

+

+

+

+

CD2

-

+/-

+

+

+

+

+

CD5

-

+/-

+

+

+

+

+

CD7

+

+

+

+

+

+

+

CD4

-

-

-

+

+

+

+

CD8α

-

-

-

-

-

+

+

CD8β

-

-

-

-

-

-

+

CD45RA

+

+

-

-

-

-

-

CD45RO

-

-

-

-

+

+/-

+

IL-7Rα

-

+

+

+

+

+

+

DJβ

-

+

+

+

+

+

+

V-DJβ

-

-

-

+

+

+

+

V-Jα

-

-

-

-

-

-

+

Several lines of evidence indicate that selection for productive, in-frame TCRβ rearrangements and production of a TCRβ protein, a process referred to as β-selection, can occur in distinct populations of cells that differ in CD4 and CD8 expression. A very low frequency of productive TCRβ V-DJ rearrangements was detected in the CD34+ CD1a+ CD4− population, but expression of TCRβ protein was not analyzed (19).

Figure 3 Intermediate stages in development of early thymocyte subsets into double-positive cells. The TCR rearrangement status of the various subpopulations are based on data presented in References 33 and 34. However, it should be noted that a recent study (19) positions the onset of these rearrangements at earlier stages. This is discussed in the text. (Abbreviation: EDP, early double-positive.)

Nonetheless, these results suggest that a few cells may undergo β-selection before CD4 is expressed. However, not all populations downstream of the CD34+ CD1a+ CD4− cells are post-β-selection cells because intracytoplasmic (ic) TCRβ− populations have been found in the CD4 ISP cells (33; S. Ligthart, Y. Yasuda, H. Spits, and B. Blom, manuscript submitted) and in the CD4+ CD8α + β− early double-positive cells (33, 35). Ten to twenty

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percent of the CD4 ISP are icTCRβ+ . In contrast to the TCRβ− CD4 ISP, the icTCRβ+ CD4 ISP were in cycle; expressed elevated levels of CD1a, CD4, CD28, CD45RO and CD71; and differentiated into TCRαβ+ T cells in a FTOC with faster kinetics than the icTCRβ− CD4 ISP (S. Ligthart, Y. Yasuda, H. Spits, and B. Blom, manuscript submitted). These data indicate that β-selection can occur within the CD4 ISP population. On the basis of the results of our studies and those of others, we propose that expression of a TCRβ protein and the ensuing β-selection occur within a certain developmental window and are not tightly coupled to regulation of CD4, CD8α, and CD8β expression. Thus, a few cells already undergo β-selection before CD4 is expressed (19). A larger proportion is β-selected after upregulation of CD4 (33; S. Ligthart, Y. Yasuda, H. Spits, and B. Blom, manuscript submitted), and a third group of the pre-T cells upregulate CD4 and CD8α before initiating and completing TCR rearrangements (35, 36).

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The Role of Cytokines in Early T Cell Development The important role of IL-7 in T cell development is well documented. The IL-7 receptor consists of two chains, IL-7Rα and gamma common (γc), which is also part of the receptors for IL-2, IL-4, IL-9, IL-15, and IL-21. Genetic defects in the genes encoding for γc (37, 38), IL-7Rα (39, 40), or the Janus kinase Jak3, a component of the IL-7-induced signal transduction pathway (41, 42), account for the majority of severe combined immune deﬁciencies (SCID) characterized by strongly reduced numbers of T cells. The most frequent form of SCID is caused by mutations in the γc-encoding gene (reviewed in 43). In these patients, T and NK cells are absent, whereas in contrast to what is observed in γc-deﬁcient mice, B cell development is normal (37, 38, 43). IL-7Rα-deﬁcient patients also display a profound T cell deﬁciency and have near normal B cell numbers. In contrast to γc-deﬁcient 294

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patients, IL-7Rα-deﬁcient patients have normal frequencies of NK cells in the periphery (39, 40). These manifestations argue against an important role of IL-7 in survival and proliferation of lymphoid precursors. Rather, they indicate that human T cell development speciﬁcally and critically depends on IL-7. The precise function of IL-7 in human T cell development is not fully understood. Inhibition of IL-7R signaling by blocking anti-IL-7 and anti-IL-7R antibodies prevents expansion and differentiation of developing T cells in a FTOC (44, 45), indicating an important role of IL-7 in mediating survival and proliferation of human T cell precursors. This effect is mediated by IL-7-induced PI3K activation through one tyrosine residue at position 449 in the cytoplasmic tail of the IL-7Rα chain (45). In the mouse, IL-7 does not appear to be critical for differentiation of TCRαβ cells in the thymus, as near normal distributions of subsets were observed in the thymuses of IL-7Rα-deﬁcient mice (46). There is evidence, however, that IL-7 is important for differentiation of human T cells. First, differentiation of CD34+ precursors in a FTOC in the presence of anti-IL-7R antibody is almost completely blocked at the transition of CD34+ CD1a+ cells into CD4 ISP (44). Second, according to one report, thymocytes from γc-deﬁcient infants possess TCRβ D-J but lack V-DJβ rearrangements (47). Although a function of IL-7 in TCRβ rearrangements in human pre-T cells has yet to be conﬁrmed, this ﬁnding may explain the complete lack of T cells in many γc- and IL7Rα-deﬁcient patients (40, 48). On the other hand, IL-7 is required for peripheral T cell homeostasis in humans (49), raising the alternative possibility that the absolute T cell deﬁciency in γc-deﬁcient patients is caused by a combination of defects in early T cell development and in T cell homeostasis. Taken together, the data indicate that IL7 is indispensable for human T cell development. The data also suggest that IL-7 is more critical for human than for mouse T cell development, which may be attributed to a

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role for IL-7 in TCRβ V-DJ rearrangements, but more data are required to conﬁrm this speculation.

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Transcription Factors Involved in Human T Cell Development Recently, it has become clear that Notch receptor ligand interactions play crucial roles in T cell development. Notch receptors control differentiation and proliferation in response to ligands on neighboring cells (50). Interaction of Notch receptors (Notch1, 2, 3, 4) with one of their ligands (DL1, 2, 4, and Jagged-1, -2) results in proteolytical cleavage of its cytoplasmic portion (icNotch), which migrates to the nucleus and interacts with C promoter binding factor (CBF)-1/recombination signal–binding protein (RBP)-J to mediate transcription of target genes. In the absence of icNotch, RBP-J represses transcription by interacting with various corepressors. When icNotch binds to RBP-J, it recruits the coactivator mastermind-like (MAML)-1, which binds to icNotch in the icNotch-CSLDNA complex, thereby converting the CSL (CBF-1/suppressor of Hairless/Lag1) complex into a transcriptional activator. Radtke et al. (51) were the ﬁrst to demonstrate that conditional deletion of Notch1 resulted in a switch in development in that, in the thymus, T cell development was inhibited and B cell development stimulated (51). More recently, Jaleco et al. (52) demonstrated that coculture of human cord blood CD34+ cells with the murine stromal cell line S17 expressing the Notch ligand DL1 resulted in generation of CD7+ cells with strong NK cell precursor activity while inhibiting B cell development (52). The cells generated in this system expressed high levels of CD7 and cytoplasmic CD3ε and may represent T/NK precursors (52). These data were conﬁrmed and extended by Zuniga-Pﬂucker and collaborators (53), who documented that cord blood and bone marrow CD34+ cells cocultured with another DL1-expressing bone marrow stro-

mal cell line, OP9-DL1, IL-7, and Flt3-L, mediated full T cell development (53, 54), similar to murine HSCs (55). Recently, the function of Notch in development of T and non-T cells in a FTOC system (26a) and in the OP9-DL1 system (55a) was investigated using the γ-secretase inhibitor 7 (N-[N-(3,5-diﬂuorophenyl)-L-alanyl]-Sphenylglycinet-butyl ester) (DAPT), which prevents proteolytic cleavage of Notch, thereby inhibiting Notch signaling. As expected, DAPT strongly inhibited T cell development, whereas development of non-T cells was stimulated. As expected, inhibition of TCRαβ development by DAPT was rescued by forced expression of icNotch1 by retrovirus-mediated gene transfer (55a). TCRβ V-DJ but not D-Jβ rearrangements were strongly inhibited in cells that developed from CD34+ CD1a− thymocytes in a FTOC in the presence of DAPT (26a), as in the mouse (56). This is not, however, the only mechanism of inhibition of T cell development by Notch1 because introduction of TCRαβ-encoding cDNA by retrovirus-mediated gene transfer into CD34+ CD1a− fails to overcome the requirement of Notch for development of these precursors into T cells (N. Legrand & H. Spits, unpublished observations). These ﬁndings indicate that Notch is required for induction and maintenance of T cell speciﬁcation, as found previously in the mouse (57). We and others also observed that, in the presence of DAPT, thymic CD34+ CD1a− cells generate more NK cells, monocytic/DCs (26a), and pDCs (55a), strongly suggesting that Notch drives lineage decisions in thymus of humans, similar to what occurs in the mouse thymus (58). Two research groups have documented that ectopic expression of icNotch1 in CD34+ cells affected T cell development in an in vitro FTOC (59, 60). icNotch1 favored development of human TCRγδ above TCRαβ cells when expressed in precursor cells before the TCRβ selection checkpoint, suggesting that a

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γc: gamma common chain of the IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 receptor DL1: Delta-like 1

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HEB

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Ets-1

Id2

γδ TCRγδ

NK

Figure 4 Transcription factors involved in T and NK cell development. The model assumes that NK cells develop from common T/NK cell precursors. It is possible, as indicated in Figure 2, that NK cells can also develop from NK/DC precursors that are unable to develop into T cells. E2A (i.e., E47) blocks NK cell development. Because Id2 sequesters E47, ablation of Id2 leads to inhibition of NK cell development, which can be neutralized by ablation of E47 (B. Kee, personal communication).

strong Notch1 signal may promote TCRγδ at the expense of TCRαβ development (59, 60). These data contrast with earlier ﬁndings in the mouse system that indicate the reverse, that diminished Notch1 signaling favors TCRγδ above TCRαβ development (61). However, De Smedt et al. (59) found that CD34+ expressing icNotch1 developed into TCRαβ cells in the bone marrow, similarly to the effect found in the mouse, indicating that in an in vivo setting there is no preferential TCRγδ development from icNotch1 overexpressing CD34+ precursor cells. Thus, the role of Notch1 in TCRαβ and γδ diversiﬁcation in humans remains to be established. Other transcription factors that have been implicated in human T cell development are members of the basic helix-loop-helix (bHLH) subfamily of E-box binding (E) proteins (Figure 4). There are four E proteins, E12, E47, HEB, and E2-2, all of which are involved in both T and/or B cell development in the mouse (62). Ectopic expression

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of natural antagonists of the E proteins, the Id (inhibitors of DNA binding)-2 and Id3, in human CD34+ cells inhibits T cell development and promotes NK cell development in a FTOC (63, 64). Stimulation of NK cells by Id2 and Id3 overexpression is consistent with the fact that Id2−/− mice have few NK cells (65), and together these data indicate that the balance of Id and E proteins plays a role in T/NK lineage diversiﬁcation both in humans and mice. Interestingly, introduction of Id3 in CD4 ISP cells results in their inhibition of TCRαβ development, but not of TCRγδ development, in a FTOC (64), which could be reversed by coexpressing HEB (R. Schotte, Y. Yasuda, and H. Spits, manuscript submitted). Studies in the mouse have shown that HEB controls pTα expression (66, 67). These data, combined with the demonstration in mice that β-selection commits precursor cells to the TCRαβ lineage (68), suggest that HEB stimulates TCRαβ development by control of pTα expression (Figure 4).

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Another transcription factor that has been associated with various stages of T cell development is GATA3 (69). Taghon et al. (70) observed that overexpression of GATA3 in human CD34+ thymic precursors resulted initially in stimulation of development into CD4+ CD8+ cells in a FTOC, consistent with a role for GATA3 in early development. However, further development of GATA3overexpressing cells was compromised, as the absolute numbers of TCRαβ cells were severely reduced at later time points. Taghon et al. (70) also found a strongly reduced expression of TCRβ protein by GATA3 overexpression and suggested that deregulated expression of GATA3 interferes with appropriate TCRβ rearrangement or translation. However, these ﬁndings are difﬁcult to reconcile with observations of conditional GATA3deﬁcient mice that suggested that GATA3 is required for TCRβ expression and pre-TCR signaling (69). Experiments, yet to be performed, to test the effect of GATA3 knockdown on T cell development should reveal the role of this transcription factor in human T cell development. In summary, the role of Notch1 in human T cell development has been established. In addition, it is clear that E proteins are required for early T cell development. It remains to be investigated how Notch1 and E proteins are linked in the control of human T cell development. Also, it has yet to be established that GATA3 is essential for early T cell development in humans.

DEVELOPMENT OF NATURAL KILLER (NK) CELLS Upon their discovery, NK cells were operationally deﬁned as leukocytes able to kill transformed cells without prior sensitization. Today, NK cells are known to play essential roles in the innate immune response, as well as in the generation of an adaptive immune response. Initially, investigators believed that NK cells were related to other cells of the

innate immune system, such as monocytes; however, work in the 1990s established that NK cells are more closely related to T cells, with whom they can share a common precursor. NK cells are functionally similar in particular to CTL, as both types of cells have the capacity to kill other cells and to produce cytokines such as interferon (IFN)-γ. NK cells probably coevolved with other cell types of the lymphoid system, in particular with T cells, because both of these lymphocytes recognize conventional and nonconventional major histocompatibility complex (MHC) molecules. Functional MHC molecules are present in cartilaginous ﬁsh, but not in more primitive species. Similarly, NK cells, as they are currently deﬁned, have not been identiﬁed in species lower than ﬁsh (71). In humans, there are two major subsets of NK cells: one expressing high levels of CD56 and low or no CD16 (CD56hi CD16+/− ), and a second that is CD56+ CD16hi (72, 73). The distribution of the cells and the functional properties of these populations are different in that CD56hi CD16+/− cells have relatively lower cytolytic activity and produce more cytokines than the CD56+ CD16hi cells (72, 73). As discussed below, the developmental relationship between these populations has yet to be resolved.

Sites of NK Cell Development NK cells can develop at multiple sites. In the fetus, NK cell precursors have been found in the bone marrow (6), liver (74), thymus (75), spleen, lymph nodes (T. Cupedo & H. Spits, manuscript in preparation), and possibly in the intestine (76). However, identiﬁcation of an NK precursor does not prove that NK cells actively develop in those organs. Such proof would require in situ identiﬁcation of committed NK cell precursors and other downstream intermediate stages. Indirect evidence suggests that the population of fetal liver CD34+ CD38+ cells contains committed NK cell precursors

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because these cells have robust NK cell precursor activity but are unable to develop into T cells (74). The fetal thymus contains an immature CD34− CD5− CD56− population with clonogenic NK cell potential (75), as well as bipotential T/NK precursors (77). Together, these data suggest that the fetal liver and thymus are sites for NK cell development. The biological function of fetal NK cells is currently unknown. The fact that patients deﬁcient in γc, most of whom lack NK cells at birth, are born at full term suggests that these cells are not essential for the fetus during pregnancy. Whether the thymus is required for the development of human fetal NK cells is also questionable because infants with DiGeorge syndrome have normal numbers of NK cells (78). The presence of NK cells in the thymus may be a consequence of the fact that some bipotential T/NK precursors, which form obligatory intermediates in the development of multipotent precursors of T cells, by chance develop into NK cells and that these cells have no function in the thymus. The general consensus is that the bone marrow is the site for NK cell development in children and adults. One study in the mouse has identiﬁed a committed NK cell precursor, unable to develop into other lymphoid cells, in the mouse bone marrow (79), but the exact phenotype of the human bone marrow equivalent has yet to be determined. Whether NK cells fully develop and mature in the bone marrow before entering the circulation is unclear. A recent study suggests that human NK cells can develop and mature via another route. A CD34lo CD45RAhi cell that expresses high levels of the integrin α4β7 was identiﬁed in adult peripheral blood and in lymph nodes (80). These cells express c-kit, CD2, low levels of CD7, L-selectin, and IL-2Rα, but were CD10− and were able to develop into CD56hi CD16dim/− NK cells with high efﬁciency in vitro. Because of their location in the lymph nodes, these cells likely represent committed NK cell precursors, which

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implies that CD34lo CD45RAhi cells lack T cell-, pDC-, or B cell–differentiating activities, although this was not directly veriﬁed in this study. Therefore, proof that these cells are NK lineage–restricted is lacking (80). In the lymph nodes, CD34lo CD45RAhi α4β7+ cells are located within T cell–rich regions, where CD56hi cells also reside (81). Because CD34lo CD45RAhi lymph node cells could develop into CD56hi NK cells when cultured in IL-2, it was speculated that in vivo T cell activation in the lymph nodes may promote the development of new NK cells from these precursors (80). The study by Freud et al. (80) left unresolved the question of where the CD56+ CD16hi NK cells develop because the committed NK precursors in the lymph nodes did not give rise to CD56+ CD16hi precursors. However, results of another study suggest that highly cytolytic CD56+ CD16+ lymph node NK cells arise from IL-2-activated noncytolytic CD56hi CD16+/− cells (82). In addition, two research groups have reported that IL-21, in concert with Flt3-L, SCF, and IL-15, stimulated the generation of CD56+ CD16+ NK cells in vitro (83, 84), but whether this is a reﬂection of what happens in vivo is unknown.

Early Stages in NK Cell Development Experiments both in mouse and human systems have established that T and NK cells share a common precursor (75, 85–89; reviewed in 27, 90). Despite the strong evidence for a common origin of T and NK cells, there are data suggesting that NK cells can be descendants of myeloid precursors. Marquez et al. (91) demonstrated that CD34+ CD33lo thymocyte precursors, which are probably identical to ETP, have the capacity of developing into CD34+ CD44hi CD33+ myeloid precursors. These cells were unable to develop into T cells but contained bipotent NK/DC precursors (91). Furthermore, Perez et al. (92) recently reported that a

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CD14+ CD11b+ CD13+ CD33+ cell isolated from cord blood can develop into cytolytic CD56+ NK cells after incubation with Flt3L and IL-15. The frequency of NK precursors within the CD14+ cord blood population was low, however (1/50) (92), raising the possibility that the CD14+ cells were contaminated with CD56− NK precursors. The idea that NK cells can derive from myeloid precursors is not supported by studies that examined the lymphoid and myeloid precursor potential of CLP, common myeloid precursors (CMP), and granulocyte/monocyte precursors (GMP) both in humans and in mice. Rather, these studies concluded that NK cells develop predominantly from lymphoid precursors (7, 9, 16, 93). However, as putative human lymphoid precursors retain some GM potential and lack erythrocyte precursor activity (10), the issue of whether in humans there are bipotential precursors able to develop into NK cells and monocytes and/or DC, but not into T cells, remains to be settled. Although early cellular stages in human T and B cell development have been reasonably well deﬁned, our knowledge about early stages of human NK development is very limited. Roughly, three stages can be deﬁned

in NK cell development: lineage commitment, NK receptor repertoire selection, and functional maturation (94, 95). Only very recently has a putative committed NK precursor within the CD34+ cell compartment been identiﬁed (80). These CD34lo CD45RA+ α4β7hi CD7+/− CD10− NK precursors that give rise to CD56hi CD16− NK cells in vitro may be downstream from CD34+ CD45RA+ CD7+ CD10− (10) or CD34+ CD45RA+ CD7+ CD10+ (15) lymphocyte precursors. The immature NK cells developing from committed NK cell precursors are deﬁned by expression of CD161 (NKR-P1) (96). These cells do not express CD56 or CD16. Immature NK cells can be induced to express these markers as well as the activating and inhibitory receptors, CD94NKG2A and killer inhibitory receptors (KIR), upon culture with stromal cells and cytokines such as IL-15 (97) or Flt3-L (98). Figure 5 shows a hypothetical model of NK cell development that is deduced from phenotypic and functional analyses of small populations of immature NK cells isolated from different sources and from in vitro experiments (74, 98, 99; reviewed in 94). As discussed in the previous paragraph, the model, which is adapted

COMMITMENT

T/NK

CLP

precursor

CD34+ CD45RA+ CD7+ (CD10+)

CD34+ CD45RA+ CD7+ CD10 –

CMP: common myeloid precursor GMP: granulocyte/ monocyte precursor

MATURATION

NKp

NKi

NK

CD34lo CD45RA+ CD7lo CD10– CD122lo α4β7hi

CD34CD161+ (NKR-P1) CD16– CD56– CD122+

CD161+ CD56hi CD16 – ?

?

?

CD161+ CD56+ CD16+

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Figure 5 Model of NK cell development modiﬁed from Reference 94. Committed NK precursors develop into CD56hi CD16− NK cells (80), but the immediate precursor of the CD56+ CD16+ NK cells is not yet known. The dashed lines indicate the various possibilities. 299

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from Reference 94, does not resolve the developmental pathway of CD56hi CD16−/+ and CD56+ CD16hi subsets. There is no consensus whether CD56hi NK cells are precursors of CD56+ CD16hi NK cells. These subsets may arise from different precursors, but so far no evidence supports this speculation.

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A number of cytokines have been identiﬁed that can support development of human NK cells from CD34+ cells in vitro, notably SCF, Flt3-L, IL-7, IL-2, and IL-15. Data from mouse experiments indicate that SCF and Flt3-L may act on early lymphoid precursors, causing their development and thereby promoting NK cell development (reviewed in 94). However, the exact roles of SCF and Flt3-L in human NK cell development have not been determined. IL-2 and IL15 were previously proposed to be the most relevant factors for human NK cell development. There is now consensus that IL-15, and not IL-2, is the critical NK cell factor, a ﬁnding that is supported by the strongly reduced NK cell numbers in mice deﬁcient in IL-15 or in its receptor components, IL15Rα, β, and γc. SCID patients suffering γc deﬁciencies lack NK cells (43), whereas IL2-deﬁcient (100) and IL-2Rα-deﬁcient patients have normal numbers of NK cells (94, 101). One SCID patient, who presented with an absence of NK cells and reduced T cell numbers, was reported to have a strongly reduced expression of the IL-2/15Rβ chain and a marked decrease of signaling through this receptor (102). Data in the mouse suggest that IL-15 is not a differentiation factor but serves to maintain the viability and support proliferation of developing NK cells because NK cell precursors are present in normal numbers in mice deﬁcient for IL-15 or its receptor components, IL-15Rα, β, and γc (103). It is unknown whether NK precursors are present in γc-deﬁcient patients, partly be300

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cause our knowledge of early NK cell precursors in humans is limited. However, the limited availability of the patient’s tissues and the scarcity of NK precursors are more important roadblocks to resolving these questions. Another cytokine implicated in NK cell development and function is IL-21 (83), but this cytokine acts only in concert with others, including SCF, Flt3-L, and IL-15, and may play a role in later stages of NK development (84). Mice lacking IL-21 develop normal numbers of NK cells (104), which is consistent with the notion that this cytokine may be affecting the function, not development, of NK cells.

Transcription Factors Involved in NK Cell Development Our knowledge of transcription factors that are speciﬁcally involved in NK cell development is very limited compared with what we know about their roles in T and B cell development. Overexpression of Id2 and Id3 in human CD34+ cells blocks their development into T cells, B cells, and pDCs but by contrast promotes NK cell development (Figure 4). As discussed above, Id2 and Id3 are natural antagonists of E proteins. Id2 also controls NK development in mice: Id2deﬁcient mice lack NK cells and NK cell precursors (105). However, Id2 is not a transcription factor but rather sequesters E proteins, thereby preventing their activity (62). Thus, the ﬁndings that Id2 overexpression stimulates NK cell development and that Id2 ablation prevents this indicate that one or more of the E proteins inhibit NK cell development. Indeed, overexpression of E12 and E47 inhibits NK cell development in vitro (R. Schotte & H. Spits, manuscript in preparation). Moreover, introduction of E47 deﬁciency into Id2−/− mice rescues NK cell development (B. Kee, personal communication), indicating that Id2 stimulates NK cell development by sequestering the inhibitory factor E47. E47 is an example of a factor that is required for the development of one lineage

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(B cells) but that inhibits alternative lineage choices. Another example is Spi-B, which is required for pDC development yet inhibits the development of NK cells (106). The role of Notch1, which drives T cell commitment, is complicated in NK cell development. Triggering Notch by DL1 drives human CD34+ cord blood cells to T/NK cell precursors (52), but inhibition of Notch signaling in a FTOC system that allows for both T and NK cell development inhibits T and stimulates NK cell development (26a). A similar observation has been made in experiments that examined the effect of γ-secretase inhibitor on rat thymus development (107). Thus, although Notch signaling may be required for the early steps in the development of HSCs into NK cells, it may not be required in later stages. A master transcription factor for NK cells has not been identiﬁed. One possible candidate is Ets-1, which is predominantly expressed in lymphoid cells (108). Ets-1deﬁcient mice have reduced numbers of NK cells, and the few NK cells present in these mice are nonfunctional (109). No follow-up studies have been reported, and there is no information about the stage in which NK development is inhibited. Moreover, the role of Ets-1 in human NK cell development has not been investigated.

DEVELOPMENT OF DENDRITIC CELLS Several types of DCs, including interstitial DCs, Langerhans DCs, blood DCs, and pDCs, have been identiﬁed that, depending on their local microenvironment, mediate different types of immune responses (110, 111). The remarkable versatility and ﬂexibility of DCs, instructed by the priming signals from microbial and tissue-derived factors, distinguish them from other cell types in the immune system that exhibit more limited functions and suggests that DCs may play a central role in integrating the innate and adaptive aspects of various immune responses. In the mouse, there are

three main DC subsets: CD8α − DCs, CD8α + DCs, and B220+ Ly6C+ pDCs. The relationship between the CD8α + and CD8α − subsets is unclear, but for the sake of simplicity these cells are often categorized as conventional DCs (cDCs). The B220+ Ly6C+ pDCs, also referred to as IFN-producing cells (IPCs), are clearly a distinct subset. Human cDCs are HLA-DR+ cells that express high levels of CD11c and consist of a major BDCA3− and a minor BDCA3+ population (112). Human HLA-DR+ pDCs are deﬁned by absence of CD11c expression and by high levels of CD123 (the IL-3Rα chain) and BDCA2 (111). The CD11c+ HLADR+ BDCA3− population can be further subdivided into CD16+ and CD16− populations (112). The developmental and functional relationships of the multiple HLA-DR+ DC populations remain unclear. For the remainder of this review, we use the terms cDC and pDC to denote populations expressing CD11c and CD123, respectively.

cDC: conventional dendritic cell

Developmental Origin of cDCs and pDCs Galy and coworkers (6) showed that a fraction of bone marrow CD34+ CD45RA+ CD10+ lymphoid progenitors could develop into CD1a+ CD14− DCs. Later work of the Galy group showed that the in vitro conditions that induced development of CD1a+ CD14− DCs failed to induce differentiation of myeloid cells into cDCs, suggesting that development of DCs from lymphoid or myeloid precursors has different signaling requirements (113). The underlying mechanisms have not been elucidated. Other groups have shown that lymphoid-restricted progenitors from cord blood (CD34+ CD38− CD45RA+ CD7+ ) when cultured in standard conditions (SCF/Flt3-L/GM-CSF/TNF-α) (114) or lymphoid conditions (S17 with IL-3, IL-6, c-kit, Flt3-L) (15) were able to give rise to a similar type of DC (CD1a+ CD14− ). Because some of these CD1a+ DCs expressed S100 and low levels of CD207/Langerin,

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Canque et al. (114) suggested that they are Langerhans DCs. More recently, Manz and coworkers (16) tested cord blood CLP, CMP (CD34+ CD38+ CD123+ CD45RA− ), and GMP (CD34+ CD38+ CD123+ CD45RA+ ) for cDC precursor activity and conﬁrmed that cDCs (CD11c+ CD123lo ) can arise from CLP, CMP, and GMP. However, they also noted that the frequency of cDC precursors in CMP and GMP was higher than in CLP (16). As is discussed below, this conclusion is valid only if one assumes that the tested populations are homogeneous. Information about the early stages in development of human HSCs into pDCs is accumulating. We have postulated that the direct progenitor of pDCs is contained within the CD34lo compartment of cord blood, fetal liver, and bone marrow. These progenitors coexpress CD45RA, CD4, and high levels of CD123, and so we have denoted these cells pro-pDCs (for progenitor of pDCs) based on their phenotypic and functional similarities with pDCs (115). The pro-pDCs may be restricted to the pDC lineage, but whether the pro-pDCs lack potential to develop into other lineages has not formally been tested. In early studies, several research groups found that pDCs express lymphoid-related genes, including CD2, CD5, CD7, pTα, Spi-B, and λ5; contain IgH D-J rearrangements (116– 120); and lack myeloid-associated markers, including CD11c, CD11b, CD13, CD33, and Mannose receptor (121). On the basis of these ﬁndings, it was postulated that pDCs derive from lymphoid progenitors. This idea was tested recently by Chicha et al. (16). They observed that single cells among CD34+ CD7+ and/or CD34+ CD10+ cells from cord blood considered to be lymphoid precursors were able to develop into pDCs, cDCs (CD11c+ CD123lo ), and B cells (16). Cord blood–derived CMP and GMP (or CD34+ CD38+ CD123+ CMP/GMP together when tested in parallel with CLP) also gave rise to both pDCs and cDCs but not to B cells (16). By using a limiting dilution assay, Chicha et al. observed that sin-

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gle cells with pDC and cDC potential are more frequent within CMP/GMP (16) than within CLP. These results were taken as evidence that human pDCs are predominantly myeloid derived (16). However, in this study (16) lymphoid precursors were not puriﬁed on the basis of CD45RA expression, ignoring that a proportion of CD34+ CD7+ cord blood cells lacks expression of CD45RA and is unable to develop into NK cells (114). Therefore, these CD34+ CD7+ CD45RA− cells cannot qualify as lymphoid precursors. Nonetheless, it seems likely that, akin to cDCs, human pDCs can develop from both lymphoid- and myeloid-restricted precursors, which is consistent with what was observed earlier in mouse models (9, 16, 122–124). Whether lymphoid-derived cDCs and pDCs are identical to their myeloid-derived counterparts remains unclear. Relative differences in levels of expression of pDC surface antigens (114) and pTα and Spi-B transcripts (16) have been observed, but none of these markers was discriminating. There are data indicating that different subsets of pDCs and cDCs exist in vivo. Differences between thymic and peripheral pDCs have been noted in expression of CD2, CD5, and CD7 (125, 126). Mutually exclusive pDC subsets were identiﬁed in Flt3-L-treated healthy volunteers (127). In their blood, CD7+ CD56− and CD7− CD56− pDCs express pTα, whereas CD56+ CD7− pDCs lack pTα expression (127). In addition, several distinct cDC subsets have been described (112). Whether the various cDC and pDC subsets, most of which are poorly characterized, have different developmental origins is, however, unclear.

Growth Factors Involved in pDC Development It has now been well established that Flt3-L is important for human, mouse, and rhesus monkey DC development in vitro (115, 128, 129) and in vivo (130–134). Mouse pDCs can be generated only from Flt3+ CLP or Flt3+ CMP (135, 136). In humans, Flt3 mRNA has

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been observed in both CLP and CMP populations (9, 16). Moreover, CD34+ CD45RA− early progenitors are responsive to Flt3-L alone, without feeder cells to support pDC development, conﬁrming Flt3 cell surface expression on at least some of these cells (115). The observation that Flt3-L-deﬁcient mice were severely depleted of all DC subsets underscores the signiﬁcance of Flt3-L in DC development (137). Moreover, mice deﬁcient for STAT3, which mediates the checkpoint of Flt3-L but not GM-CSF-induced DC development, and mice lacking Gﬁ1, a zinc ﬁnger repressor molecule that regulates levels of STAT3, demonstrate a severe block in DC development (138, 139). Development of cDCs and pDCs can be differentially regulated by various cytokines. For example, cytokines that promote cDC development, including GM-CSF and TNF-α, have a strong inhibitory effect on the Flt3-L-induced generation of both human and mouse pDCs (115, 128). Thrombopoietin, which alone does not induce pDC development, enhances the effect of Flt3-L on human pDC development (140). Another cytokine, G-CSF, signiﬁcantly increased the number of blood pDCs when injected into healthy volunteers (134). Because G-CSF alone did not induce pDC development from HSCs (115), these data suggest that it increases pDC levels by stimulating mobilization of pDC or its progenitors from the bone marrow.

Transcriptional Control of Human DC Development Several transcription factors involved in differentiation of DCs have now been identiﬁed. These include Ikaros (141) and members of the Rel family of transcription factors (142). Information about transcription factors in development of cDCs in humans is scarce. Wu and collaborators (141) observed that mice homozygous for an Ikaros dominantnegative mutation that lack all cells of lymphoid origin, including T, B, and NK cells, also show a defect in development of a sub-

set of cDCs, presumably lymphoid-derived cDCs. Consistent with these observations, Galy et al. (113) reported that ectopic expression of this dominant-negative form of Ikaros interfered with development of lymphoid but not of myeloid cDCs. These ﬁndings suggest that Ikaros is involved in development of lymphoid-derived cDCs in both humans and mice. More is known about the role of transcription factors in human pDC development (Figure 6). CD34+ hematopoietic precursor cells ectopically expressing either Id2 or Id3 are blocked in their development into pDCs but not into myeloid cells (126). These ﬁndings are in line with observations in mice lacking Id2, which have increased numbers of pDCs (143). As discussed above, Id proteins are not transcription factors but act by sequestering E proteins. It remains to be elucidated, however, which member(s) of the bHLH family is (are) required for pDC development. We recently found that the Ets family member Spi-B is expressed in pDCs and its mature progeny but not in cDCs (119, 144), a ﬁnding that has been conﬁrmed in the mouse (118). Moreover Spi-B transcripts have been observed in in vitro CLP-/CMPderived pDC, but not in other DC subsets (16). Spi-B is involved in development of human pDCs because downregulation of Spi-B by RNA interference in human CD34+ progenitor cells compromises their development into pDCs in vitro and in vivo (145). In contrast, knockdown of Spi-B stimulates myeloid development (145). RNA interference experiments further indicate that another Ets family transcription factor, PU.1, which is highly homologous to Spi-B, is needed for development of both pDCs and CD14+ myeloid cells (145). These data suggest that PU.1 acts at an earlier stage of pDC development than does Spi-B, which is consistent with data in the mouse that show that PU.1 is required for development and survival of myeloid and lymphoid precursors (146) (Figure 6). Studies in mice have shown that pDC development also depends on members of the

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Figure 6 Transcription factors involved in pDC development. The Ets transcription factor Spi-B is involved in development of human pDCs, as shown by the fact that its ablation results in inhibition of human pDCs both in vivo and in vitro (145). Ectopic expression of Spi-B results in inhibition of T, B, and NK cell development. Knocking down Spi-B leads to an increase in B cell and monocyte development. These ﬁndings suggest that Spi-B can also play a role in regulation of pDC/monocyte diversiﬁcation. Based on inhibition of pDC development by Id2 or Id3 (126), an E protein should be involved in pDC development, but the nature of this E protein is unknown. (Abbreviation: EBF, early B cell factor.)

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IFN regulatory factor (IRF) family, including IRF-8 [or IFN consensus sequence-binding protein (ICSBP)] and to a lesser degree IRF-4 (147, 148). To date no studies have been performed to determine their role in human pDC development. Interestingly, IRF-8, in addition to binding to other members of the IRF family, can also bind to Spi-B and to members of the bHLH family to form transcriptional complexes apparently critical for the regulation of the immune system (149–151). Therefore, it is tempting to speculate that pDC development is driven by a complex of IRF-8, Spi-B, and a bHLH protein. Recently, the role of Notch signaling in development of pDCs was investigated. In agreement with an observation by Kincade’s group that interaction of DL1 with Notch inhibited pDC development from murine bone marrow precursor cells in vitro (118), we Blom

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found that DL1 inhibited pDC development both from thymic and fetal liver CD34+ cell precursors (55a). Inhibition of pDC development of thymic precursors by Notch signaling could be overcome by overexpressing SpiB. Thus, Notch, in concert with Spi-B, may regulate T/pDC lineage diversiﬁcation in the thymus.

DEVELOPMENT OF B CELLS Most hematopoietic cell lineages, including B cells, are believed to originate in the bone marrow. As in mice, all cellular stages of early B cell development in humans can be detected in fetal bone marrow as well as in bone marrow of young and aged adults (see below), indicating that B cell development proceeds throughout life (152). Early in ontogeny, B cell development may take place

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in the omentum (153) and fetal liver. The presence of early B cell–biased precursors in cord blood was recently demonstrated (10, 154, 155), and, even after birth, B cells may develop at sites other than the bone marrow. Notably, pre-B cells and mature B cells have been shown to be present in the thymus of young children (26).

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Early Stages of B Cell Development In the early 1990s, Hardy et al. (156) introduced a differentiation pathway for B220+ progenitors in mouse bone marrow. They showed, on the basis of differential surface expression of CD43, BP-1, and HSA, that B220+ progenitors develop via pre-pro-B, early pro-B, late pro-B, and pre-B into B cells. Functionally similar populations of early, pro-, and pre-B cells have been identiﬁed in humans, but the transition of common lymphocytes into B cell–restricted or –committed B cell precursors is as yet poorly deﬁned. The current consensus is that human B lineage–restricted cells pass through a CD34+ CD19− CD10+ early B, CD34+ CD19+ CD10+ pro-B, large CD34+ CD19+ CD10+ pre-BI, large CD34− CD19+ CD10+ pre-BII, small CD34− CD19+ CD10+ pre-BII cell development pathway (Figure 7). As discussed above, human CLP appear to be present in CD34+ CD45RA+ cells that express CXCR4 and that also express CD10 or CD7, or both (6, 10, 13–15). The CD34+ CD45RA+ CD10+ CD7− cells express IL-7Rα, and although these cells, found in bone marrow as well as cord blood, contain precursors able to develop into other lymphoid lineages, they are biased to develop into B cells (8, 10). Other studies have documented that early B cells, which express CD34, CD10, and IL-7Rα but lack CD19, have initiated DJH rearrangements (157) and express cytoplasmic CD79a (158) and Vpre-B (159). Although these studies suggest the initiation of B cell commitment in a CD34+ CD45RA+ CD10+ IL-7Rα + cell

population, other studies have found cells with B cell characteristics in a small fraction of bone marrow and cord blood, CD34+ cells that lacked CD10 and CD19 but that expressed CD79a transcripts (3.3%) and protein (0.4%) (154, 155). In addition, upon in vitro culture of CD34+ CD10− CD19− cord blood progenitor cells with mouse stromal cells and cytokines (SCF, IL-2, IL-15), these cells ﬁrst acquire CD79a and IL-7Rα before CD10 and CD19 (154). These CD34+ CD79a+ IL-7Rα + CD10− CD19− cells had DJH but no VDJH rearrangements and faintly expressed Pax-5 transcripts, which is considered the hallmark of B cell commitment in the mouse (160). Although these ﬁndings suggest that CD79a+ IL-7Rα + cells are committed to the B cell lineage, in vitro–generated CD79a+ IL-7Rα + pro-B cells were not restricted to the B cell lineage but could still differentiate into macrophages, NK cells, and some T cells in vitro (154). The developmental potential of freshly puriﬁed CD34+ CD79a+ CD10− CD19− cord blood cells remains to be assessed. However, considering that the possibly more differentiated CD34+ CD19− CD10+ cord blood cells still have B/NK potential and even retained some myeloid-erythroid potential (15), it seems fair to assume that CD34+ CD79a+ CD10− CD19− cells are multipotential. A developmental pathway of human preB cell subpopulations in bone marrow was proposed by Ghia et al. (161), who used single-cell PCR analyses to determine the expression of TdT and RAG (recombination activating gene)-1 transcripts, Vpre-B protein, the cycling status, and the conﬁguration of the IgH and IgL chain alleles in human B lineage subpopulations (reviewed in 162) (Figure 7). Human pro-B cells in the bone marrow are a well-characterized population expressing CD34, CD10, and CD19 (163). The vast majority of pro-B cells express TdT (161, 163, 164), some express Vpre-B at the cell surface (159, 161), and V-DJH rearrangements are easily detected (157, 165). Only a

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fraction of the pro-B cells express cytoplasmic μHC (161, 166, 167). Recently, a subpopulation (20%) of CD34+ CD19+ bone marrow cells was described that lack CXCR4 (168), which is the receptor for the chemokine CXCL12 (SDF-1). CD34+ CD19+ CXCR4− cells rapidly express CXCR4 and differentiate into mature κ or λ positive B cells in vitro (PHA-stimulated PBL–conditioned medium, SDF-1, IL-6). Surprisingly, however, a fraction of these cells (12%) were also able to form myeloid colonies containing granulocytes, macrophages, or erythrocytes (168). 306

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If we assume that human CD19 expression, like in the mouse, is dependent on Pax-5 (169, 170), which restricts the developmental options of early progenitors to the B cell pathway (160), these ﬁndings suggest that lineagerestricted pro-B cells retain a certain plasticity and may develop into either B cells or myeloid cells, depending on the organism’s requirements and subsequent bone marrow microenvironmental cues. Alternatively, the observed myeloid differentiation potential of CD34+ CD10+ CD19+ bone marrow cells in the study by Hou et al. (168) may derive from

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cells that lack Pax-5 expression. In line with this, Sanz et al. (155) showed that in cord blood 35% of CD34+ CD10+ CD19+ cells do not express Pax-5. Although this remains to be conﬁrmed in bone marrow cells, it will be interesting to determine how CD19 expression is regulated in the absence of Pax-5 in these cells. Pro-B cell differentiation into pre-BI cells is characterized by loss of CD34 and TdT and by acquisition of cytoplasmic μHC in more than 95% of the cells (161, 163, 164). Based on cell-cycle analysis, human pre-B cells can be generally subdivided into large proliferating pre-BI cells, large proliferating pre-BII cells, and small resting pre-BII cells (161). Pre-BI cells that bear in-frame productive (V-DJ)H rearrangements transport the μH chains to the membrane associated with L (Vpre-B and λ5) and CD79 to form the pre-B cell receptor, which signals several rounds of division. The large cycling pre-BI cells that express functional μH downregulate CD34, TdT, RAG-1, and RAG-2 and maintain expression of L, CD10, and CD19 genes. To allow for Ig κ or λ L gene recombination, RAG-1 and RAG-2 expression are reinduced in small resting CD34− CD19+ CD10+ pre-BII cells.

Growth Factors Involved in B Cell Development In mice, the c-kit, Flt3, and IL-7R signaling systems together account for the generation of all B lymphocytes in the bone marrow of adult mice (160, 171). In conspicuous contrast to what is known about the role of cytokines in mouse B cell development, the essential growth factors required for the growth of normal human B cell precursors remain elusive, although in vitro studies do suggest some function of c-kit, Flt3, and IL-7 in proliferation and survival of early pro-B cells (167, 172; reviewed in 162). However, in contrast to the well-documented requirement of IL-7 for mouse B cell development, this cytokine is not needed for human B cell development

in vitro (173) or in vivo (39, 40). Data obtained from SCID patients lacking expression of either IL-7Rα (39, 40), γc (37), or Jak3 (42, 48, 174), which are all components involved in the IL-7R signaling pathway, have normal or even increased levels of circulating B cells, although their function is greatly impaired (175). The presence of B cells in IL-7Rαdeﬁcient patients also excludes an important role of TSLP (thymic stromal-derived lymphopoietin) in human B cell development because its receptor consists of the IL-7Rα chain, in addition to a speciﬁc γc-homolog, TSLP-R. Moreover, the addition of TSLP has no effect on in vitro development of human CD34+ cells cocultured with the murine stromal cell line S17 (B. Blom, unpublished data).

Transcriptional Control of Human B Cell Development From mice studies, a wealth of information is available on the transcriptional control of early B cell commitment and development, which has been extensively reviewed by Busslinger (160). Development of HSCs via lymphoid progenitors into early B cells requires the concerted actions of multiple transcription factors, including Ikaros, PU.1, E2A, early B cell factor (EBF), and Pax-5. PU.1 and Ikaros act in parallel pathways to control transition of HSCs into lymphoid precursors. E2A, EBF, and Pax-5 regulate development of CLP into early B cells. E2A proteins are assumed to control EBF, which in turn regulates Pax-5. Not surprisingly, the limited information about expression of transcription factors in precursor cell populations and transcriptional control of human B cell development suggests that data obtained in murine systems can be largely extrapolated to the human situation. CD34+ Lin− CD10− CD7− cord blood cells, like mouse progenitor cells (146, 176), express the Ets family member PU.1 (154). Knockdown of PU.1 in human CD34+ CD38− fetal liver progenitors

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results in inhibition of B cell, monocyte, and pDC development, indicating a role for this transcription factor in development of both myeloid and lymphoid precursors (145). Multipotent CD34+ lin− CD10− CD7− cord blood cells lack expression of Pax5 and EBF (154, 177), whereas early B cells (CD34+ lin− CD10+ CD19− CD7− ) from adult bone marrow were reported to have markedly upregulated expression of Pax-5 relative to the more primitive CD34+ lin− CD10− CD7− subset or CD34+ lin− CD10− CD7+ NK-/T-biased precursors (13) and pro-B cells (CD34+ CD19+ ) in cord blood (155). These ﬁndings correlate well with observations in the mouse, where CLP already express transcripts of various lineage-speciﬁc transcription factors, including Pax-5. Indirect evidence for a role for EBF in human B cell development comes from ﬁndings that early hematopoietic zinc ﬁnger (EHZF, the mouse homolog of Evi3), which inhibits the transcriptional activity of EBF, is highly expressed in CD34+ cells but absent in CD19+ B cells (178). A study by Jaleco et al. (179) strongly suggests that E2A proteins are required for human B cell development because forced expression of one of their antagonists, Id3, strongly inhibits B cell development in vitro. Interestingly, following ectopic expression of Id3 in CD34+ CD38− CD10− fetal liver cells, their development into CD10+ IL-7Rα + cells was almost completely inhibited (179). These data indicate that E2A proteins are required for generation and/or survival of CD10+ IL-7Rα + B cell–biased precursors (8, 10). These ﬁndings are in accord with the observation that expression of another member of the Id family, Id1, is strongly

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reduced in CD34+ CD10+ lin− Pax-5+ EBF+ cord blood progenitor cells (13).

CONCLUDING REMARKS Our understanding of human lymphocyte development has increased signiﬁcantly over the past 20 years. In particular, our understanding of human T and B cell development has improved. Nonetheless, there are many gaps in our knowledge, particularly regarding the early stages of development of HSCs into lymphoid-restricted precursors. Much remains to be learned about the phenotype of human CLP and downstream lineage-biased and -restricted precursors. Despite enormous progress in our understanding of the function of NK cells and the mechanisms underlying their activities, our knowledge about the developmental pathways and the mechanisms of development of human NK cells is still surprisingly limited. The same holds true for our knowledge of DC development. One problem is that the different subsets of cDCs and pDCs are still poorly deﬁned. Future studies in the ﬁeld of human lymphocyte development will undoubtedly make use of recently described mouse strains that support development of all human lymphocyte classes (180–182). The possibility of stably expressing siRNA-encoding DNA fragments in human precursor cells, which is transferred to the mature progeny in an in vivo setting (181), allows for mechanistic studies in the ﬁeld of human hematopoietic development that were not possible until recently. We may also expect that future investigations in patients with deﬁned deﬁciencies in lymphocyte development will yield valuable information about the underlying mechanisms.

SUMMARY POINTS 1. CLPs coexpress CD34, CD45RA, and CD7 but are still poorly deﬁned. 2. T cell development in the human thymus occurs via a series of well-deﬁned intermediate stages.

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3. T cells require IL-7 and NK cells require IL-15 for their development. The importance of IL-7 in human T cell development may be more stringent than it is in mice. 4. Our understanding of early stages in NK cell development is improving, but the identity of the earliest NK-restricted precursor is not yet ﬁrmly established. The developmental relationship of the two major human NK cell subsets, CD56hi CD16− and CD56+ CD16hi , remains unresolved.

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5. cDCs and pDCs can develop from both lymphoid and myeloid precursors. There are multiple subsets of cDCs and pDCs, but the developmental relationship of those subsets is as yet unknown. 6. Development of lymphocytes is driven by transcription factors. The essential role of Notch1 in human T cell development has been demonstrated. bHLH factors belonging to the E protein family control T cell, B cell, and pDC development and negatively regulate NK cell development. Spi-B, probably in concert with other factors, such as IRF-8 and members of the bHLH E protein family, is indispensable for pDC development. 7. We have a clear understanding of early stages of human B cell development. It is completely unknown, however, which cytokine(s) is (are) driving human B cell development. Although IL-7 is essential for B cell development in mice, it is not required for B cell development in humans.

FUTURE ISSUES TO BE RESOLVED 1. We need to develop a uniform deﬁnition of human pluripotent stem cells and CLPs and to determine their precursor potential on a clonal level. 2. Improved mouse models are needed to test the developmental potential of human lymphocyte precursors in an vivo setting. 3. We need improved methods that completely knockdown genes in primary hematopoietic precursors to elucidate the roles of key genes in human lymphoid development. 4. The developmental origin and function of various NK cell and DC subsets need to be elucidated.

LITERATURE CITED 1. Adolfsson J, Mansson R, Buza-Vidas N, Hultquist A, Liuba K, et al. 2005. Identiﬁcation of Flt3+ lympho-myeloid stem cells lacking erythro-megakaryocytic potential: a revised road map for adult blood lineage commitment. Cell 121:295–306 2. Katsura Y. 2002. Redeﬁnition of lymphoid progenitors. Nat. Rev. Immunol. 2:127–32 2a. Maillard I, Fang T, Pear WS. 2005. Regulation of lymphoid development, differentiation, and function by the Notch pathway. Annu. Rev. Immunol. 23:945–74 2b. Murre C. 2005. Helix-loop-helix proteins and lymphocyte development. Nat. Immunol. 6(11):1079–86 www.annualreviews.org • Development of Human Lymphoid Cells

309

ARI

15 February 2006

21:28

2c. Radtke F, Wilson A, Mancini SJ, MacDonald HR. 2004. Notch regulation of lymphocyte development and function. Nat. Immunol. 5(3):247–53 2d. Yokoyama WM, Kim S, French AR. 2004. The dynamic life of natural killer cells. Annu. Rev. Immunol. 22:405–29 3. Payne KJ, Crooks GM. 2002. Human hematopoietic lineage commitment. Immunol. Rev. 187:48–64 4. Gore SD, Kastan MB, Civin CI. 1991. Normal human bone marrow precursors that express terminal deoxynucleotidyl transferase include T-cell precursors and possible lymphoid stem cells. Blood 77:1681–90 5. Terstappen LW, Huang S, Picker LJ. 1992. Flow cytometric assessment of human T-cell differentiation in thymus and bone marrow. Blood 79:666–77 6. Galy A, Travis M, Cen D, Chen B. 1995. Human T, B, natural killer, and dendritic cells arise from a common bone marrow progenitor cell subset. Immunity 3:459–73 7. Kondo M, Weissman IL, Akashi K. 1997. Identiﬁcation of clonogenic common lymphoid progenitors in mouse bone marrow. Cell 91:661–72 8. Ryan DH, Nuccie BL, Ritterman I, Liesveld JL, Abboud CN, Insel RA. 1997. Expression of interleukin-7 receptor by lineage-negative human bone marrow progenitors with enhanced lymphoid proliferative potential and B-lineage differentiation capacity. Blood 89:929–40 9. Manz MG, Miyamoto T, Akashi K, Weissman IL. 2002. Prospective isolation of human clonogenic common myeloid progenitors. Proc. Natl. Acad. Sci. USA 99:11872–77 10. Haddad R, Guardiola P, Izac B, Thibault C, Radich J, et al. 2004. Molecular characterization of early human T/NK and B-lymphoid progenitor cells in umbilical cord blood. Blood 104:3918–26 11. Barcena A, Muench MO, Galy AH, Cupp J, Roncarolo MG, et al. 1993. Phenotypic and functional analysis of T-cell precursors in the human fetal liver and thymus: CD7 expression in the early stages of T- and myeloid-cell development. Blood 82:3401– 14 12. Miller JS, Alley KA, McGlave P. 1994. Differentiation of natural killer (NK) cells from human primitive marrow progenitors in a stroma-based long-term culture system: identiﬁcation of a CD34+ 7+ NK progenitor. Blood 83:2594–601 13. Rossi MI, Yokota T, Medina KL, Garrett KP, Comp PC, et al. 2003. B lymphopoiesis is active throughout human life, but there are developmental age-related changes. Blood 101:576–84 14. Ishii T, Nishihara M, Ma F, Ebihara Y, Tsuji K, et al. 1999. Expression of stromal cellderived factor-1/pre-B cell growth-stimulating factor receptor, CXC chemokine receptor 4, on CD34+ human bone marrow cells is a phenotypic alteration for committed lymphoid progenitors. J. Immunol. 163:3612–20 15. Hao Q-L, Zhu J, Price MA, Payne KJ, Barsky LW, Crooks GM. 2001. Identiﬁcation of a novel, human multilymphoid progenitor in cord blood. Blood 97:3683–90 16. Chicha L, Jarrossay D, Manz MG. 2004. Clonal type I interferon-producing and dendritic cell precursors are contained in both human lymphoid and myeloid progenitor populations. J. Exp. Med. 200:1519–24 17. Hao QL, Zhu J, Price MA, Payne KJ, Barsky LW, Crooks GM. 2001. Identiﬁcation of a novel, human multilymphoid progenitor in cord blood. Blood 97:3683–90 18. Res P, Martinez-Caceres E, Cristina Jaleco A, Staal F, Noteboom E, et al. 1996. CD34+ CD38dim cells in the human thymus can differentiate into T, natural killer, and dendritic cells but are distinct from pluripotent stem cells. Blood 87:5196–206

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

310

Blom

·

Spits

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

ARI

15 February 2006

21:28

19. Dik WA, Pike-Overzet K, Weerkamp F, de Ridder D, de Haas EFE, et al. 2005. New insights on human T cell development by quantitative T cell receptor gene rearrangement studies and gene expression proﬁling. J. Exp. Med. 201:1715–23 20. Bhandoola A, Sambandam A, Allman D, Meraz A, Schwarz B. 2003. Early T lineage progenitors: new insights, but old questions remain. J. Immunol. 171:5653–58 21. Porritt HE, Rumfelt LL, Tabrizifard S, Schmitt TM, Zuniga-Pﬂucker JC, Petrie HT. 2004. Heterogeneity among DN1 prothymocytes reveals multiple progenitors with different capacities to generate T cell and non-T cell lineages. Immunity 20:735–45 22. Petrie HT, Kincade PW. 2005. Many roads, one destination for T cell progenitors. J. Exp. Med. 202:11–13 23. Blom B, Res P, Noteboom E, Weijer K, Spits H. 1997. Prethymic CD34+ progenitors capable of developing into T cells are not committed to the T cell lineage. J. Immunol. 158:3571–77 24. Ktorza S, Blanc C, Laurent C, Sarun S, Verpilleux MP, et al. 1996. Complete TCR-δ rearrangements and partial (D-J) recombination of the TCR-β locus in CD34+ 7+ precursors from human cord blood. J. Immunol. 156:4120–27 25. Klein F, Feldhahn N, Lee S, Wang H, Ciufﬁ F, et al. 2003. T lymphoid differentiation in human bone marrow. Proc. Natl. Acad. Sci. USA 100:6747–52 26. Weerkamp F, de Haas EF, Naber BA, Comans-Bitter WM, Bogers AJ, et al. 2005. Agerelated changes in the cellular composition of the thymus in children. J. Allergy Clin. Immunol. 115:834–40 26a. De Smedt M, Hoebeke I, Reynvoet K, Leclercq G, Plum J. 2005. Different thresholds of Notch signaling bias human precursor cells towards B, NK, monocytic/dendritic cell or T cell lineage in thymus microenvironment. Blood. In press 27. Spits H, Blom B, Jaleco AC, Weijer K, Verschuren MC, et al. 1998. Early stages in the development of human T, natural killer and thymic dendritic cells. Immunol. Rev. 165:75–86 28. Spits H. 2002. Development of αβ T cells in the human thymus. Nat. Rev. Immunol. 2:760–72 29. Galy A, Verma S, Barcena A, Spits H. 1993. Precursors of CD3+ CD4+ CD8+ cells in the human thymus are deﬁned by expression of CD34. Delineation of early events in human thymic development. J. Exp. Med. 178:391–401 30. Dalloul AH, Patry C, Salamero J, Canque B, Grassi F, Schmitt C. 1999. Functional and phenotypic analysis of thymic CD34+ CD1a− progenitor-derived dendritic cells: predominance of CD1a+ differentiation pathway. J. Immunol. 162:5821–28 31. Res P, Spits H. 1999. Developmental stages in the human thymus. Semin. Immunol. 11:39–46 32. Hori T, Cupp J, Wrighton N, Lee F, Spits H. 1991. Identiﬁcation of a novel human thymocyte subset with a phenotype of CD3− CD4+ CD8α + β-1. Possible progeny of the CD3− CD4− CD8− subset. J. Immunol. 146:4078–84 33. Blom B, Verschuren MC, Heemskerk MH, Bakker AQ, van Gastel-Mol EJ, et al. 1999. TCR gene rearrangements and expression of the pre-T cell receptor complex during human T-cell differentiation. Blood 93:3033–43 34. Verschuren MC, Blom B, Bogers AJ, Spits H, van Dongen JJ. 1998. PJA-BP expression and TCR δ deletion during human T cell differentiation. Int. Immunol. 10:1873–80 35. Carrasco YR, Trigueros C, Ramiro AR, de Yebenes VG, Toribio ML. 1999. β-selection is associated with the onset of CD8β chain expression on CD4+ CD8αα + pre-T cells during human intrathymic development. Blood 94:3491–98 www.annualreviews.org • Development of Human Lymphoid Cells

311

ARI

15 February 2006

21:28

36. Trigueros C, Ramiro AR, Carrasco YR, de Yebenes VG, Albar JP, Toribio ML. 1998. Identiﬁcation of a late stage of small noncycling pTα − pre-T cells as immediate precursors of T cell receptor α/β+ thymocytes. J. Exp. Med. 188:1401–12 37. Noguchi M, Yi H, Rosenblatt HM, Filipovich AH, Adelstein S, et al. 1993. Interleukin-2 receptor gamma chain mutation results in X-linked severe combined immunodeﬁciency in humans. Cell 73:147–57 38. Russell SM, Keegan AD, Harada N, Nakamura Y, Noguchi M, et al. 1993. Interleukin-2 receptor γ chain: A functional component of the interleukin-4 receptor. Science 262:1880– 83 39. Puel A, Ziegler SF, Buckley RH, Leonard WJ. 1998. Defective IL7R expression in T− B+ NK+ severe combined immunodeﬁciency. Nat. Genet. 20:394–97 40. Giliani S, Mori L, de Saint Basile G, Le Deist F, Rodriguez-Perez C, et al. 2005. Interleukin-7 receptor α (IL-7Rα) deﬁciency: cellular and molecular bases. Analysis of clinical, immunological, and molecular features in 16 novel patients. Immunol. Rev. 203:110–26 41. Macchi P, Villa A, Gillani S, Sacco MG, Frattini A, et al. 1995. Mutations of Jak-3 gene in patients with autosomal severe combined immunodeﬁciency. Nature 377:65– 68 42. Russell SM, Tayebi N, Nakajima H, Riedy MC, Roberts JL, et al. 1995. Mutation of Jak3 in a patient with SCID: essential role of Jak3 in lymphoid development. Science 270:797–800 43. Fischer A, Le Deist F, Hacein-Bey-Abina S, Andre-Schmutz I, Basile GD, et al. 2005. Severe combined immunodeﬁciency. A model disease for molecular immunology and therapy. Immunol. Rev. 203:98–109 44. Plum J, De Smedt M, Leclercq G, Verhasselt B, Vandekerckhove B. 1996. Interleukin-7 is a critical growth factor in early human T-cell development. Blood 88:4239–45 45. Pallard C, Stegmann AP, van Kleffens T, Smart F, Venkitaraman A, Spits H. 1999. Distinct roles of the phosphatidylinositol 3-kinase and STAT5 pathways in IL-7-mediated development of human thymocyte precursors. Immunity 10:525–35 46. Peschon JJ, Morrissey PJ, Grabstein KH, Ramsdell FJ, Maraskovsky E, et al. 1994. Early lymphocyte expansion is severely impaired in interleukin 7 receptor-deﬁcient mice. J. Exp. Med. 180:1955–60 47. Sleasman JW, Harville TO, White GB, George JF, Barrett DJ, Goodenow MM. 1994. Arrested rearrangement of TCR V beta genes in thymocytes from children with X-linked severe combined immunodeﬁciency disease. J. Immunol. 153:442–48 48. Pesu M, Candotti F, Husa M, Hofmann SR, Notarangelo LD, O’Shea JJ. 2005. Jak3, severe combined immunodeﬁciency, and a new class of immunosuppressive drugs. Immunol. Rev. 203:127–42 49. Napolitano LA, Grant RM, Deeks SG, Schmidt D, De Rosa SC, et al. 2001. Increased production of IL-7 accompanies HIV-1-mediated T-cell depletion: implications for T-cell homeostasis. Nat. Med. 7:73–79 50. Artavanis-Tsakonas S, Rand MD, Lake RJ. 1999. Notch signaling: cell fate control and signal integration in development. Science 284:770–76 51. Radtke F, Wilson A, Stark G, Bauer M, van Meerwijk J, et al. 1999. Deﬁcient T cell fate speciﬁcation in mice with an induced inactivation of Notch1. Immunity 10:547–58 52. Jaleco AC, Neves H, Hooijberg E, Gameiro P, Clode N, et al. 2001. Differential effects of Notch ligands Delta-1 and Jagged-1 in human lymphoid differentiation. J. Exp. Med. 194:991–1002

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

312

Blom

·

Spits

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

ARI

15 February 2006

21:28

53. La Motte-Mohs RN, Herer E, Zuniga-Pﬂucker JC. 2005. Induction of T-cell development from human cord blood hematopoietic stem cells by Delta-like 1 in vitro. Blood 105:1431–39 54. De Smedt M, Hoebeke I, Plum J. 2004. Human bone marrow CD34+ progenitor cells mature to T cells on OP9-DL1 stromal cell line without thymus microenvironment. Blood Cells Mol. Dis. 33:227–32 55. Schmitt TM, Zuniga-Pﬂucker JC. 2002. Induction of T cell development from hematopoietic progenitor cells by Delta-like-1 in vitro. Immunity 17:749–56 55a. Dontje W, Schotte R, Cupedo T, Nagasawa M, Scheeren F, et al. 2006. DeltaLike1 induced Notch1 signalling regulates the human plasmacytoid dendritic cell versus T cell lineage decision through control of GATA-3 and Spi-B. Blood. In press 56. Wolfer A, Wilson A, Nemir M, MacDonald HR, Radtke F. 2002. Inactivation of Notch2 impairs VDJβ rearrangement and allows pre-TCR-independent survival of early αβ lineage thymocytes. Immunity 16:869–79 57. Schmitt TM, Ciofani M, Petrie HT, Zuniga-Pﬂucker JC. 2004. Maintenance of T cell speciﬁcation and differentiation requires recurrent Notch receptor-ligand interactions. J. Exp. Med. 200:469–79 58. Radtke F, Ferrero I, Wilson A, Lees R, Aguet M, MacDonald HR. 2000. Notch1 deﬁciency dissociates the intrathymic development of dendritic cells and T cells. J. Exp. Med. 191:1085–94 59. De Smedt M, Reynvoet K, Kerre T, Taghon T, Verhasselt B, et al. 2002. Active form of Notch imposes T cell fate in human progenitor cells. J. Immunol. 169:3021–29 60. Garcia-Peydro M, de Yebenes VG, Toribio ML. 2003. Sustained Notch1 signaling instructs the earliest human intrathymic precursors to adopt a γδ T-cell fate in fetal thymus organ culture. Blood 102:2444–51 61. Washburn T, Schweighoffer E, Gridley T, Chang D, Fowlkes BJ, et al. 1997. Notch activity inﬂuences the αβ versus γδ T cell lineage decision. Cell 88:833–43 62. Engel I, Murre C. 2001. The function of E- and Id proteins in lymphocyte development. Nat. Rev. Immunol. 1:193–99 63. Heemskerk MH, Blom B, Nolan G, Stegmann AP, Bakker AQ, et al. 1997. Inhibition of T cell and promotion of natural killer cell development by the dominant negative helix loop helix factor Id3. J. Exp. Med. 186:1597–602 64. Blom B, Heemskerk MH, Verschuren MC, van Dongen JJ, Stegmann AP, et al. 1999. Disruption of αβ but not of γδ T cell development by overexpression of the helix-loophelix protein Id3 in committed T cell progenitors. EMBO J. 18:2793–802 65. Yokota Y, Mansouri A, Mori S, Sugawara S, Adachi S, et al. 1999. Development of peripheral lymphoid organs and natural killer cells depends on the helix-loop-helix inhibitor Id2. Nature 397:702–6 66. Herblot S, Steff AM, Hugo P, Aplan PD, Hoang T. 2000. SCL and LMO1 alter thymocyte differentiation: inhibition of E2A-HEB function and pre-Tα chain expression. Nat. Immunol. 1:138–44 67. Reizis B, Leder P. 2001. The upstream enhancer is necessary and sufﬁcient for the expression of the pre-T cell receptor α gene in immature T lymphocytes. J. Exp. Med. 194:979–90 68. Aifantis I, Azogui O, Feinberg J, Saint Ruf C, Buer J, von Boehmer H. 1998. On the role of the pre-T cell receptor in αβ versus γδ T lineage commitment. Immunity 9:649–55 69. Pai SY, Truitt ML, Ting CN, Leiden JM, Glimcher LH, Ho IC. 2003. Critical roles for transcription factor GATA-3 in thymocyte development. Immunity 19:863–75 www.annualreviews.org • Development of Human Lymphoid Cells

313

ARI

15 February 2006

21:28

70. Taghon T, De Smedt M, Stolz F, Cnockaert M, Plum J, Leclercq G. 2001. Enforced expression of GATA-3 severely reduces human thymic cellularity. J. Immunol. 167:4468– 75 71. Lanier LL. 2005. NK cell recognition. Annu. Rev. Immunol. 23:225–74 72. Nagler A, Lanier LL, Cwirla S, Phillips JH. 1989. Comparative studies of human FcRIIIpositive and negative natural killer cells. J. Immunol. 143:3183–91 73. Cooper MA, Fehniger TA, Turner SC, Chen KS, Ghaheri BA, et al. 2001. Human natural killer cells: a unique innate immunoregulatory role for the CD56bright subset. Blood 97:3146–51 74. Jaleco AC, Blom B, Res P, Weijer K, Lanier LL, et al. 1997. Fetal liver contains committed NK progenitors, but is not a site for development of CD34+ cells into T cells. J. Immunol. 159:694–702 75. S´anchez MJ, Spits H, Lanier LL, Philips JH. 1993. Human natural killer cell committed thymocytes and their relationship to the T cell lineage. J. Exp. Med. 178:1857–66 76. Gunther U, Holloway JA, Gordon JG, Knight A, Chance V, et al. 2005. Phenotypic characterization of CD3− 7+ cells in developing human intestine and an analysis of their ability to differentiate into T cells. J. Immunol. 174:5414–22 77. S´anchez MJ, Muench MO, Roncarolo MG, Lanier LL, Phillips JH. 1994. Identiﬁcation of a common T/natural killer cell progenitor in human fetal thymus. J. Exp. Med. 180:569– 76 78. Sirianni MC, Businco L, Seminara R, Aiuti F. 1983. Severe combined immunodeﬁciencies, primary T-cell defects and DiGeorge syndrome in humans: characterization by monoclonal antibodies and natural killer cell activity. Clin. Immunol. Immunopathol. 28:361–70 79. Rosmaraki EE, Douagi I, Roth C, Colucci F, Cumano A, Di Santo JP. 2001. Identiﬁcation of committed NK cell progenitors in adult murine bone marrow. Eur. J. Immunol. 31:1900–9 80. Freud AG, Becknell B, Roychowdhury S, Mao HC, Ferketich AK, et al. 2005. A human CD34+ subset resides in lymph nodes and differentiates into CD56bright natural killer cells. Immunity 22:295–304 81. Fehniger TA, Cooper MA, Nuovo GJ, Cella M, Facchetti F, et al. 2003. CD56bright natural killer cells are present in human lymph nodes and are activated by T cell-derived IL-2: a potential new link between adaptive and innate immunity. Blood 101:3052–57 82. Ferlazzo G, Thomas D, Lin SL, Goodman K, Morandi B, et al. 2004. The abundant NK cells in human secondary lymphoid tissues require activation to express killer cell Ig-like receptors and become cytolytic. J. Immunol. 172:1455–62 83. Parrish-Novak J, Dillon SR, Nelson A, Hammond A, Sprecher C, et al. 2000. Interleukin 21 and its receptor are involved in NK cell expansion and regulation of lymphocyte function. Nature 408:57–63 84. Sivori S, Cantoni C, Parolini S, Marcenaro E, Conte R, et al. 2003. IL-21 induces both rapid maturation of human CD34+ cell precursors towards NK cells and acquisition of surface killer Ig-like receptors. Eur. J. Immunol. 33:3439–47 85. Rodewald HR, Moingeon P, Lucich JL, Dosiou C, Lopez P, Reinherz EL. 1992. A population of early fetal thymocytes expressing FcγRII/III contains precursors of T lymphocytes and natural killer cells. Cell 69:139–50 86. Phillips JH, Hori T, Nagler A, Bhat N, Spits H, Lanier LL. 1992. Ontogeny of human natural killer (NK) cells: Fetal NK cells mediate cytolytic function and express cytoplasmic CD3δ proteins. J. Exp. Med. 175:1055–66

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

314

Blom

·

Spits

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-10

ARI

15 February 2006

21:28

87. S´anchez MJ, Muench MO, Roncarolo MG, Lanier LL, Phillips JH. 1994. Identiﬁcation of a common T/NK cell progenitor in human fetal thymus. J. Exp. Med. 180:569– 76 88. Carlyle JR, Michie AM, Furlonger C, Nakano T, Lenardo MJ, et al. Identiﬁcation of a novel developmental stage marking lineage commitment of progenitor thymocytes. J. Exp. Med. 186:173–82 89. Ikawa T, Kawamoto H, Fujimoto S, Katsura Y. 1999. Commitment of common T/natural killer (NK) progenitors to unipotent T and NK progenitors in the murine fetal thymus revealed by a single progenitor assay. J. Exp. Med. 190:1617–26 90. Spits H, Lanier LL, Phillips JH. 1995. Development of human T and natural killer cells. Blood 85:2654–70 91. Marquez C, Trigueros C, Franco JM, Ramiro AR, Carrasco YR, et al. 1998. Identiﬁcation of a common developmental pathway for thymic natural killer cells and dendritic cells. Blood 91:2760–71 92. Perez SA, Sotiropoulou PA, Gkika DG, Mahaira LG, Niarchos DK, et al. 2003. A novel myeloid-like NK cell progenitor in human umbilical cord blood. Blood 101:3444– 50 93. Akashi K, Traver D, Miyamoto T, Weissman IL. 2000. A clonogenic common myeloid progenitor that gives rise to all myeloid lineages. Nature 404:193–97 94. Colucci F, Caligiuri MA, Di Santo JP. 2003. What does it take to make a natural killer? Nat. Rev. Immunol. 3:413–25 95. Vosshenrich CA, Samson-Villeger SI, Di Santo JP. 2005. Distinguishing features of developing natural killer cells. Curr. Opin. Immunol. 17:151–58 96. Bennett IM, Zatsepina O, Zamai L, Azzoni L, Mikheeva T, Perussia B. 1996. Deﬁnition of a natural killer NKR− P1A+ /CD56− /CD16− functionally immature human NK cell subset that differentiates in vitro in the presence of interleukin 12. J. Exp. Med. 184:1845– 56 97. Miller JS, McCullar V. 2001. Human natural killer cells with polyclonal lectin and immunoglobulinlike receptors develop from single hematopoietic stem cells with preferential expression of NKG2A and KIR2DL2/L3/S2. Blood 98:705–13 98. Sivori S, Falco M, Marcenaro E, Parolini S, Biassoni R, et al. 2002. Early expression of triggering receptors and regulatory role of 2B4 in human natural killer cell precursors undergoing in vitro differentiation. Proc. Natl. Acad. Sci. USA 99:4526–31 99. Zamai L, Ahmad M, Bennett IM, Azzoni L, Alnemri ES, Perussia B. 1998. Natural killer (NK) cell-mediated cytotoxicity: differential use of TRAIL and Fas ligand by immature and mature primary human NK cells. J. Exp. Med. 188:2375–80 100. Di Santo JP, Keever CA, Small T, Nichols GL, O’Reilly RJ, Flomenberg N. 1990. Absence of interleukin 2 production in a severe combined immunodeﬁciency disease syndrome with T cells. J. Exp. Med. 171:1697–704 101. Roifman CM. 2005. Studies of patients’ thymi aid in the discovery and characterization of immunodeﬁciency in humans. Immunol. Rev. 203:143–55 102. Gilmour KC, Fujii H, Cranston T, Davies EG, Kinnon C, Gaspar HB. 2001. Defective expression of the interleukin-2/interleukin-15 receptor β subunit leads to a natural killer cell-deﬁcient form of severe combined immunodeﬁciency. Blood 98:877–79 103. Vosshenrich CAJ, Ranson T, Samson SI, Corcuff E, Colucci F, et al. 2005. Roles for common cytokine receptor γ-chain-dependent cytokines in the generation, differentiation, and maturation of NK cell precursors and peripheral NK cells in vivo. J. Immunol. 174:1213–21 www.annualreviews.org • Development of Human Lymphoid Cells

315

ARI

15 February 2006

21:28

104. Kasaian MT, Whitters MJ, Carter LL, Lowe LD, Jussif JM, et al. 2002. IL-21 limits NK cell responses and promotes antigen-speciﬁc T cell activation: a mediator of the transition from innate to adaptive immunity. Immunity 16:559–69 105. Ikawa T, Fujimoto S, Kawamoto H, Katsura Y, Yokota Y. 2001. Commitment to natural killer cells requires the helix-loop-helix inhibitor Id2. Proc. Natl. Acad. Sci. USA 98:5164– 69 106. Schotte R, Rissoan MC, Bendriss-Vermare N, Bridon JM, Duhen T, et al. 2003. The transcription factor Spi-B is expressed in plasmacytoid DC precursors and inhibits T-, B-, and NK-cell development. Blood 101:1015–23 107. van den Brandt J, Voss K, Schott M, Hunig T, Wolfe MS, Reichardt HM. 2004. Inhibition of Notch signaling biases rat thymocyte development towards the NK cell lineage. Eur. J. Immunol. 34:1405–13 108. Bories JC, Willerford DM, Grevin D, Davidson L, Camus A, et al. 1995. Increased T-cell apoptosis and terminal B-cell differentiation induced by inactivation of the Ets-1 proto-oncogene. Nature 377:635–38 109. Barton K, Muthusamy N, Fischer C, Ting CN, Walunas TL, et al. 1998. The Ets1 transcription factor is required for the development of natural killer cells in mice. Immunity 9:555–63 110. Banchereau J, Briere F, Caux C, Davoust J, Lebecque S, et al. 2000. Immunobiology of dendritic cells. Annu. Rev. Immunol. 18:767–811 111. Liu Y-J. 2005. IPC: professional type 1 interferon-producing cells and plasmacytoid dendritic cell precursors. Annu. Rev. Immunol. 23:275–306 112. MacDonald KPA, Munster DJ, Clark GJ, Dzionek A, Schmitz J, Hart DNJ. 2002. Characterization of human blood dendritic cell subsets. Blood 100:4512–20 113. Galy A, Christopherson I, Ferlazzo G, Liu G, Spits H, Georgopoulos K. 2000. Distinct signals control the hematopoiesis of lymphoid-related dendritic cells. Blood 95:128–37 114. Canque B, Camus S, Dalloul A, Kahn E, Yagello M, et al. 2000. Characterization of dendritic cell differentiation pathways from cord blood CD34+ CD7+ CD45RA+ hematopoietic progenitor cells. Blood 96:3748–56 115. Blom B, Ho S, Antonenko S, Liu YJ. 2000. Generation of interferon α-producing predendritic cell (pre-DC)2 from human CD34+ hematopoietic stem cells. J. Exp. Med. 192:1785–96 116. Bendriss-Vermare N, Barthelemy C, Durand I, Bruand C, Dezutter-Dambuyant C, et al. 2001. Human thymus contains IFN-α-producing CD11c− , myeloid CD11c+ , and mature interdigitating dendritic cells. J. Clin. Invest. 107:835–44 117. Res PC, Couwenberg F, Vyth-Dreese FA, Spits H. 1999. Expression of pTα mRNA in a committed dendritic cell precursor in the human thymus. Blood 94:2647–57 118. Pelayo R, Hirose J, Huang J, Garrett KP, Delogu A, et al. 2005. Derivation of 2 categories of plasmacytoid dendritic cells in murine bone marrow. Blood 105:4407–15 119. Rissoan MC, Duhen T, Bridon JM, Bendriss-Vermare N, Peronne C, et al. 2002. Subtractive hybridization reveals the expression of immunoglobulin-like transcript 7, EphB1, granzyme B, and 3 novel transcripts in human plasmacytoid dendritic cells. Blood 100:3295–303 120. Corcoran L, Ferrero I, Vremec D, Lucas K, Waithman J, et al. 2003. The lymphoid past of mouse plasmacytoid cells and thymic dendritic cells. J. Immunol. 170:4926–32 121. Grouard G, Rissoan MC, Filgueira L, Durand I, Banchereau J, Liu YJ. 1997. The enigmatic plasmacytoid T cells develop into dendritic cells with interleukin (IL)-3 and CD40ligand. J. Exp. Med. 185:1101–11

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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ANRV270-IY24-10

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122. Shigematsu H, Reizis B, Iwasaki H, Mizuno S, Hu D, et al. 2004. Plasmacytoid dendritic cells activate lymphoid-speciﬁc genetic programs irrespective of their cellular origin. Immunity 21:43–53 123. Traver D, Akashi K, Manz M, Merad M, Miyamoto T, et al. 2000. Development of CD8α-positive dendritic cells from a common myeloid progenitor. Science 290:2152– 54 124. Martin CH, Aifantis I, Scimone ML, von Andrian UH, Reizis B, et al. 2003. Efﬁcient thymic immigration of B220+ lymphoid-restricted bone marrow cells with T precursor potential. Nat. Immunol. 4:866–73 125. Blom B, Ligthart SJ, Schotte R, Spits H. 2002. Developmental origin of pre-DC2. Hum. Immunol. 63:1072–80 126. Spits H, Couwenberg F, Bakker AQ, Weijer K, Uittenbogaart CH. 2000. Id2 and Id3 inhibit development of CD34+ stem cells into pre-dendritic cell (pre-DC)2 but not into pre-DC1. Evidence for a lymphoid origin of pre-DC2. J. Exp. Med. 192:1775–84 127. Comeau MR, de Vries ARV, Maliszewski CR, Galibert L. 2002. CD123bright plasmacytoid predendritic cells: progenitors undergoing cell fate conversion? J. Immunol. 169:75– 83 128. Gilliet M, Boonstra A, Paturel C, Antonenko S, Xu XL, et al. 2002. The development of murine plasmacytoid dendritic cell precursors is differentially regulated by FLT3-ligand and granulocyte/macrophage colony-stimulating factor. J. Exp. Med. 195:953–58 129. Naik SH, Proietto AI, Wilson NS, Dakic A, Schnorrer P, et al. 2005. Cutting edge: generation of splenic CD8+ and CD8− dendritic cell equivalents in Fms-like tyrosine kinase 3 ligand bone marrow cultures. J. Immunol. 174:6592–97 130. Bjorck P. 2001. Isolation and characterization of plasmacytoid dendritic cells from Flt3 ligand and granulocyte-macrophage colony-stimulating factor-treated mice. Blood 98:3520–26 131. Coates PT, Barratt-Boyes SM, Zhang L, Donnenberg VS, O’Connell PJ, et al. 2003. Dendritic cell subsets in blood and lymphoid tissue of rhesus monkeys and their mobilization with Flt3 ligand. Blood 102:2513–21 132. Manfra DJ, Chen SC, Jensen KK, Fine JS, Wiekowski MT, Lira SA. 2003. Conditional expression of murine Flt3 ligand leads to expansion of multiple dendritic cell subsets in peripheral blood and tissues of transgenic mice. J. Immunol. 170:2843–52 133. Maraskovsky E, Daro E, Roux E, Teepe M, Maliszewski CR, et al. 2000. In vivo generation of human dendritic cell subsets by Flt3 ligand. Blood 96:878–84 134. Pulendran B, Banchereau J, Burkeholder S, Kraus E, Guinet E, et al. 2000. Flt3-ligand and granulocyte colony-stimulating factor mobilize distinct human dendritic cell subsets in vivo. J. Immunol. 165:566–72 135. D’Amico A, Wu L. 2003. The early progenitors of mouse dendritic cells and plasmacytoid predendritic cells are within the bone marrow hemopoietic precursors expressing Flt3. J. Exp. Med. 198:293–303 136. Karsunky H, Merad M, Cozzio A, Weissman IL, Manz MG. 2003. Flt3 ligand regulates dendritic cell development from Flt3+ lymphoid and myeloid-committed progenitors to Flt3+ dendritic cells in vivo. J. Exp. Med. 198:305–13 137. McKenna HJ, Stocking KL, Miller RE, Brasel K, De Smedt T, et al. 2000. Mice lacking ﬂt3 ligand have deﬁcient hematopoiesis affecting hematopoietic progenitor cells, dendritic cells, and natural killer cells. Blood 95:3489–97 138. Laouar Y, Welte T, Fu XY, Flavell RA. 2003. STAT3 is required for Flt3L-dependent dendritic cell differentiation. Immunity 19:903–12 www.annualreviews.org • Development of Human Lymphoid Cells

317

ARI

15 February 2006

21:28

139. Rathinam C, Geffers R, Yucel R, Buer J, Welte K, et al. 2005. The transcriptional repressor Gﬁ1 controls STAT3-dependent dendritic cell development and function. Immunity 22:717–28 140. Chen W, Antonenko S, Sederstrom JM, Liang X, Chan AS, et al. 2004. Thrombopoietin cooperates with FLT3-ligand in the generation of plasmacytoid dendritic cell precursors from human hematopoietic progenitors. Blood 103:2547–53 141. Wu L, Nichogiannopoulou A, Shortman K, Georgopoulos K. 1997. Cell-autonomous defects in dendritic cell populations of Ikaros mutant mice point to a developmental relationship with the lymphoid lineage. Immunity 7:483–92 142. Wu L, D’Amico A, Winkel KD, Suter M, Lo D, Shortman K. 1998. RelB is essential for the development of myeloid-related CD8α − dendritic cells but not of lymphoid-related CD8α + dendritic cells. Immunity 9:839–47 143. Hacker C, Kirsch RD, Ju XS, Hieronymus T, Gust TC, et al. 2003. Transcriptional proﬁling identiﬁes Id2 function in dendritic cell development. Nat. Immunol. 4:380–86 144. Schotte R, Rissoan MC, Bendriss-Vermare N, Bridon JM, Duhen T, et al. 2002. The transcription factor Spi-B is expressed in plasmacytoid DC precursors and inhibits T, B, and NK cell development. Blood 101:1015–23 145. Schotte R, Nagasawa M, Weijer K, Spits H, Blom B. 2004. The ETS transcription factor Spi-B is required for human plasmacytoid dendritic cell development. J. Exp. Med. 200:1503–9 146. Iwasaki H, Somoza C, Shigematsu H, Duprez EA, Iwasaki-Arai J, et al. 2005. Distinctive and indispensable roles of PU.1 in maintenance of hematopoietic stem cells and their differentiation. Blood 106:1590–600 147. Tamura T, Tailor P, Yamaoka K, Kong HJ, Tsujimura H, et al. 2005. IFN regulatory factor-4 and -8 govern dendritic cell subset development and their functional diversity. J. Immunol. 174:2573–81 148. Tsujimura H, Tamura T, Ozato K. 2003. Cutting edge: IFN consensus sequence binding protein/IFN regulatory factor 8 drives the development of type I IFN-producing plasmacytoid dendritic cells. J. Immunol. 170:1131–35 149. Brass AL, Kehrli E, Eisenbeis CF, Storb U, Singh H. 1996. Pip, a lymphoid-restricted IRF, contains a regulatory domain that is important for autoinhibition and ternary complex formation with the Ets factor PU.1. Genes Dev. 10:2335–47 150. Nakano N, Nishiyama C, Masuoka N, Nishiyama M, Yamane H, et al. 2005. Analysis of PU.1/ICSBP (IRF-8) complex formation with various PU.1 mutants: molecular cloning of rat Icsbp (Irf-8) cDNA. Immunogenetics 56:871–77 151. van der Stoep N, Quinten E, Rezende MM, van den Elsen PJ. 2004. E47, IRF-4, and PU.1 synergize to induce B-cell-speciﬁc activation of the class II transactivator promoter III (CIITA-PIII). Blood 104:2849–57 152. Nunez C, Nishimoto N, Gartland GL, Billips LG, Burrows PD, et al. 1996. B cells are generated throughout life in humans. J. Immunol. 156:866–72 153. Solvason N, Kearney JF. 1992. The human fetal omentum: a site of B cell generation. J. Exp. Med. 175:397–404 154. Reynaud D, Lefort N, Manie E, Coulombel L, Levy Y. 2003. In vitro identiﬁcation of human pro-B cells that give rise to macrophages, natural killer cells, and T cells. Blood 101:4313–21 155. Sanz E, Alvarez-Mon M, Martinez AC, de la Hera A. 2003. Human cord blood CD34+ Pax-5+ B-cell progenitors: single-cell analyses of their gene expression proﬁles. Blood 101:3424–30

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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Blom

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Spits

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ANRV270-IY24-10

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15 February 2006

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156. Hardy RR, Carmack CE, Shinton SA, Kemp JD, Hayakawa K. 1991. Resolution and characterization of pro-B and pre-pro-B cell stages in normal mouse bone marrow. J. Exp. Med. 173:1213–25 157. Bertrand FE 3rd, Billips LG, Burrows PD, Gartland GL, Kubagawa H, et al. 1997. Ig DH gene segment transcription and rearrangement before surface expression of the pan-B-cell marker CD19 in normal human bone marrow. Blood 90:736–44 158. Dworzak MN, Fritsch G, Froschl G, Printz D, Gadner H. 1998. Four-color ﬂow cytometric investigation of terminal deoxynucleotidyl transferase-positive lymphoid precursors in pediatric bone marrow: CD79a expression precedes CD19 in early B-cell ontogeny. Blood 92:3203–9 159. Wang YH, Nomura J, Faye-Petersen OM, Cooper MD. 1998. Surrogate light chain production during B cell differentiation: differential intracellular versus cell surface expression. J. Immunol. 161:1132–39 160. Busslinger M. 2004. Transcriptional control of early B cell development. Annu. Rev. Immunol. 22:55–79 161. Ghia P, ten Boekel E, Sanz E, de la Hera A, Rolink A, Melchers F. 1996. Ordering of human bone marrow B lymphocyte precursors by single-cell polymerase chain reaction analyses of the rearrangement status of the immunoglobulin H and L chain gene loci. J. Exp. Med. 184:2217–29 162. LeBien TW. 2000. Fates of human B-cell precursors. Blood 96:9–23 163. Loken MR, Shah VO, Dattilio KL, Civin CI. 1987. Flow cytometric analysis of human bone marrow. II. Normal B lymphocyte development. Blood 70:1316–24 164. LeBien TW, Wormann B, Villablanca JG, Law CL, Steinberg LM, et al. 1990. Multiparameter ﬂow cytometric analysis of human fetal bone marrow B cells. Leukemia 4:354–58 165. Davi F, Faili A, Gritti C, Blanc C, Laurent C, Sutton L, et al. 1997. Early onset of immunoglobulin heavy chain gene rearrangements in normal human bone marrow CD34+ cells. Blood 90:4014–21 166. Lemmers B, Gauthier L, Guelpa-Fonlupt V, Fougereau M, Schiff C. 1999. The human (L+ μ− ) proB complex: cell surface expression and biochemical structure of a putative transducing receptor. Blood 93:4336–46 167. Dittel BN, LeBien TW. 1995. The growth response to IL-7 during normal human B cell ontogeny is restricted to B-lineage cells expressing CD34. J. Immunol. 154:58–67 168. Hou YH, Srour EF, Ramsey H, Dahl R, Broxmeyer HE, Hromas R. 2005. Identiﬁcation of a human B-cell/myeloid common progenitor by the absence of CXCR4. Blood 105:3488–92 169. Nutt SL, Morrison AM, Dorﬂer P, Rolink A, Busslinger M. 1998. Identiﬁcation of BSAP (Pax-5) target genes in early B-cell development by loss- and gain-of-function experiments. EMBO J. 17:2319–33 170. Nutt SL, Urbanek P, Rolink A, Busslinger M. 1997. Essential functions of Pax5 (BSAP) in pro-B cell development: difference between fetal and adult B lymphopoiesis and reduced V-to-DJ recombination at the IgH locus. Genes Dev. 11:476–91 171. Milne CD, Fleming HE, Zhang Y, Paige CJ. 2004. Mechanisms of selection mediated by interleukin-7, the preBCR, and hemokinin-1 during B-cell development. Immunol. Rev. 197:75–88 172. Namikawa R, Muench MO, de Vries JE, Roncarolo MG. 1996. The FLK2/FLT3 ligand synergizes with interleukin-7 in promoting stromal-cell-independent expansion and differentiation of human fetal pro-B cells in vitro. Blood 87:1881–90

www.annualreviews.org • Development of Human Lymphoid Cells

319

ARI

15 February 2006

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173. Prieyl JA, LeBien TW. 1996. Interleukin 7 independent development of human B cells. Proc. Natl. Acad. Sci. USA 93:10348–53 174. Macchi P, Villa A, Giliani S, Sacco MG, Frattini A, et al. 1995. Mutations of Jak-3 gene in patients with autosomal severe combined immune deﬁciency (SCID). Nature 377:65–68 175. Kovanen PE, Leonard WJ. 2004. Cytokines and immunodeﬁciency diseases: critical roles of the γc -dependent cytokines interleukins 2, 4, 7, 9, 15, and 21, and their signaling pathways. Immunol. Rev. 202:67–83 176. Nutt SL, Metcalf D, D’Amico A, Polli M, Wu L. 2005. Dynamic regulation of PU.1 expression in multipotent hematopoietic progenitors. J. Exp. Med. 201:221–31 177. Berardi AC, Meffre E, Pﬂumio F, Katz A, Vainchenker W, et al. 1997. Individual CD34+ CD38low CD19− CD10− progenitor cells from human cord blood generate B lymphocytes and granulocytes. Blood 89:3554–64 178. Bond HM, Mesuraca M, Carbone E, Bonelli P, Agosti V, et al. 2004. Early hematopoietic zinc ﬁnger protein (EHZF), the human homolog to mouse Evi3, is highly expressed in primitive human hematopoietic cells. Blood 103:2062–70 179. Jaleco AC, Stegmann AP, Heemskerk MH, Couwenberg F, Bakker AQ, et al. 1999. Genetic modiﬁcation of human B-cell development: B-cell development is inhibited by the dominant negative helix loop helix factor Id3. Blood 94:2637–46 180. Traggiai E, Chicha L, Mazzucchelli L, Bronz L, Piffaretti JC, et al. 2004. Development of a human adaptive immune system in cord blood cell-transplanted mice. Science 304:104–7 181. Gimeno R, Weijer K, Voordouw A, Uittenbogaart CH, Legrand N, et al. 2004. Monitoring the effect of gene silencing by RNA interference in human CD34+ cells injected into newborn RAG2−/− γc−/− mice: functional inactivation of p53 in developing T cells. Blood 104:3886–93 182. Ishikawa F, Yasukawa M, Lyons B, Yoshida S, Miyamoto T, et al. 2005. Development of functional human blood and immune systems in NOD/SCID/IL2 receptor γ chainnull mice. Blood 106:1565–73

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Contents

Annual Review of Immunology Volume 24, 2006

Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:287-320. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Genetic Disorders of Programmed Cell Death in the Immune System∗ Nicolas Bid`ere,† Helen C. Su,† and Michael J. Lenardo Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892; email: [email protected]

Annu. Rev. Immunol. 2006. 24:321–52

Key Words

First published online as a Review in Advance on December 12, 2005

cytokine, Fas/APO-1/CD95, caspase, tumor necrosis factor receptor, autoimmune lymphoproliferative syndrome (ALPS), caspase-eight deﬁciency state (CEDS)

The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090513 c 2006 by Copyright Annual Reviews. All rights reserved ∗ The U.S. Government has the right to retain a nonexclusive, royalty-free license in and to any copyright covering this paper. †

These authors contributed equally to this work.

Abstract Human genetics offers new possibilities for understanding physiological regulatory mechanisms and disorders of the immune system. Genetic abnormalities of lymphocyte cell death programs have provided insights into mechanisms of receptor biology and principles of immune homeostasis and tolerance. Thus far, there are two major diseases of programmed cell death associated with inherited human mutations: the autoimmune lymphoproliferative syndrome and the caspase-eight deﬁciency state. We describe the details of their molecular pathogenesis and discuss how these diseases illustrate important concepts in immune regulation and genetics.

0732-0582/06/0423-0321$20.00

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INTRODUCTION

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ALPS: autoimmune lymphoproliferative syndrome Apoptosis: programmed cell death characterized by cell shrinkage, nuclear condensation with DNA fragmentation, membrane blebbing, and breakdown into rapidly phagocytosed apoptotic bodies Caspases: a family of highly conserved intracellular cysteine proteases involved in apoptosis, development, cytokine maturation, and normal cell function

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Human genetics has entered a new era. Technological advances allow investigators to probe inherited diseases with unprecedented molecular precision and thus shed light on normal physiological pathways. The complete DNA sequence of the human genome, new techniques to introduce DNA into primary and transformed cells, and RNA interference for selectively inhibiting gene expression now allow experimentalists to examine human cells as powerfully as cells from model organisms. The major advantage of human research has always been the widespread detection and documentation of unusual genetic conditions. The marriage of new technology and disease investigation brings humans to the proscenium and may make experimental animal systems supporting actors in future genetic research. How the immune system maintains homeostasis and tolerance remains a question of fundamental importance. On the low side, lymphocyte depletion and immunodeﬁciency can result. On the high side, a failure of regulatory mechanisms can cause an overabundance of lymphocytes and possibly cancer, as well as autoimmune and allergic reactions. The autoimmune lymphoproliferative syndrome (ALPS) is one of the ﬁrst human inherited disorders of apoptosis to be described (1). Investigations of this disease led to the ﬁrst descriptions of inherited mutations in genes encoding caspase proteins in humans (2, 3). It is clear that deﬁciencies in the death receptor Fas/APO-1/CD95, Fas ligand (FasL or CD95L), caspase-8, and caspase-10 primarily affect the immune system. In fact, it has been difﬁcult to ﬁnd abnormalities in any other organ system associated with mutations in these genes in humans. In this context, these mutations reveal the central role of apoptosis in immune homeostasis. Recent work also may point to a key role for these molecules and an associated form of nonapoptotic death in immune regulation (N. Bid`ere & M. Lenardo, unpublished observations). One key obser-

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vation is that dominant-negative mutations in genes that encode death factors predominate in ALPS. This type of mutation predominates because the transduction of a death signal involves speciﬁc stoichiometric protein complexes that are susceptible to interference by the inclusion of mutant signaling components. Given the widespread involvement of noncovalent protein complexes in cellular processes, dominant interference is likely to be a common mode for genetic abnormalities in humans, who are typically not consanguineous. Thus, disease mechanisms due to heterozygous mutations may be more prominent than those attributable to homozygous mutations. Finally, another conceptual advance stemming from these diseases is that an overabundance of lymphocytes does not necessarily equate with increased function. Caspase-8 deﬁciency causes, for example, lymphocytosis due to impaired apoptosis together with immunodeﬁciency due to defective antigen receptor signaling (2, 4).

IMMUNE REGULATION BY PROGRAMMED CELL DEATH A simpliﬁed view of lymphocyte regulation by apoptosis is useful for thinking about the molecular mechanisms involved (Figure 1). The basic principles of intrinsic (lymphokine withdrawal) and extrinsic (death receptor) death pathways have been reviewed previously (5, 6). In essence, high antigen load stimulates the extrinsic death pathway as a means to restrain the immune response and prevent immunotoxicity. On the other hand, the clearance of antigen leads to decreased trophic cytokine production and cell death by the intrinsic pathway. This event rids the body of excess, unnecessary lymphocytes and restores homeostasis. Genetic analysis in mice shows that an imbalance of either pathway can lead to a loss of lymphocyte control and tolerance (7). Antigen-speciﬁc cells that escape both pathways and reenter a resting state contribute to the pool of memory cells. The memory pool can be later called upon for anamnestic

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Figure 1 The propriocidal death pathway in lymphocytes. Antigen stimulation of resting T lymphocytes leads to their activation and proliferation. Activated T cells express Fas and its cognate ligand, FasL, which regulates Fas-expressing B cells and APC. Similar effects might be mediated by other death receptors. When T cells reencounter high antigen levels, the extrinsic pathway of cell death (active apoptosis) is triggered. After cessation of antigen and cytokine production, passive apoptosis ensues through the intrinsic pathway. The remaining T cells constitute the memory population.

responses. The key concept is that mature T and B lymphocytes can be programmed to die in an antigen-speciﬁc fashion. This antigencontrolled cell death affords dynamic control of both lymphocyte cell number and immunological tolerance in the peripheral immune system. Because variation in antigen levels stimulates T cells to kill either themselves or B cells, this constitutes a self-regulatory or propriocidal pathway of death. Recently, it has been proposed that an individual lineage of regulatory T cells can carry out suppressive functions in immune responses (8). We and others have observed that the suppressive effects of such cells is due to the induction of target cell death (9; P. Pandiyan & M. Lenardo, unpublished results). However, little has been done to examine possible immune protective functions of death induced by this variety of T cells. Therefore, it is difﬁcult to know whether the suppressive effects contribute to self-tolerance by deletion or through bystander effects of an immunological response involving cell killing. Although it was initially believed that the extrinsic pathway caused apoptosis mediated by death receptors in the tumor necrosis factor

(TNF) receptor superfamily, subsequent data has revealed a more complex situation. Antigen stimulation of T cells causes the induction of FasL and TNF, which can trigger the corresponding death domain (DD)-containing receptors (10–13). Moreover, the triggering of FasL on T cells can induce the death of B cells that have been activated and express Fas (Figure 1). The requirement for antigen stimulation to provoke the pathway—though often confused with a means to remove lymphocytes at the end of an immune response— indicates its importance for negative feedback regulation during a proliferative lymphocyte response to antigen. One complexity is that the blockade of death receptors does not completely eliminate the ability of antigen receptor stimulation to kill T cells (14). This suggests that alternative death mechanisms exist that have not yet been clearly identiﬁed. A second complexity is that the blockade of Fas-induced apoptosis does not prevent death. In fact, the addition of the pan-caspase inhibitor zVAD to Fas stimulation leads to increased alternative cell death in activated human peripheral blood T cells (N. Bid`ere & M. Lenardo, unpublished observations). There www.annualreviews.org • Genetic Disorders and Cell Death

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TNFR: tumor necrosis factor receptor Necrosis: nonapoptotic cell death characterized by cell swelling, early membrane damage, nuclear lysis without condensation, and a strong inﬂammatory response DD: death domain FADD: Fas-associated death domain DISC: death-inducing signaling complex DED: death effector domain

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have been several observations that triggering either Fas or TNF receptor 1 (TNFR1) can lead to programs of necrosis (15, 16). Presumably, this provides an important alternative in cases in which viruses or other infectious agents can impede apoptosis. However, a more thorough understanding of these complexities will be an important focus of further work in this area.

GENETIC INSIGHTS INTO PROGRAMMED DEATH BY FAS Physiological elimination of activated T cells during an immune response can occur via an active extrinsic pathway triggered by the engagement of specialized transmembrane receptors called death receptors. These death receptors belong to the TNFR superfamily and include Fas (also called CD95, TNFRSF6, or APO-1), TNFR1 (also known as p60 or TNFRSF1A), death receptor 3 (also known as DR3, or TNFRSF25), death receptor 4 [also known as DR4, TNFrelated apoptosis-inducing ligand receptor 1 (TRAIL-R1), or TNFRSF10A], death receptor 5 (DR5, or TRAIL-R2 or TNFRSF10B), and death receptor 6 (DR6 or TNFRSF21). The TNFR superfamily is characterized by arrays of two to ﬁve extracellular cysteine-rich domains (CRDs) (17). Members of the death receptor subgroup of TNFRs share a moderately conserved region of 80 amino acids in the cytoplasmic portion termed the death domain (DD), as it is required for death signaling. Fas, the prototypical death receptor, plays a critical role in lymphocyte regulation by apoptosis and, potentially, nonapoptotic death. Fas is activated by its cognate ligand, FasL, a type 2 transmembrane protein that can be released in soluble form by metalloproteinases (18). Both the ligand and the receptor can be potently upregulated by antigenic stimulation of T cells, especially if the cells are activated and cultured in interleukin (IL)-2 (19). Stimulation of B cells can induce death receptors and TNFα, but not expression of FasL. Rather, B cells appear to be killed Bid`ere

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by conjugation to T cells that express FasL (20) (Figure 1). The fundamental mode of signaling by death receptors is protein complex formation at two levels. First, the receptor assembly is triggered by ligand to form stoichiometric complexes with cytoplasmic signaling proteins. Second, higher order aggregates of receptors, ligands, and signaling proteins immediately follow and are crucial for signal transmission. As an example, we review Fas signaling. Within seconds after FasL stimulation, Fas recruits the adapter molecule FADD and the initiators caspases-8 and -10 to form a large complex termed death-inducing signaling complex (DISC) (21). DISC formation is crucial for generating caspase activity to initiate the apoptotic process. Both Fas and FasL operate as homotrimers. Their interaction triggers changes in conformation and/or orientation of the receptor trimer that allow homotypic interactions between the DDs in Fas and FADD. In addition to its COOHterminal DD, FADD also possesses a NH2 terminal protein-protein-interaction domain of approximately 80 residues called the death effector domain (DED), which is also found in the initiator (or apical) caspases-8 and -10. Remarkably, the DD and DED are both hexahelical bundles with a similar overall topology that is also shared by the caspase-recruitment domain (CARD) (22). Hence, these domains may be specializations of a common ancestor. It is generally the case, however, that each domain interacts in a homotypic manner, i.e., DD to DD, but do not cross bind one another (DD to DED, etc.). Therefore, as illustrated in Figure 2, FADD is brought to the DISC by its DD association with Fas and then recruits caspases-8 and -10 to the complex through DED interactions (23, 24). Within this multiprotein DISC aggregate, the increased local concentration of caspase8/10 zymogens presumably promotes an altered conformation that leads to their autoprocessing and activation. Caspases-8 and -10 each contain three parts: a double-DEDcontaining prodomain, a large enzymatic

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Figure 2 Caspase-8 is recruited to the plasma membrane and integrates two different signaling complexes, with two different outcomes. As described in the text, Fas triggers DISC formation with the release of highly active, processed caspase-8 into the cytosol that leads to the demise of the cells. By contrast, antigen receptor stimulation triggers activation-receptor induced signalosome (ARIS) formation. In that case, active caspase-8 remains unprocessed and allows the recruitment of IκB kinase (IKK) complex to promote NF-κB activation.

subunit, and a small enzymatic subunit. Each part is ﬂanked by aspartate residues that form caspase cleavage sites especially sensitive to its own activity. Initially, it was believed that autoprocessing and release of the two enzymatic subunits were essential for death signaling (25). However, recent data has suggested that apical caspases can, under certain conditions, transmit signals without cleavage by relying only on dimerization or other rearrangements of inactive unprocessed monomers (26). Nevertheless, processing of apical caspases stabilizes the catalytic activity as heterotetramers (two each of the large and small subunits) and allows its release into the cytosol (26). The released, highly active caspases cleave multiple

downstream targets, including effector caspases that initiate a caspase ampliﬁcation loop leading to the cell’s demise. In the so-called type I cells, including thymocytes and peripheral T cells, a high quantity of active caspase8/10 is generated at the DISC that directly activates effector caspases, i.e., caspases-3, -6, and -7. By contrast, in type II cells, including tumor cell lines such as Jurkat and CEM, the DISC is weaker, and caspase activation occurs more slowly and to a lesser extent than in type I cells. In type II cells, robust apoptosis is achieved by caspase-8 processing of Bid, a proapoptotic BH3-only Bcl-2 family member. The truncated active form of Bid causes cytochrome c release from the mitochondria www.annualreviews.org • Genetic Disorders and Cell Death

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SPOTS: signaling protein oligomerization transduction structures

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c-FLIP: cellular FLICE inhibitory protein DNT cells: double-negative T cells

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and activation of the intrinsic apoptosis pathway (27). Because of mitochondrial involvement, death receptor–mediated death in type II cells, but not type I cells, can be inhibited by anti-apoptotic Bcl-2 family members (28). DISC formation is followed by the formation of Fas clusters recently designated as signaling protein oligomerization transduction structures (SPOTS) (29, 30). SPOTS are composed of perhaps thousands of individual receptor complexes (making them visible microscopically) that further increase the local concentration of caspase-8 and promote its autoprocessing and activation (30). This receptor oligomerization depends on the Fas DD and FADD but does not require caspase activity. Once SPOTS are formed, a further caspase-dependent clustering and capping of Fas complexes is followed by receptor internalization in vesicles bearing endosomal markers. It is unclear whether internalization downregulates Fas-induced death or is a critical step for death signaling, as recently shown for the TNF receptosomes (31). Alternatively, since Fas internalization depends on caspase activation, it may be a post-lethal event that has no regulatory role at all. One potentially important regulator of Fas at the DISC level is the cellular FLICE inhibitory protein (c-FLIP) that is present in two isoforms: c-FLIPS and c-FLIPL (32). The c-FLIPL protein is very similar to caspases-8 and -10, and the genes for all three are found in a gene cluster on chromosome 2q33-34; this observation suggests that the genes may have arisen by tandem duplication of one ancestral gene (33–35). Although a double DED is intact in c-FLIPL , the part of the gene that encodes the enzymatic subunits contains numerous mutations that prevent caspase activity (34). c-FLIPS is closer in structure to a viral counterpart, v-FLIP, which consists only of the double DED and a short COOH-terminal addition (34). Both c-FLIPL and c-FLIPS can be recruited to the DISC, where they typically exert an inhibitory effect on caspase-8 activation and cell death. c-FLIPL has been observed to have different, sometimes conBid`ere

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tradictory functions. c-FLIPL in the DISC is processed by caspase-8 but cannot process caspase-8 in return, so that the latter is kept in an uncleaved form at the receptor level (36). B cell receptor stimulation can induce c-FLIPL expression and block Fas- and TRAIL-mediated killing; this observation indicates a potential role in B cell selection and tolerance (37). On the other hand, small amounts of c-FLIPL can cause caspase-8 activation and cell death and have also been proposed to promote Fas-induced lymphocyte proliferation (32).

THE AUTOIMMUNE LYMPHOPROLIFERATIVE SYNDROME (ALPS) Clinical Presentation ALPS is a rare human disorder deﬁned by lymphoproliferation, peripheral expansion of double-negative αβ T (DNT) cells, and impaired lymphocyte apoptosis. Autoimmune disease and an increased risk of lymphoma are also observed. The clinical presentation of ALPS in humans, which has been summarized in previous excellent reviews, reﬂects impaired lymphocyte homeostasis, particularly of the DNT cells (38, 39). The diagnostic criteria are listed in Table 1. Lymphoproliferation apparently results from the gradual accumulation of lymphocytes that have not undergone normal programmed cell death. This proliferation leads to chronic enlargement of the lymph nodes, thymus, liver, and/or spleen, beginning in early childhood. Both B cells, including CD5+ B cells, and T cells are elevated. Not all lymphocyte subsets are affected, as CD4+ CD25+ absolute numbers are reduced and CD4+ CD25− absolute numbers are not increased. By contrast, expansion of an unusual population of peripheral CD4− CD8− T cells is striking. These peripheral DNT cells express αβ T cell receptor (TCR) chains as well as the CD45R isoform B220 (typically expressed on B cells); this expression distinguishes them from

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Table 1 Diagnostic criteria for autoimmune lymphoproliferative syndrome (adapted from Reference 39)

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Criteria

Description

Required features

Chronic nonmalignant lymphadenopathy and/or splenomegaly Peripheral expansion of CD4− CD8− TCRα/β T lymphocytes (DNT cells) Impaired lymphocyte apoptosis

Supporting features

Family history of ALPS Characteristic histopathology Autoimmune disease Germline mutations in Fas, Fas ligand, or caspase-10 genes Somatic mutation in Fas gene in DNT cells

peripheral γδ T cells, which naturally lack CD4 and CD8 coreceptors (40). DNTs also differ from double-negative thymocytes, an immature stage of T cells in which TCR genes have not yet completely rearranged (41). The DNT cells could be either previously activated, mature T cells that have lost CD8 or CD4 coreceptor expression, or a special minor cell lineage (42). In the blood, DNT cells constitute less than 1% in normal individuals, but can reach up to 40% in patients with ALPS (40). The DNT expansion is also evident histologically by nearly pathognomic abnormalities of the architecture of secondary lymphoid tissues, with paracortical and follicular hyperplasia (39). DNT cells appear to play a crucial role in disease development. In certain ALPS patients lacking germline mutations in the death receptor Fas, Holzelova et al. (43) found a population of DNT cells harboring somatic dominantly interfering Fas mutations. Thus, unregulated and excessive DNT cells are seemingly sufﬁcient to cause disease. Two previously described characteristics of DNT cells may explain how. First, the DNT cells are primary producers of strikingly elevated IL10 observed in ALPS patients. By contrast, DNT cells from normal individuals do not produce IL-10. Moreover, healthy relatives of ALPS patients bearing both Fas mutations and in vitro apoptotic defects have only modestly elevated DNTs and IL-10 (44). Besides elevated IL-10, ALPS patients also exhibit in-

creased IL-4 and IL-5, but decreased IL-2 and IFN-γ (44, 45). This cytokine proﬁle is characteristic of T helper type 2 (Th2) cells, which inhibit cell-mediated immunity and promote humoral immune responses. Thus, the overall cytokine environment may favor autoantibody production in ALPS. Second, DNT cells exhibit an unusual phenotype including B220 expression and altered cell surface O-glycans (40). This phenotype could change the trafﬁcking pattern and/or potential interactions of DNT cells with other cell types. There are some important and unresolved paradoxes regarding DNTs. First, they are very difﬁcult to culture in vitro and almost immediately die despite an ostensible resistance to apoptosis in vivo. Their lifespan is not extended by IL10. Second, although DNTs are believed to play a role in hyperactive immune responses, they are generally unresponsive to proliferative and activating stimuli (46). Finally, the antigen speciﬁcities recognized by the TCRs on DNTs have not been deﬁned, but the general assumption is that they will include selfantigens. Autoimmunity accompanies lymphoproliferation in ALPS. Most patients have elevated serum immunoglobulin levels and autoantibodies. Anti-cardiolipin antibodies are frequent but not usually associated with thromboembolic disease (39). Many patients exhibit Coombs’ positive hemolytic anemia and/or immune thrombocytopenia (39). These two features are also observed in

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Evans syndrome, a diagnosis that can be confused with ALPS (47). Less common autoimmune manifestations in ALPS include anti-nuclear antibodies, rheumatoid factor, autoimmune neutropenia, glomerulonephritis, uveitis, autoimmune hepatitis, primary biliary cirrhosis, Guillain-Barr´e, vasculitis, linear IgA dermopathy, and anti-Factor VIII antibodies with coagulopathy (39, 48). It is not known why, in contrast to other autoimmune diseases, antibody-mediated autoimmune disease is primarily directed against the hematopoietic system in ALPS patients. However, autoimmune phenotyping is imprecise, as illustrated by the discovery of a patient, initially diagnosed with systemic lupus erythematosus (SLE), who was later found to have a Fas ligand mutation with defective apoptosis (49). An interesting question is whether the reduced CD4+ CD25+ cells seen in ALPS patients might represent a defect in regulatory cells that contributes to autoimmune disease. Although lymphoproliferation is initially nonmalignant, ALPS patients have a marked propensity to develop B or T cell malignancies. To date, we observed lymphomas in type I ALPS patients bearing dominant Fas DD mutations that cause severe apoptosis defects. We found a 14-fold and 51-fold increased incidence of non-Hodgkin lymphoma and Hodgkin lymphoma, respectively (50). The lymphomas were found anywhere between 15 to 48 years after onset of ALPS symptoms, with a median age of 21 for non-Hodgkin lymphoma and 11 for Hodgkin lymphoma (50; J. Dale & S. Straus, personal communication). Analysis of lymphoma tissue from ALPS patients showed no loss of Fas heterozygosity or increased apoptosis resistance. Interestingly, the responsible mutations impair apoptosis, but not Fas-induced NF-κB and mitogenactivated protein kinase (MAPK) signaling (51). Hence, the contribution of Fas mutations to lymphomagenesis may involve blocking death and releasing growth-promoting effects. It is also possible that the increased number of lymphocytes in general

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may provide a larger pool for tumorigenesis (52). In an early clinical description of ALPS, Sneller and colleagues (46) recognized that ALPS closely resembled the lpr/lpr (lpr) and gld/gld (gld) mouse strains. On certain inbred genetic backgrounds, homozygosity of these alleles causes mice to develop profound lymphoproliferation involving DNT cells, hypergammaglobulinemia, autoantibodies, autoimmune disease, and lymphomas (52, 53). Not surprisingly, these naturally arising mouse strains have molecular defects in some of the same genes as ALPS patients. The lpr and gld mice have autosomal recessive mutations in Fas and FasL, respectively, resulting in deﬁcient expression, whereas the lprcg variant has a point mutation in Fas that renders it nonfunctional (54, 55). Although the Fas locus is the primary genetic determinant, there is a strong effect of the inbred mouse strain background on the autoimmune phenotype. Severe disease occurs on the MRL background, whereas disease is substantially reduced on a B6 or BALB/c background. These mice do not develop the autoimmune hemolytic anemia and thrombocytopenia seen in human patients with ALPS. Instead, they display certain features resembling SLE (anti-dsDNA, anti-Sm, anti-immunoglobulin autoantibodies) with a predilection for developing glomerulonephritis, polyarteritis, sialoadenitis, and, with lesser frequency, arthritis or primary biliary cirrhosis (56, 57). Using intercrosses and backcrosses of diseasesusceptible and disease-resistant lpr strains, several groups have performed genome-wide linkage analysis (56). Mapping of genetic modiﬁers of disease revealed loci for lymphoproliferation and autoantibody production (i.e., Lmb1, Lmb2, and Lmb3), sialoadenitis (i.e., Asm and Asm2), glomerulonephritis (i.e., Agnm1, Agnm2, and Agnm3), and arthritis (i.e., Paam1 and Paam2) (56, 58). The Agnm3 locus associated with glomerulonephritis corresponds to allelic polymorphisms of the osteopontin gene. Polymorphisms of this gene, which is highly expressed in

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T cells, correspond to functional differences in activating macrophages and B cell antibody production (59). Interestingly, allelic polymorphisms of osteopontin may inﬂuence disease penetrance in humans who have a variant of ALPS termed ALD (described below) (60). In all but the most severely affected patients, the clinical course of ALPS waxes and wanes over time and often improves with age. Thymic involution may decrease the output of T cells, and it is known that neonatal thymectomy in lpr mice prevents disease (61, 62). Clinical complications relate mainly to the severity of autoimmune disease. Infections may occur after splenectomy or secondary to immunosuppressive treatment. Newer agents, such as mycophenolate mofetil, may bypass the need for prolonged courses of corticosteroids or splenectomy (39). Notably, pyrimethamine plus sulphadoxine has also ameliorated symptoms in some ALPS patients, possibly through a mechanism involving induction of lymphocyte apoptosis via an alternative mitochondrial pathway (63). There are published reports of curative bone marrow transplant in patients with Fas mutations on both alleles and intractable lymphoproliferative and autoimmune disease (64). However, conservative treatment generally remains sufﬁcient to control disease in most patients. Finally, minor clinical variants of ALPS may reﬂect different and uncharacterized molecular defects in lymphocyte homeostasis mechanisms. A signiﬁcant fraction of ALPS or ALPS-like patients has no mutations in the genes encoding proteins involved in the Fas signaling. Approximately half of these patients have no apoptosis defect following direct challenge with an agonist anti-Fas antibody (H. Su & M. Lenardo, unpublished results). Although the ALPS diagnosis requires documentation of impaired apoptosis of mature lymphocytes, most laboratories rely on assessment of apoptosis induced through the Fas death receptor. Defects in Fas-mediated apoptosis usually coincide with defects in TCR-restimulation-induced death;

however, an ALPS patient with normal Fasmediated apoptosis but impaired apoptosis following stimulation with phytohemagglutin plus IL-2 has been described (65). ALPS could also result from defects involving non– death receptor–induced intrinsic pathways of apoptosis. We have recently characterized an ALPS patient who displays the usual clinical features, but has cytokine withdrawal apoptotic defects with intact Fas-mediated death ( J.B. Oliveira & M. Lenardo, unpublished results). Patients can also exhibit the main clinical features as ALPS, with lymphoproliferation, apoptosis defects, autoimmune disease, and cancer, but lack DNT cell expansion. This ALPS variation has been termed autoimmune lymphoproliferative disease (ALD) (66). Although the responsible molecular defect(s) is (are) yet undiscovered, ALD patients likely share with ALPS patients a common affected pathway downstream of the Fas receptor. In summary, there are number of variant and overlapping clinical subtypes that will be extremely interesting to explore at the molecular level.

ALD: autoimmune lymphoproliferative disease

Molecular Diagnosis The molecular diagnosis of ALPS encompasses the following three, possibly four, major categories as well as some selected molecular subtypes (summarized in Table 2 and on the web at http://research.nhgri.nih. gov/ALPS/). For type Ia, the heterozygous mutations may be found in any part Table 2 Molecular classification of autoimmune lymphoproliferative syndrome ALPS classification

Gene mutated

Ia

Fas (TNFRSF6)

Ib

FasL (TNFSF6)

Ic or Im

Fas somatic mutation in DNT cells

II

Caspase-10

III

Molecularly undeﬁned

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PLAD: pre-ligand assembly domain

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of the Fas coding sequence, but over half are found in exon 9, which encodes the intracytoplasmic DD (see http://research.nhgri.nih.gov/ALPS/dist fas mutations.shtml). In general, the mutations permit the expression of a pre-ligand assembly domain (PLAD)-containing abnormal version of the Fas protein that has a dominant-interfering effect (see below; for review of the PLAD domain, see References 67 and 68). Nevertheless, there are exceptions to this rule. For example, some early exon 2 mutations essentially eliminate protein expression; this observation raises the question of whether haploinsufﬁciency may cause disease in certain circumstances. Recently, evidence supporting this hypothesis has been put forward (69). Type Ib involving FasL is much less common, but several cases are known (38, 70; S. Straus & M. Lenardo, unpublished observations). Molecular analysis shows that point mutations generate mutant versions of the FasL protein that are dominant interfering. Type Ic (or Im) is ALPS that results from somatic mutations in DNT cells. The possibility of somatic mutation in DNT should be investigated in any ALPS-like patient who does not seem to have a Fas, FasL, or caspase-10 mutation, especially those that do not have a clear-cut apoptosis defect in normal lymphocytes. Type II ALPS involves mutations in the caspase-10 gene (see http://research. nhgri.nih.gov/ALPS/alpsII mut.shtml and Reference 3). Originally, two cases were reported, although reinvestigation of the V410I mutation has revealed that it is widely distributed in Western European populations and brought into question its disease-causing potential (71). However, at least two more caspase-10 mutations have been detected in ALPS patients lacking Fas or FasL mutations ( J. Puck, S. Straus & M. Lenardo, unpublished observations). Originally, there was a suggestion that caspase-8 mutations may also cause type II ALPS. However, in the single family identiﬁed so far with caspase-8 mutations, the disease features constitute a distinct Bid`ere

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entity, as no dramatic increases in DNTs are observed; this fact may account for the lack of many hallmarks of ALPS. Additional families will be required to determine whether caspase8 mutations can cause a true ALPS phenotype. Until now, type III has been used to describe ALPS with an unknown molecular pathogenesis. This terminology is likely to change rapidly as new molecular abnormalities are uncovered. Hence, the cases with an unknown molecular pathogenesis will become type IV, etc. These undiagnosed cases are likely to harbor a wealth of interesting molecular insights into lymphocyte regulation and autoimmunity.

ALPS Type Ia Unveils Dominant Interference, Fas Preassociation, and Cooperativity ALPS Ia patients have heterozygous Fas mutations that impair apoptosis. More than 200 ALPS Ia patients have been described so far. The vast majority of them harbor mutations located in the region of the gene that encodes the intracellular domain of Fas, with a preponderance (60%) in the DD (38). These mutations are mostly missense or nonsense mutations that generate mutant proteins, as opposed to full loss-of-expression mutations. The mutant proteins have been shown to greatly reduce FADD binding and consequently impair recruitment and activation of caspases even in the presence of the wildtype protein (72). Why do dominant mutations have this effect? It does not appear to be due to selective expression of the mutant chains. Rather, the mutant chains enter complexes with the good chains, and these mixed complexes do not transduce the death signal effectively. In genetic terms, this constitutes dominant interference (73). It was presumed that the association of good and bad chains was due to the recruitment or crosslinking of the chains into mixed trimeric complexes by ligand binding. Mutations in the region of the gene that encodes the Fas extracellular domain also

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occur in ALPS type Ia. Roughly half of these mutations lead to truncated forms of Fas, and the other half result in an amino acid sequence modiﬁcation. Again, most of these mutations proved to be dominant interfering despite the puzzling fact that they failed to bind ligand (74, 75). These unanticipated observations led to the theory that Fas receptor chains formed trimers in a ligand-independent fashion (75, 76). It has now been directly demonstrated that Fas, TNFR1, and TNFR2 selfassociate as oligomers, independently of any ligand, through the PLAD, an approximately 50 amino acid N-terminal domain (75, 76). For Fas, the PLAD and ligand binding site are distinct. This fact explains why ALPS mutant chains with impaired FasL binding can form mixed trimers with wild-type Fas chains. The PLAD was found to be intact in all ALPS type Ia patients with dominant-interfering mutations. Moreover, Fas preassembly is necessary for FasL binding and correct DISC formation (75, 76). How is Fas preassociation linked to downstream signaling, and why does preassembled Fas not form a DISC in the absence of FasL? One can imagine two major allosteric conformations (or chain orientations): inactive and active. Binding of FasL would mediate an activating change that would propagate to the intracellular portion of Fas to induce a stoichiometric 3:3 Fas:FADD complex. Evidence suggests that this complex is cooperatively formed and highly stable (H. Wu & M. Lenardo, unpublished observations). The preassociation model has important implications in addition to explaining dominant interference in ALPS. It explains how distinct receptor chains that bind the same ligand, such as TNFR1 and TNFR2, can form homotypic complexes, as the PLAD from each receptor appears to be speciﬁc for self binding. Conformational change of preassembled dimeric receptors upon stimulation has been described for the erythropoietin receptor and other surface receptors (77). The fact that preassembly was also observed for TNFR1 and TNFR2 suggests that other members of the TNFR superfamily may have PLADs and are there-

fore susceptible to dominant-interfering mutations (76). Besides the dynamics of protein interactions in the DISC, higher order interactions such as those leading to SPOTS may also participate in ALPS signaling defects. Higher order complex formation may require cooperative interactions between individual receptor complexes in addition to cooperative recruitment of signaling proteins. For example, transient transfection of increasing amounts of Fas mutant alleles in Jurkat cells exerts a powerful transdominant interfering effect in preventing FADD and caspase association (74). The disruption of cooperative interactions may explain why dominant mutations—some of which only partially inhibit FADD:Fas binding—can markedly decrease apoptosis signaling. A DD mutation was found in an ALPS patient that permitted DISC formation but impaired SPOTS and subsequent caspase-8 cleavage. Cooperativity may serve to make signaling an all or none response to the level of receptor stimulation by ligand. Holler et al. (78) recently showed that two trimers of FasL are required to trigger a potent DISC formation. However, supercluster formation is also dependent on signals transduced from the intracellular portion of Fas to the extracellular domain (“inside-out” signaling) as reduced FADD and caspase-8 recruitment in ALPS Ia patients also abolishes SPOTS formation (30). Thus, cooperativity makes signaling efﬁcient but also vulnerable to dominant interference.

Caspase-10 as a Major Player in Death Receptor–Induced Apoptosis The recent report of two ALPS patients who bear caspase-10 mutations has unveiled the pivotal role of this protease during death receptor–induced apoptosis (3). The caspase-10 mutants show signiﬁcantly reduced enzymatic and autoprocessing activities, and lymphocytes from these patients exhibit an impaired Fas-, CD3-, and www.annualreviews.org • Genetic Disorders and Cell Death

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TRAIL-induced death. In line with these observations, caspases-8 and -10 are recruited and activated at the DISC level in FasLtreated PBL or Jurkat cells (79, 80). It is likely that these caspases cooperate, as mutants for caspases-8 and -10 could cross-inhibit one another and can enter the same receptor complex (80, 81). However, the relationships between these two very close caspases remain unclear. Although one study suggests that caspase-10 cannot substitute for caspase-8 in Fas- or TRAIL-induced death (82), some reports demonstrate that the two enzymes can trigger apoptosis independently from each other (79, 80). Two different roles for these proteases are conceivable, as their regions involved in substrate recognition and binding clearly differ (33). Moreover, caspase-10 as well as caspase-8 triggers cell death via Bid processing and caspase activation, but caspase-10 does not cleave receptor interacting protein (RIP), a canonical substrate for caspase-8 (80). Interestingly, one patient with a deleterious caspase-10 mutation exhibited an unprecedented defect in dendritic cell apoptosis in response to TRAIL that is associated with an overaccumulation of dendritic cells in the T cell areas of the lymph nodes (3). Other apoptosis-inactivating caspase-10 mutations were also described in non-Hodgkin lymphomas as well as in gastric cancers (83, 84).

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Type III and Variant ALPS There are two important categories of patients within the spectrum of ALPS that do not easily fall into the aforementioned groups. The ﬁrst group is referred to as type III ALPS. These patients meet ALPS clinical criteria and exhibit a mild resistance to Fasinduced apoptosis but do not carry any mutations in the genes that encode Fas, FasL, FADD, caspase-8, or caspase-10. In some of these patients, FADD and caspase-8 are normally recruited to the DISC following Fas crosslinking. However, DISC formation is abortive, as caspase-8 remains largely un332

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processed at the receptor level (N. Bid`ere, J.B. Oliveira & M. Lenardo, unpublished results). This observation suggests that the processing and release of active caspase-8 from the DISC is tightly regulated, although the complete mechanism is unknown. We hypothesize that mutations in such regulators might also lead to ALPS development. Among possible regulators, c-FLIP is an appealing candidate as it blocks caspase-8 activation at the receptor level. However, transgenic mice that overexpress FLIPL in the T cell compartment do not exhibit T cell accumulation in secondary lymphoid organs as in MLR/lpr mice (85). It is also plausible that defects in the molecules controlling the conformational change of Fas or its aggregation and internalization might lead to the disease. Other less-characterized cytoplasmic partners of Fas, such as Daxx, RIP, or FAF1, might be involved. The DD-containing serine/threonine-containing kinase RIP has been described as a component of the Fas DISC (86). RIP seems to play a critical role in the nonapoptotic death following Fas stimulation when caspases are inhibited (16). Another category of patients, called variant ALPS, exhibits the phenotypic characteristics of ALPS despite their lymphocytes’ normal sensitivity to Fas killing.

Genetic Modifiers of ALPS Disease ALPS exhibits variable disease penetrance and severity. Over two thirds of patients have heterozygous mutations in Fas and exhibit a range of phenotypes from no symptoms at all to lifethreatening autoimmunity. A major factor in disease phenotype appears to be the nature of the Fas mutation and how it affects apoptosis signaling. One patient was found to be a compound heterozygote of two Fas mutations and presented a severe clinical phenotype (87). Among patients with heterozygous Fas mutations, the location of the mutation affects penetrance and severity. For instance, dominantinterfering alleles encoding DD alterations that severely affect apoptosis signaling are

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associated with a greater penetrance and disease severity (75, 88). By contrast, mutations in the region of the gene that encodes the extracellular domain, even if dominant interfering, are less penetrant and cause milder disease. A minority of patients who have haploinsufﬁcient mutations in the region of the gene that encodes the extracellular domain tends also to have milder disease. Nonetheless, even within these broad groups, there is signiﬁcant heterogeneity. For any given patient, it is not uncommon that multiple family members possessing the same mutation manifest impaired apoptosis in vitro but do not have ALPS disease. This situation has been mimicked by generating mice transgenic for human Fas DD mutations that were backcrossed onto FVB/N or MRL strains (89). Compared to transgenic FVB/N or MRL strains, FVB/N X MRL mice transgenic for identical Fas mutations had incompletely penetrant autoimmune disease that correlated with degree of apoptotic defect (89). The existence of healthy family members of ALPS patients and the inﬂuence of the inbred genetic background on the phenotype of lpr and gld mice strongly suggests the existence of modiﬁer genes. A modiﬁer gene could be deﬁned as a gene encoded by a locus unlinked to the primary ALPS mutation that determines the ultimate disease phenotype. Several modiﬁer or susceptibility genes have been linked to other autoimmune diseases and are therefore candidates for ALPS modiﬁers. Perhaps the most common is the association of speciﬁc human leukocyte antigen (HLA) alleles with autoimmune diseases (90). However, analyses of large numbers of ALPS patients and their relatives have not yet yielded any speciﬁc HLA associations with disease exacerbation, although HLA-B44 may have a protective effect (M. Vacek & J. Puck, unpublished observations). This result is surprising as HLA-B44 had been previously linked to Crohn’s disease (91). Certain inherited forms of caspase-10 may also act as genetic modiﬁers of disease in ALPS patients (S. Zhu, L. Zheng, M. Lenardo & J. Puck, unpublished

results). In addition to caspase-10, other cellkilling molecules may modulate disease in the presence of Fas mutations. A patient was recently described with a compound heterozygous Fas splicing mutation and a perforin mutation, which were inherited separately from healthy parents (92). However, it was unclear if the Perforin alteration represented a polymorphism or functional mutation. In a separate family, the identical Perforin alteration was found together with a Fas mutation, yet that individual did not have ALPS (93). Cytotoxic T lymphocyte–associated antigen4 (CTLA-4) polymorphisms have been associated with autoimmune thyroid disease and type 1 diabetes mellitus (94). However, analysis of a small number of ALPS patients revealed no relationship to CTLA-4 polymorphisms (L. Wicker, personal communication). Besides modifying ALPS disease penetrance, other genes may affect the clinical presentation of disease. Of 151 ALPS patients currently followed at the National Institutes of Health, several have other superimposed inﬂammatory conditions. Periodic fevers suggestive of TNF-receptor-associated periodic syndrome (TRAPS) were observed in six ALPS patients, and two of these individuals were found to have a R92Q functional polymorphism in the p55 TNF-R1 gene (I. Aksentijevich, R. Siegel, S. Straus & D. Kastner, personal communication). The carrier frequency of this polymorphism is higher in TRAPS and other disorders such as rheumatoid arthritis when compared with the general population (95). The biochemical mechanism by which the R92Q polymorphism contributes to disease is unclear. A similar situation exists for another ALPS patient, who also has Familial Mediterranean Fever with an associated functional polymorphism of the pyrin gene (E148Q) (96). Moreover, one of the ALPS patients with the R92Q polymorphism for p55 TNF-R1 has a caspase10 gene alteration; this fact suggests that, by modifying coexisting inﬂammatory conditions, these genes may inﬂuence ALPS disease penetrance and severity. www.annualreviews.org • Genetic Disorders and Cell Death

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CASPASE-8-DEFICIENCY STATE (CEDS)

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Clinical Presentation The study of a wide spectrum of patients exhibiting some features of ALPS has revealed new and unexpected biological roles for molecules that function in programmed cell death. We found two siblings with several ALPS features who surprisingly also fulﬁlled diagnostic criteria for common variable immunodeﬁciency (CVID) (2). Remarkably, these children were found to have homozygous inactivating mutations in caspase-8 (2). It was surprising that the caspase-8-deﬁciency state (CEDS) was compatible with embryonic development in humans as the caspase-8 gene knockout caused embryonic lethality in mice (97). One explanation could be the presence in humans, but not in mice, of the highly related caspase-10. The CVID-affected children had recurrent sinopulmonary infections and mucocutaneous herpesvirus infections. Despite elevated lymphocyte numbers, serum immunoglobulin levels were reduced, and immunization with polysaccharide antigen yielded poor antibody responses. Moreover, activation of B and T cells (through antigen receptors) or natural killer (NK) cells (through Fc - or NK-activating receptors) were all impaired. Although immunodeﬁciency was their most striking clinical feature, the patients also had secondary lymphoid enlargement, impaired lymphocyte apoptosis in vitro, and occasional borderline elevations in DNT cells (2). Although caspase-8 mutations explained abnormal apoptosis and lymphocyte homeostasis, the clinical ﬁndings of immunodeﬁciency also pointed to an unappreciated role for caspase-8 in lymphocyte activation. By using a novel transfection technology to introduce caspase-8-speciﬁc small-interferingRNAs into primary human lymphocytes, we discovered that caspase-8 was indeed required in normal lymphocytes for activation through the T and B cell antigen receptors, the Fc and 2B4 receptors of NK cells, and TollBid`ere

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like receptor 4 (TLR4) (2, 4). Our observations explained the puzzling ﬁnding that pharmacologic caspase-inhibitors could interfere with T lymphocyte activation in vitro (98). These ﬁndings have also been essentially conﬁrmed in mice in which T cells or B cells were conditionally rendered caspase-8 deﬁcient by homologous recombination (99; R. Hakem, personal communication). The T cell–knockout mice eventually succumb to an unusual autoimmune disease characterized by lymphocytic inﬁltration of lung, kidney, and liver parenchyma (L. Salmena & R. Hakem, personal communication). Clinical follow-up of the caspase-8-deﬁcient patients has revealed new bronchiectasis, as well as autoantibodies or a compensated autoimmune hemolytic anemia (H. Su, S. Straus & K. Rao, unpublished observations). Whether these new symptoms reﬂect complications of CVID, which in itself can be associated with autoimmune disease, and/or a superimposed forme fruste of ALPS is presently unclear. The resulting clinical syndrome shares certain features with ALPS, but clearly has unique features. Its striking clinical hallmark is a primary combined lymphoid immunodeﬁciency with susceptibility to infection. Importantly, despite an extensive literature on caspase-8 and Fas-induced programmed cell death in nonlymphoid tissues, so far CEDS patients have not manifested primary abnormalities in other organ systems. In summary, these ﬁndings underscore the importance of examining critical pathways in humans rather than relying upon mice or other model organisms. Clearly, there are important limitations to the use of studies in mice for the elucidation of human disease.

Molecular Effects of Caspase-8 Deficiency The possible involvement of caspase-8 in antigen receptor–induced lymphocyte activation was puzzling at ﬁrst glance because the known functions of caspases were limited to cytokine maturation and mediation of programmed

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cell death (25). The pivotal role of caspase8 in both activation and death of lymphocytes was ﬁnally clearly demonstrated (2, 4, 99). These observations extended the functions of caspases to include roles as intracellular signaling mediators during cell activation and proliferation. The implications of the evolution of these dual roles for caspases and other death molecules have been previously discussed (23). These observations also raised the question of the molecular pathway on which caspase-8 exerts its effect during lymphocyte activation. Recently, we conclusively demonstrated that caspase-8 is essential for antigen receptor–induced NF-κB activation (4). During the antigenic stimulation of T cells, the transcription factor NF-κB induces genes that regulate cellular proliferation and cell survival, such as IL-2 and CD25 (highafﬁnity IL-2 receptor α-chain) (100). The DNA-binding components of NF-κB are constitutively present in the cytosol bound to speciﬁc inhibitory proteins called inhibitors of κB (IκBs). NF-κB activation involves the phosphorylation and ubiquitin-mediated degradation of IκBs. The degradation of IκB enables NF-κB to translocate to the nucleus and govern the transcription of a large number of genes by binding to speciﬁc DNA sites in gene regulatory regions. The phosphorylation of IκBs is carried out by the IκB kinase (IKK) complex, which is composed of two catalytically active kinases (IKKα and IKKβ) and a catalytically inactive regulatory subunit (IKKγ, also known as NEMO) (101). How the signal is conveyed from antigen receptor to IKK complex has not been fully elucidated. However, the recent discovery of three critical proteins, CARMA1, BCL10, and MALT1, upstream of IKK has shed new light on the NF-κB activation pathway (102). It is now accepted that the Ca2+ -independent protein kinase C (PKC) θ is rapidly recruited to membrane lipid rafts at the immunological synapse following antigen receptor stimulation (Figure 2). PKCθ then triggers IKK acti-

vation by initiating the formation of a large complex that includes CARMA1, BCL10, and MALT1, termed the CBM complex. Genetic and biochemical approaches have established the pivotal role of CARMA1 upstream of a preassociated BCL10 and MALT1 complex (102). Recently, Lee et al. (103) have shown that the 3-phosphoinositide-dependent kinase 1 (PDK1) is a bridge linking the IKK and CBM complexes to PKCθ. How the CBM complex activates IKK is still unclear. It is thought that IKK activation requires the noncanonical K63-linked ubiquitylation of NEMO (102). This could be achieved by the binding of the ubiquitin-conjugating E2 complex (composed of UBC13 and MMS2) to MALT1 after its oligomerization or, alternatively, through the binding of the RING domain ubiquitin ligase TRAF6. This modiﬁcation of NEMO would induce the phosphorylation and activation of IKK. Our evidence now shows that caspase-8 appears to be an essential nexus between the CBM and IKK complexes (Figure 2). This protease physically interacts with both the CBM and IKK, and its absence precludes the entire formation of the activation-receptor induced signalosome (ARIS) and prevents de facto IKK activation (4). This unexpected role for caspase-8 in lymphocyte activation explains the characteristic immunodeﬁciency in CEDS patients. Defective induction of NFκB during the activation of T, B, and NK cells will severely incapacitate critical gene expression events. How caspase-8 protein assembles the ARIS and what comprises the ﬁnal signaling complex is still unknown. Mutation of the active site cysteine inhibits the NF-κB activating function of caspase-8; this result clearly implies that the complex includes a crucial proteolytic substrate. The identiﬁcation of this substrate will provide considerable insight into how the pathway works. The adaptor FADD is also involved, although there is no evidence that it is cleaved. FADD is transiently recruited to the CBM, and whether it plays a key role in the assembly of the ARIS needs to be further explored. www.annualreviews.org • Genetic Disorders and Cell Death

IKK: IκB kinase CBM complex: CARMA1-BCL10MALT1 complex ARIS: activation-receptor induced signalosome

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Genetic deﬁciency of FADD or transgenic expression of a dominant-interfering mutant of FADD have been found to impair both death receptor–induced and mitogen- and antigeninduced T cell activation (24, 104). Future work should also be aimed at determining whether the caspase-8 regulator FLIP is part of ARIS and whether it regulates caspase-8 activation or downstream signaling events, as has been previously suggested (32). Interestingly, enzymatic activity but not autoprocessing of caspase-8 is required during antigen receptor–induced NF-κB activation (4). It is likely that caspase-8 activity generated within the ARIS is weaker than during DISC formation, as the enzyme remains unprocessed. Furthermore, evidence suggests that there is no release of the active caspase-8 from the ARIS, as compared with DISC formation, where the active enzyme is proteolytically released from the prodomain. The local and restricted caspase-8 activity in the ARIS likely triggers pro-survival events and avoids release of the protease into the cytosol, where it might ﬁnd apoptosis substrates. Thus, depending on its location, its concentration, and its partners, caspase-8 controls the fragile balance between life and death in lymphocytes during immune responses.

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RELATED DISORDERS OF LYMPHOCYTE HOMEOSTASIS The use of knockout mice has enabled the identiﬁcation of several other genes that regulate programmed cell death during normal lymphocyte homeostasis. These genes may provide clues to human diseases that have not yet been uncovered. Table 3 lists mouse strains that exhibit lymphocyte apoptotic defects with lymphoproliferation or other characteristics suggestive of ALPS in humans. Because approximately 20% of ALPS (type III) patients have no identiﬁed gene defect, the mouse studies may provide candidates as to potential defects in these individuals. Several themes emerge, which we discuss below. 336

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Abnormal IL-2 signaling results in an ALPS-like picture in mice. Mice genetically deﬁcient in IL-2 or the IL-2 receptor subunits IL-2Rα (CD25) and IL-2Rβ (CD122) develop autoimmune hemolytic anemia, colitis, lymphoproliferation, and early death (105– 107). The one reported human with a deleterious IL-2Rα mutation had severe multisystem autoimmune disease (108). Despite its growth-promoting properties, IL-2 is also required for programmed death of T cells after antigen receptor restimulation. Defective death following TCR restimulation was directly shown when T cells were initially treated with limiting doses of IL-2 (109) and was conﬁrmed using cells from IL-2−/− or IL2Rα−/− mice (19, 110). Defective death mediated through the Fas death receptor was also observed in T cells from IL-2−/− or IL-2Rα−/− mice (112, 113). Apoptotic defects occurred in vivo, as administration of anti-IL-2 neutralizing antibodies with antigen resulted in persistence of adoptively transferred antigen-speciﬁc TCR-transgenic T cells (110), and impaired peripheral deletion of Vβ8+ T cells was seen after administration of the bacterial superantigen Staphylococcus enterotoxin B (SEB) in mice treated with anti-IL-2Rα blocking antibodies (109). These results were conﬁrmed in similar mouse models (107, 112, 114). IL-2 regulates numerous genes known to affect apoptosis (19, 115), and slight differences between these mouse models may reﬂect variations in the molecular targets perturbed by impaired signaling downstream of IL-2. Subsequent studies seemed to challenge the idea that an intrinsic inability of T cells to die in the absence of proper IL-2 signaling contributes to disease, but they failed to consider the possibility of compensatory mechanisms for cell death. T cell hyperactivation and hematopoietic abnormalities in IL-2Rβ −/− mice can be corrected by adoptive transfer of highly puriﬁed CD8+ CD122+ cells expressing TNF, Perforin, and Granzyme B but not requiring Fas or FasL (116, 117). By contrast, adoptive

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Table 3

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Mutant mouse strains with phenotypes resembling human autoimmune lymphoproliferative syndrome

Mouse strain

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Molecular target affected

Lymphoproliferation

Apoptotic defect

Other features, including those suggestive of ALPS

References

lpr/lpr, lprcg

Fas

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation or Fas stimulation

Autoantibodies Hypergammaglobulinemia Glomerulonephritis Sialitis Arthritis

(144–146)

gld/gld

FasL

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation Impaired apoptosis after stimulation with FasL from gld mice

Autoantibodies Hypergammaglobulinemia Glomerulonephritis

(144, 147, 148)

IL-2−/−

Cytokine

Splenomegaly Lymphadenopathy

Impaired Fas-mediated apoptosis or peripheral deletion after SEB Persistence after TCR stimulation Increased c-Flip expression

Autoimmune hemolytic anemia Colitis Hypergammaglobulinemia

(105, 110, 112)

IL-2Rα−/−

IL-2 receptor subunit

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation Impaired peripheral deletion after SEB administration Impaired Fas-mediated apoptosis (in IL-2Rα−/− X DO.11)

Autoimmune hemolytic anemia Colitis Hypergammaglobulinemia

(19, 107, 113)

IL-2Rβ−/−

IL-2/IL-15 receptor subunit

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation

Autoimmune hemolytic anemia Colitis Hypergammaglobulinemia Lack of NK cells

(106)

p65PI3K transgene under T cellspeciﬁc p56 lck promoter

Constitutively active phosphoinositide 3-kinase truncation mutation of p85α regulatory subunit

Splenomegaly Lymphadenopathy

Decreased Annexin V and subG1 staining of expanded CD4 cells

Autoantibodies Hypergammaglobulinemia Glomerulonephritis Lymphoma

(124)

(Continued )

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(Continued )

Mouse strain

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Molecular target affected

Lymphoproliferation

Apoptotic defect

Other features, including those suggestive of ALPS

References

PKB transgene under T cellspeciﬁc CD2 or lck promoter

Constitutively active myristolated protein kinase

Splenomegaly Lymphadenopathy

Increased viability after γ irradiation Increased viability after dexamethasone Increased viability in culture Impaired Fas-mediated death

Hypergammaglobulinemia Autoantibodies Glomerulonephritis

(125–127)

PTEN+/− , PTEN−/− , − PTENﬂox/ (in T cells), bPTENﬂox/ﬂox

Phosphatase

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation or Fas stimulation Decreased baseline B cell Annexin V staining Impaired apoptosis after Fas stimulation Impaired apoptosis after γ or UV irradiation Impaired apoptosis after IL-2 (±serum) withdrawal Impaired apoptosis after anti-IgM stimulation (in conditionally deleted mice)

Autoantibodies Glomerulonephritis Hypergammaglobulinemia Heterogeneous tumors including lymphomas

(128–132)

caspase-3−/− reconstituted into RAG2−/−

Effector caspase

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation Impaired Fas-mediated apoptosis

Central nervous system defects in caspase-3−/− mice B cell hyperproliferation due to dysregulated cell cycle in caspase-3−/− mice

(140, 141)

tcaspase-8ﬂ/ﬂ

Initiator caspase

Splenomegaly Lymphadenopathy

Impaired Fas-mediated apoptosis

T cell immunodeﬁciency with impaired NF-κB activation

(4, 99)

Bim−/−

Proapoptotic Bcl-2 family member

Splenomegaly Lymphadenopathy

Impaired apoptosis after IL-2 withdrawal Impaired peripheral deletion after SEB Impaired BCR-induced death

Autoantibodies Glomerulonephritis Hypergammaglobulinemia

(133, 150, 151)

(Continued ) 338

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(Continued )

Mouse strain

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Molecular target affected

Lymphoproliferation

Apoptotic defect

Other features, including those suggestive of ALPS

References

Bax−/−

Proapoptotic Bcl-2 family member

Splenomegaly

Impaired apoptosis after anti-IgM stimulation Improved viability of Bax−/− T cells (without mitogens)

Aspermatogenesis Increased atrophic granulosa cells in ovaries

(134, 135)

Bax−/− X Bak−/−

Proapoptotic Bcl-2 family members

Splenomegaly Lymphadenopathy

Impaired death in culture (without mitogens)

Persistence of interdigital webs Imperforate vaginal introitis Accumulation of cells in central nervous system

(136)

Bcl-2 transgene under B cell-speciﬁc immunoglobulin promoter

Anti-apoptotic Bcl-2 family member

Splenomegaly Lymphadenopathy

Increased viability in culture

Hypergammaglobulinemia Autoantibodies Glomerulonephritis Lymphoma

(137, 138)

Mutant cytochrome c K72A/K72A, reconstituted into RAG1−/−

Mitochondrial component of the apoptosome

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation Impaired apoptosis after γ or UV irradiation

Central nervous system defects in cytochrome c K72A/K72A mice

(139)

Stra13−/−

Transcription factor affecting IL-2

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation Decreased FasL expression

Autoantibodies Glomerulonephritis

(152)

T cell-speciﬁc adapter protein (TSAd)−/−

Adaptor-like molecule in T cells, transcriptional regulator affecting IL-2

Splenomegaly Lymphadenopathy

Impaired peripheral deletion after SEB

Autoantibodies Glomerulonephritis Hypergammaglobulinemia

(153)

IRF-4−/−

Transcription factor

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation or peripheral deletion after SEB

T cell functional defect Hypogammaglobulinemia

(154)

(Continued )

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Table 3

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(Continued ) Molecular target affected

Lymphoproliferation

Apoptotic defect

E2F-1−/−

Transcription factor

Splenomegaly Lymphadenopathy

Impaired apoptosis after TCR restimulation

Lymphoma and other tumors Testicular atrophy Exocrine gland dysplasia Anemia Glomerulonephritis

(155–157)

Par4−/−

Atypical protein kinase C inhibitor Inhibits NF-κB activation as well

Splenomegaly

Impaired apoptosis after TCR restimulation

T cell hyperproliferation and Th2 differentiation

(143)

Mutant LAT Y136F/Y136F

Adaptor protein in T cells

Splenomegaly Lymphadenopathy

Impaired Fas-mediated apoptosis Decreased Fas and FasL expression Decreased IL-2 production

Lymphoproliferation differs from the T cell functional defect in LAT−/− mice

(158)

NFAT1−/− X NFAT4−/−

Transcription factors

Splenomegaly Lymphadenopathy (greater than in NFAT1−/− mice)

Impaired apoptosis after TCR restimulation Decreased FasL expression also seen in NFAT1−/− mice or NFAT1−/− X NFAT2−/−

Allergic blepharitis Pneumonitis Hypergammaglobulinemia

(159–161)

Act1−/−

Adaptor molecule

Splenomegaly Lymphadenopathy

Decreased Annexin V staining of B cells after CD40L stimulation

Hypergammaglobulinemia Autoantibodies

(162)

Mouse strain

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transfer of highly puriﬁed CD8+ CD122− cells exacerbated disease. Since in vivo antibody depletion of CD4 T cells had previously been shown to reduce autoantibody production and hemolytic anemia (106), these results suggest that CD8+ CD122+ cells constitute a potent regulatory population equipped to kill disease-causing T cells. IL-2Rβ −/− mice lack CD4+ CD25+ cells. Although adoptive transfer of populations enriched in CD4+ CD25+ cells corrected disease in IL-2Rβ −/− mice (118), highly puriﬁed CD4+ CD25+ cells 340

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Other features, including those suggestive of ALPS

References

were clearly less capable than CD8+ CD122+ cells (116). Recent studies have shown that CD4+ CD25+ regulatory T cells can exert suppressive effects through Granzyme B or Perforin (8, 9); these studies raise the possibility that these cells may act by killing disease-causing T cells. IL-2−/− mice also lack CD4+ CD25+ cells, and adoptive transfer of wild-type CD4 cells seemed equally or slightly more efﬁcacious than CD8 cells in ameliorating disease (119). However, this study was limited by small sample sizes and large

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mouse-to-mouse variability, as well as by the failure to separate CD122+ and CD122− cells in the CD8 preparation used for adoptive transfer. Hence, these studies did not refute the importance of IL-2 for induction of apoptosis in T cells. The role of regulatory cells as a potential alternative mode of lymphocyte deletion and tolerance will be an interesting topic for future investigation. These results collectively lead to the prediction that related cytokines using the common gamma chain (γc ) receptor component, which also participates in IL-2 signaling, promote apoptosis in T cells. Although several groups initially reported that other cytokines could not substitute for IL-2, we showed that given sufﬁciently high concentrations to drive T cells into cell cycle, prior exposure of IL2Rα−/− cells to IL-4 or IL-7 restored death after TCR restimulation (19). Other investigators have also shown that IL-2, IL-4, IL7, and IL-15 could promote death of IL-2−/− cells and that, as expected, IL-4 and IL-7 but not IL-2 or IL-15 could promote death of IL-2Rβ −/− cells (IL-2Rβ also is IL-15Rβ and serves as a component of IL-15R) (120). Mice or humans genetically deﬁcient in γc display severe immunodeﬁciency and generally die of infections early in life unless curative bone marrow transplantation is undertaken. However, aging mice do develop splenomegaly with CD4 T cell accumulation, as well as inﬂammatory bowel disease lesions (121). Similarly, there is a case report of an incompletely immunodeﬁcient human patient with a splicing mutation in the γc chain who had peripheral lymphocytes and developed severe colitis (122). Mice deﬁcient in γc were found to have impaired peripheral deletion after SEB administration, with decreased Bcl-2 and FasL expression (although they maintained Fas sensitivity) (123). Interestingly, mice deﬁcient in IL-4, IL-4Rα, IL-7, IL-7Rα, IL-15, or IL-15Rα have not been reported to develop lymphoproliferative or autoimmune disease. Taken together, these ﬁndings underscore the critical importance of IL-2 signaling, in particular, for lymphocyte homeostasis in vivo.

Although studies in ALPS patients have established the physiologic importance of molecules involved in the extrinsic pathway of apoptosis controlled by death receptors, no comparable studies exist in humans for molecules participating in the intrinsic mitochondrial pathway of apoptosis. Mice rendered genetically deﬁcient in many of these candidate molecules have been generated, and several display phenotypes that suggest these genes may contribute to uncharacterized forms of ALPS in humans. For instance, following growth factor receptor stimulation, phosphoinositide-3 kinase (PI3K) generates the lipid second messenger PIP3 , which activates the serine/threonine protein kinase B (PKB)/Akt. PKB/Akt in turn promotes cell survival through multiple effects, including inhibition of translocation of the pro-death Bcl-2 family members Bax and Bak to the mitochondria. Bax and Bak activate the intrinsic apoptosis pathway by permeabilizing the mitochondria and thus releasing cytochrome c to associate with Apaf-1 and caspase-9, which leads to caspase activation. Other Bcl-2 family members such as Bim or Bid can regulate the activity of Bax and Bak. Bid is particularly interesting because it can feed the extrinsic pathway of death into the intrinsic pathway through the ability of caspase-8 to activate Bid. In this general framework, mice transgenic for T cell–speciﬁc, constitutively active mutant forms of PI3K or PKB/Akt developed lymphoproliferation, autoimmune disease, lymphomas, and defective apoptosis (124–127). Phosphatase and tensin homolog (PTEN) limits the available pool of PIP3 and thus has proapoptotic effects. Mice in which PTEN was heterozygously or conditionally deleted in lymphocytes developed a similar ALPS-like phenotype (128–132). Molecules acting further downstream in the intrinsic pathway have been evaluated by rendering mice genetically deﬁcient in various proapoptotic Bcl-2 family members. Mice deﬁcient in Bax or Bim develop lymphoproliferative disease, and, in the case of Bim, autoimmune disease as well (133–135). Impaired www.annualreviews.org • Genetic Disorders and Cell Death

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lymphocyte apoptosis could be demonstrated when Bax-deﬁcient mice were intercrossed with Bak-deﬁcient mice (136). Transgenic expression of the anti-apoptotic Bcl-2 in B cells also caused an ALPS-like disease (137, 138). For genes that cause embryonic lethality, a role in mature lymphocyte homeostasis was assessed by reconstitution into recombinase activating gene (RAG)-deﬁcient mice, which lack endogenous T and B cells. Using this approach, adoptive transfer of mutant cytochrome c knockin cells gave rise to lymphoproliferative disease and impaired apoptosis following TCR restimulation, gamma-, or UV-irradiation (139). Adoptive transfer of caspase-3-deﬁcient cells also resulted in lymphoproliferative disease and impaired apoptosis to TCR restimulation or Fas (140, 141), as would be expected for an effector caspase that participates in both the intrinsic and extrinsic pathways of apoptosis. Finally, other knockout mice have been generated in which the role of certain transcription factors and other regulators for programmed cell death have been less well characterized. Based on the discussion above, these proteins may regulate IL-2 or other various proapoptotic and anti-apoptotic molecules functioning in extrinsic or intrinsic apoptosis pathways. In this light, it is interesting that signaling through the Fas death receptor or through TCR results in NF-κB activation, which normally antagonizes simultaneous death effects (142). This mechanism may contribute to lymphomagenesis in ALPS patients (51). Thus, gene mutations that affect NF-κB activation can perturb lymphocyte homeostasis through their effects on apoptosis. For example, mice rendered genetically deﬁcient in Par-4, an atypical PKC inhibitor that normally inhibits NF-κB activation, exhibit hyperproliferation (143). In summary, mice rendered genetically deﬁcient by homologous recombination have been useful in identifying potential genes that govern lymphocyte homeostasis and tol-

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erance. In particular, genes involved in IL2 signaling, genes involved in the intrinsic and extrinsic pathways for apoptosis, and certain transcription factors, including those that regulate NF-κB activation, are likely to be responsible for some cases of ALPS type III.

CONCLUSION The study of humans with rare genetic defects has been made possible by new technological breakthroughs that have enabled unprecedented molecular manipulation and rigorous scientiﬁc analyses. These studies are complemented by knowledge of disease pathogenesis gleaned from medical studies of individual patients and patient populations. Two such examples of this powerful new approach are discussed in this review. Studies in patients with ALPS and CEDS have shed considerable light into the normal signal transduction events that lead to programmed cell death, activation, and lymphocyte homeostasis in humans. These studies have led to a number of insights not previously appreciated from work done in model organisms. ALPS patients show that the consequences of defective apoptosis are lymphoproliferative disease, autoimmunity, and lymphoma. Although identiﬁcation of genetic disease modiﬁers remains unresolved, CD4− CD8− αβ T cells clearly play an important role for this disease pathogenesis. An understanding of the biochemical events leading to apoptosis explains how dominant mutations cause disease in most ALPS patients. At the molecular level, we now know that Fas death receptor preassociation and its higher order interactions are essential for apoptosis induction. Caspase-10, which is found in humans but not in mice, is also a critical component. By contrast, its related molecule, caspase-8, though contributing to death, serves a more important role in the activation of lymphocytes through antigen receptors and other immunoreceptors

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through its effects on NF-κB. The study of other ALPS patients with as yet unidentiﬁed genetic defects and patients with defects in other pathways of programmed cell

death may prove to be equally illuminating for understanding the molecular underpinnings of this basic biological process in humans.

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ACKNOWLEDGMENTS We thank our long-term collaborators Stephen Straus, Janet Dale, Jennifer Puck, Joie Davis, Elaine Jaffe, Thomas Fleisher, Faith Dugan, Koneti Rao, and Roxanne Fischer. We thank Richard Siegel for helpful suggestions and discussions and Lixin Zheng, Joao Bosco Oliveira, and Keiko Sakai for critically reading the manuscript. We apologize to many investigators whose important work was not cited owing to editorial constraints. This work was supported by the Intramural Research Program of the NIH, NIAID, as well as the Cancer Research Institute (HS).

LITERATURE CITED 1. Lenardo MJ. 2003. Molecular regulation of T lymphocyte homeostasis in the healthy and diseased immune system. Immunol. Res. 27:387–98 2. Chun HJ, Zheng LX, Ahmad M, Wang J, Speirs CK, et al. 2002. Pleiotropic defects in lymphocyte activation caused by caspase-8 mutations lead to human immunodeﬁciency. Nature 419:395–99 3. Wang J, Zheng L, Lobito A, Chan FK, Dale J, et al. 1999. Inherited human caspase 10 mutations underlie defective lymphocyte and dendritic cell apoptosis in autoimmune lymphoproliferative syndrome type II. Cell 98:47–58 4. Su H, Bid`ere N, Zheng LX, Cubre A, Sakai K, et al. 2005. Requirement for caspase-8 in NF-κB activation by antigen receptor. Science 307:1465–68 5. Lenardo M, Chan FK-M, Hornung F, McFarland H, Siegel R, et al. 1999. Mature T lymphocyte apoptosis–immune regulation in a dynamic and unpredictable antigenic environment. Annu. Rev. Immunol. 17:221–53 6. Lenardo MJ. 1996. Fas and the art of lymphocyte maintenance. J. Exp. Med. 183:721–24 7. Strasser A, Pellegrini M. 2004. T-lymphocyte death during shutdown of an immune response. Trends Immunol. 25:610–15 8. Grossman WJ, Verbsky JW, Barchet W, Colonna M, Atkinson JP, et al. 2004. Human T regulatory cells can use the perforin pathway to cause autologous target cell death. Immunity 21:589–601 9. Gondek DC, Lu LF, Quezada SA, Sakaguchi S, Noelle RJ. 2005. Cutting edge: contactmediated suppression by CD4+ CD25+ regulatory cells involves a granzyme B-dependent, perforin-independent mechanism. J. Immunol. 174:1783–86 10. Brunner T, Mogil RJ, LaFace D, Yoo NJ, Mahboubi A, et al. 1995. Cell-autonomous Fas (CD95)/Fas-ligand interaction mediates activation-induced apoptosis in T-cell hybridomas. Nature 373:441–44 11. Dhein J, Walczak H, Baumler C, Debatin KM, Krammer PH. 1995. Autocrine T-cell suicide mediated by APO-1/(Fas/CD95). Nature 373:438–41 12. Ju ST, Panka DJ, Cui H, Ettinger R, el-Khatib M, et al. 1995. Fas(CD95)/FasL interactions required for programmed cell death after T-cell activation. Nature 373:444–48 13. Zheng L, Fisher G, Miller RE, Peschon J, Lynch DH, et al. 1995. Induction of apoptosis in mature T cells by tumour necrosis factor. Nature 377:348–51 www.annualreviews.org • Genetic Disorders and Cell Death

343

ARI

23 February 2006

15:22

14. Muppidi JR, Siegel RM. 2004. Ligand-independent redistribution of Fas (CD95) into lipid rafts mediates clonotypic T cell death. Nat. Immunol. 5:182–89 15. Chan FK-M, Shisler J, Bixby JG, Felices M, Zheng LX, Lenardo MJ. 2003. A role for tumor necrosis factor receptor-2 and receptor-interacting protein in programmed necrosis and antiviral responses. J. Biol. Chem. 278:51613–21 16. Holler N, Zaru R, Micheau O, Thome M, Attinger A, et al. 2000. Fas triggers an alternative, caspase-8-independent cell death pathway using the kinase RIP as effector molecule. Nat. Immunol. 1:489–95 17. Locksley RM, Killeen N, Lenardo MJ. 2001. The TNF and TNF receptor superfamilies: integrating mammalian biology. Cell 104:487–501 18. Suda T, Takahashi T, Golstein P, Nagata S. 1993. Molecular cloning and expression of the Fas ligand, a novel member of the tumor necrosis factor family. Cell 75:1169–78 19. Zheng L, Trageser CL, Willerford DM, Lenardo MJ. 1998. T cell growth cytokines cause the superinduction of molecules mediating antigen-induced T lymphocyte death. J. Immunol. 160:763–69 20. Wang J, Lenardo MJ. 1997. Essential lymphocyte function associated 1 (LFA-1): intercellular adhesion molecule interactions for T cell-mediated B cell apoptosis by Fas/APO1/CD95. J. Exp. Med. 186:1171–76 21. Kischkel FC, Hellbardt S, Behrmann I, Germer M, Pawlita M, et al. 1995. Cytotoxicitydependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor. EMBO J. 14:5579–88 22. Fesik SW. 2000. Insights into programmed cell death through structural biology. Cell 103:273–82 23. Tibbetts MD, Zheng LX, Lenardo MJ. 2003. The death effector domain protein family: regulators of cellular homeostasis. Nat. Immunol. 4:404–9 24. Zhang J, Cado D, Chen A, Kabra NH, Winoto A. 1998. Fas-mediated apoptosis and activation-induced T-cell proliferation are defective in mice lacking FADD/Mort1. Nature 392:296–300 25. Nicholson DW, Thornberry NA. 1997. Caspases: killer proteases. Trends Biochem. Sci. 22:299–306 26. Boatright KM, Salvesen GS. 2003. Mechanisms of caspase activation. Curr. Opin. Cell Biol. 15:725–31 27. Jiang X, Wang X. 2004. Cytochrome C-mediated apoptosis. Annu. Rev. Biochem. 73:87– 106 28. Krueger A, Fas SC, Baumann S, Krammer PH. 2003. The role of CD95 in the regulation of peripheral T-cell apoptosis. Immunol. Rev. 193:58–69 29. Algeciras-Schimnich A, Shen L, Barnhart BC, Murmann AE, Burkhardt JK, et al. 2002. Molecular ordering of the initial signaling events of CD95. Mol. Cell. Biol. 22:207–20 30. Siegel RM, Muppidi JR, Sarker M, Lobito A, Jen M, et al. 2004. SPOTS: signaling protein oligomeric transduction structures are early mediators of death receptor-induced apoptosis at the plasma membrane. J. Cell Biol. 167:735–44 31. Schneider-Brachert W, Tchikov V, Neumeyer J, Jakob M, Winoto-Morbach S, et al. 2004. Compartmentalization of TNF receptor 1 signaling: internalized TNF receptosomes as death signaling vesicles. Immunity 21:415–28 32. Thome M, Tschopp J. 2001. Regulation of lymphocyte proliferation and death by FLIP. Nat. Rev. Immunol. 1:50–58 33. Fernandes-Alnemri T, Armstrong RC, Krebs J, Srinivasula SM, Wang L, et al. 1996. In vitro activation of CPP32 and Mch3 by Mch4, a novel human apoptotic cysteine

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-11

344

Bid`ere

· · Su

Lenardo

ANRV270-IY24-11

34. 35.

36.

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

37.

38. 39. 40.

41. 42.

43.

44.

45.

46.

47.

48.

49.

ARI

23 February 2006

15:22

protease containing two FADD-like domains. Proc. Natl. Acad. Sci. USA 93:7464– 69 Irmler M, Thome M, Hahne M, Schneider P, Hofmann K, et al. 1997. Inhibition of death receptor signals by cellular FLIP. Nature 388:190–95 Rasper DM, Vaillancourt JP, Hadano S, Houtzager VM, Seiden I, et al. 1998. Cell death attenuation by ‘Usurpin,’ a mammalian DED-caspase homologue that precludes caspase8 recruitment and activation by the CD-95 (Fas, APO-1) receptor complex. Cell Death Differ. 5:271–88 Krueger A, Schmitz I, Baumann S, Krammer PH, Kirchhoff S. 2001. Cellular FLICEinhibitory protein splice variants inhibit different steps of caspase-8 activation at the CD95 death-inducing signaling complex. J. Biol. Chem. 276:20633–40 Wang J, Lobito AA, Shen F, Hornung F, Winoto A, Lenardo MJ. 2000. Inhibition of Fas-mediated apoptosis by the B cell antigen receptor through c-FLIP. Eur. J. Immunol. 30:155–63 Rieux-Laucat F, Fischer A, Deist FL. 2003. Cell-death signaling and human disease. Curr. Opin. Immunol. 15:325–31 Sneller MC, Dale JK, Straus SE. 2003. Autoimmune lymphoproliferative syndrome. Curr. Opin. Immunol. 15:417–21 Bleesing JJ, Brown MR, Dale JK, Straus SE, Lenardo MJ, et al. 2001. TcR-α/β+ CD4− CD8− T cells in humans with the autoimmune lymphoproliferative syndrome express a novel CD45 isoform that is analogous to murine B220 and represents a marker of altered O-glycan biosynthesis. Clin. Immunol. 100:314–24 Zuniga-Pﬂucker JC, Lenardo MJ. 1996. Regulation of thymocyte development from immature progenitors. Curr. Opin. Immunol. 8:215–24 Bleesing JJ, Brown MR, Novicio C, Guarraia D, Dale JK, et al. 2002. A composite picture of TcR α/β+ CD4− CD8− T cells (α/β-DNTCs) in humans with autoimmune lymphoproliferative syndrome. Clin. Immunol. 104:21–30 Holzelova E, Vonarbourg C, Stolzenberg MC, Arkwright PD, Selz F, et al. 2004. Autoimmune lymphoproliferative syndrome with somatic Fas mutations. N. Engl. J. Med. 351:1409–18 Lopatin U, Yao X, Williams RK, Bleesing JJ, Dale JK, et al. 2001. Increases in circulating and lymphoid tissue interleukin-10 in autoimmune lymphoproliferative syndrome are associated with disease expression. Blood 97:3161–70 Fuss IJ, Strober W, Dale JK, Fritz S, Pearlstein GR, et al. 1997. Characteristic T helper 2 T cell cytokine abnormalities in autoimmune lymphoproliferative syndrome, a syndrome marked by defective apoptosis and humoral autoimmunity. J. Immunol. 158:1912–18 Sneller MC, Straus SE, Jaffe ES, Jaffe JS, Fleisher TA, et al. 1992. A novel lymphoproliferative/autoimmune syndrome resembling murine lpr/gld disease. J. Clin. Invest. 90:334–41 Teachey DT, Manno CS, Axsom KM, Andrews T, Choi JK, et al. 2004. Unmasking Evans Syndrome: T cell phenotype and apoptotic response reveal autoimmune lymphoproliferative syndrome (ALPS). Blood 105:2443–48 Wong CS, Arkwright PD, Rieux-Laucat F, Cant AJ, Stevens RF, et al. 2004. Childhood linear IgA disease in association with autoimmune lymphoproliferative syndrome. Br. J. Dermatol. 150:578–80 Wu J, Wilson J, He J, Xiang L, Schur PH, Mountz JD. 1996. Fas ligand mutation in a patient with systemic lupus erythematosus and lymphoproliferative disease. J. Clin. Invest. 98:1107–13 www.annualreviews.org • Genetic Disorders and Cell Death

345

ARI

23 February 2006

15:22

50. Straus SE, Jaffe ES, Puck JM, Dale JK, Elkon KB, et al. 2001. The development of lymphomas in families with autoimmune lymphoproliferative syndrome with germline Fas mutations and defective lymphocyte apoptosis. Blood 98:194–200 51. Legembre P, Barnhart BC, Zheng LX, Vijayan S, Straus SE, et al. 2004. Induction of apoptosis and activation of NF-κB by CD95 require different signalling thresholds. EMBO Rep. 5:1084–89 52. Davidson WF, Giese T, Fredrickson TN. 1998. Spontaneous development of plasmacytoid tumors in mice with defective Fas-Fas ligand interactions. J. Exp. Med. 187:1825–38 53. Cohen PL, Eisenberg RA. 1991. Lpr and gld: single gene models of systemic autoimmunity and lymphoproliferative disease. Annu. Rev. Immunol. 9:243–69 54. Kimura M, Matsuzawa A. 1994. Autoimmunity in mice bearing lprcg: a novel mutant gene. Int. Rev. Immunol. 11:193–210 55. Nagata S. 1998. Human autoimmune lymphoproliferative syndrome, a defect in the apoptosis-inducing Fas receptor: a lesson from the mouse model. J. Hum. Genet. 43:2– 8 56. Nose M, Nishihara M, Kamogawa J, Terada M, Nakatsuru S. 2000. Genetic basis of autoimmune disease in MRL/lpr mice: dissection of the complex pathological manifestations and their susceptibility loci. Rev. Immunogenet. 2:154–64 57. Tsuneyama K, Nose M, Nisihara M, Katayanagi K, Harada K, et al. 2001. Spontaneous occurrence of chronic non-suppurative destructive cholangitis and antimitochondrial autoantibodies in MRL/lpr mice: possible animal model for primary biliary cirrhosis. Pathol. Int. 51:418–24 58. Kamogawa J, Terada M, Mizuki S, Nishihara M, Yamamoto H, et al. 2002. Arthritis in MRL/lpr mice is under the control of multiple gene loci with an allelic combination derived from the original inbred strains. Arthritis Rheum. 46:1067–74 59. Miyazaki T, Ono M, Qu WM, Zhang MC, Mori S, et al. 2005. Implication of allelic polymorphism of osteopontin in the development of lupus nephritis in MRL/lpr mice. Eur. J. Immunol. 35:1510–20 60. Chiocchetti A, Indelicato M, Bensi T, Mesturini R, Giordano M, et al. 2004. High levels of osteopontin associated with polymorphisms in its gene are a risk factor for development of autoimmunity/lymphoproliferation. Blood 103:1376–82 61. Hang L, Theoﬁlopoulos AN, Balderas RS, Francis SJ, Dixon FJ. 1984. The effect of thymectomy on lupus-prone mice. J. Immunol. 132:1809–13 62. Wofsy D, Ledbetter JA, Roubinian JR, Seaman WE, Talal N. 1982. Thymic inﬂuences on autoimmunity in MRL-lpr mice. Scand. J. Immunol. 16:51–58 63. van der Werff Ten Bosch J, Schotte P, Ferster A, Azzi N, Boehler T, et al. 2002. Reversion of autoimmune lymphoproliferative syndrome with an antimalarial drug: preliminary results of a clinical cohort study and molecular observations. Br. J. Dermatol. 117:176–88 64. Benkerrou M, Le Deist F, de Villartay JP, Caillat-Zucman S, Rieux-Laucat F, et al. 1997. Correction of Fas (CD95) deﬁciency by haploidentical bone marrow transplantation. Eur. J. Immunol. 27:2043–47 65. Hundt M, Posovszky C, Schmidt RE. 2002. A new disorder of lymphocyte apoptosis: combination of autoimmunity, infectious lymphadenopathy, double negative T cells, and impaired activation-induced cell death. Immunobiology 206:514–18 66. Ramenghi U, Bonissoni S, Migliaretti G, DeFranco S, Bottarel F, et al. 2000. Deﬁciency of the Fas apoptosis pathway without Fas gene mutations is a familial trait predisposing to development of autoimmune diseases and cancer. Blood 95:3176–82

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-11

346

Bid`ere

· · Su

Lenardo

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-11

ARI

23 February 2006

15:22

67. Chan FK-M, Siegel RM, Lenardo MJ. 2000. Signaling by the TNF receptor superfamily and T cell homeostasis. Immunity 13:419–22 68. Siegel RM, Chan FK-M, Chun HJ, Lenardo MJ. 2000. The multifaceted role of Fas signaling in immune cell homeostasis and autoimmunity. Nat. Immunol. 1:469–74 69. Roesler J, Izquierdo JM, Ryser M, Rosen-Wolff A, Gahr M, et al. 2005. Haploinsufﬁciency rather than the effect of an excessive production of soluble CD95 (CD95 Delta TM) is the basis for ALPS Ia in a family with duplicated 3 splice site AG in CD95 intron 5 on one allele. Blood 106:1652–59 70. Rieux-Laucat F, Le Deist F, Fischer A. 2003. Autoimmune lymphoproliferative syndromes: genetic defects of apoptosis pathways. Cell Death Differ. 10:124–33 71. Gronbaek K, Dalby T, Zeuthen J, Ralfkiaer E, Guidberg P. 2000. The V410I (G1228A) variant of the caspase-10 gene is a common polymorphism of the Danish population. Blood 95:2184–85 72. Fisher GH, Rosenberg FJ, Straus SE, Dale JK, Middleton LA, et al. 1995. Dominant interfering Fas gene mutations impair apoptosis in a human autoimmune lymphoproliferative syndrome. Cell 81:935–46 73. Herskowitz I. 1987. Functional inactivation of genes by dominant negative mutations. Nature 329:219–22 74. Martin DA, Zheng L, Siegel RM, Huang B, Fisher GH, et al. 1999. Defective CD95/APO-1/Fas signal complex formation in the human autoimmune lymphoproliferative syndrome, type Ia. Proc. Natl. Acad. Sci. USA 96:4552–57 75. Siegel RM, Frederiksen JK, Zacharias DA, Chan FK-M, Johnson M, et al. 2000. Fas preassociation required for apoptosis signaling and dominant inhibition by pathogenic mutations. Science 288:2354–57 76. Chan FK-M, Chun HJ, Zheng LX, Siegel RM, Bui KL, Lenardo MJ. 2000. A domain in TNF receptors that mediates ligand-independent receptor assembly and signaling. Science 288:2351–54 77. Remy I, Wilson IA, Michnick SW. 1999. Erythropoietin receptor activation by a ligandinduced conformation change. Science 283:990–93 78. Holler N, Tardivel A, Kovacsovics-Bankowski M, Hertig S, Gaide O, et al. 2003. Two adjacent trimeric Fas ligands are required for Fas signaling and formation of a deathinducing signaling complex. Mol. Cell. Biol. 23:1428–40 79. Kischkel FC, Lawrence DA, Tinel A, LeBlanc H, Virmani A, et al. 2001. Death receptor recruitment of endogenous caspase-10 and apoptosis initiation in the absence of caspase8. J. Biol. Chem. 276:46639–46 80. Wang J, Chun HJ, Wong W, Spencer DM, Lenardo MJ. 2001. Caspase-10 is an initiator caspase in death receptor signaling. Proc. Natl. Acad. Sci. USA 98:13884–88 81. Siegel RM, Chan FK-M, Zacharias DA, Swofford R, Holmes KL, et al. 2000. Measurement of molecular interactions in living cells by ﬂuorescence resonance energy transfer between variants of the green ﬂuorescent protein. Science STKE 2000:PL1 82. Sprick MR, Rieser E, Stahl H, Grosse-Wilde A, Weigand MA, et al. 2002. Caspase-10 is recruited to and activated at the native TRAIL and CD95 death-inducing signalling complexes in a FADD-dependent manner but can not functionally substitute caspase-8. EMBO J. 21:4520–30 83. Park WS, Lee JH, Shin MS, Park JY, Kim HS, et al. 2002. Inactivating mutations of the caspase-10 gene in gastric cancer. Oncogene 21:2919–25 84. Shin MS, Kim HS, Kang CS, Park WS, Kim SY, et al. 2002. Inactivating mutations of CASP10 gene in non-Hodgkin lymphomas. Blood 99:4094–99 www.annualreviews.org • Genetic Disorders and Cell Death

347

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85. Lens SM, Kataoka T, Fortner KA, Tinel A, Ferrero I, et al. 2002. The caspase 8 inhibitor c-FLIP(L) modulates T-cell receptor-induced proliferation but not activation-induced cell death of lymphocytes. Mol. Cell. Biol. 22:5419–33 86. Stanger BZ, Leder P, Lee TH, Kim E, Seed B. 1995. RIP: a novel protein containing a death domain that interacts with Fas/APO-1 (CD95) in yeast and causes cell death. Cell 81:513–23 87. Rieux-Laucat F, Le Deist F, Hivroz C, Roberts IA, Debatin KM, et al. 1995. Mutations in Fas associated with human lymphoproliferative syndrome and autoimmunity. Science 268:1347–49 88. Jackson CE, Fischer RE, Hsu AP, Anderson SM, Choi Y, et al. 1999. Autoimmune lymphoproliferative syndrome with defective Fas: genotype inﬂuences penetrance. Am. J. Hum. Genet. 64:1002–14 89. Choi Y, Ramnath VR, Eaton AS, Chen A, Simon-Stoos KL, et al. 1999. Expression in transgenic mice of dominant interfering Fas mutations: a model for human autoimmune lymphoproliferative syndrome. Clin. Immunol. 93:34–45 90. Klein J, Sato A. 2000. The HLA system. Second of two parts. N. Engl. J. Med. 343:782–86 91. Purrmann J, Bertrams J, Knapp M, Cleveland S, Hengels KJ, et al. 1990. Gene and haplotype frequencies of HLA antigens in 269 patients with Crohn’s disease. Scand. J. Gastroenterol. 25:981–85 92. Clementi R, Dagna L, Dianzani U, Dupre L, Dianzani I, et al. 2004. Inherited perforin and Fas mutations in a patient with autoimmune lymphoproliferative syndrome and lymphoma. N. Engl. J. Med. 351:1419–24 93. Rieux-Laucat F, Le Deist F, de Saint Basile G. 2005. Autoimmune lymphoproliferative syndrome and perforin. N. Engl. J. Med. 352:306 94. Ueda H, Howson JM, Esposito L, Heward J, Snook H, et al. 2003. Association of the T-cell regulatory gene CTLA4 with susceptibility to autoimmune disease. Nature 423:506–11 95. Hull KM, Drewe E, Aksentijevich I, Singh HK, Wong K, et al. 2002. The TNF receptorassociated periodic syndrome (TRAPS): emerging concepts of an autoinﬂammatory disorder. Medicine 81:349–68 96. Touitou I. 2001. The spectrum of Familial Mediterranean Fever (FMF) mutations. Eur. J. Hum. Genet. 9:473–83 97. Varfolomeev EE, Schuchmann M, Luria V, Chiannilkulchai N, Beckmann JS, et al. 1998. Targeted disruption of the mouse caspase 8 gene ablates cell death induction by the TNF receptors, Fas/Apo1, and DR3 and is lethal prenatally. Immunity 9:267–76 98. Kennedy NJ, Kataoka T, Tschopp J, Budd RC. 1999. Caspase activation is required for T cell proliferation. J. Exp. Med. 190:1891–96 99. Salmena L, Lemmers B, Hakem A, Matysiak-Zablocki E, Murakami K, et al. 2003. Essential role for caspase 8 in T-cell homeostasis and T-cell-mediated immunity. Genes Dev. 17:883–95 100. Li Q, Verma IM. 2002. NF-κB regulation in the immune system. Nat. Rev. Immunol. 2:725–34 101. Karin M, Delhase M. 2000. The IκB kinase (IKK) and NF-κB: key elements of proinﬂammatory signalling. Semin. Immunol. 12:85–98 102. Thome M. 2004. CARMA1, BCL-10 and MALT1 in lymphocyte development and activation. Nat. Rev. Immunol. 4:348–59 103. Lee KY, D’Acquisto F, Hayden MS, Shim JH, Ghosh S. 2005. PDK1 nucleates T cell receptor-induced signaling complex for NF-κB activation. Science 308:114–18

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-11

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· · Su

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ANRV270-IY24-11

ARI

23 February 2006

15:22

104. Newton K, Harris AW, Bath ML, Smith KG, Strasser A. 1998. A dominant interfering mutant of FADD/MORT1 enhances deletion of autoreactive thymocytes and inhibits proliferation of mature T lymphocytes. EMBO J. 17:706–18 105. Sadlack B, Merz H, Schorle H, Schimpl A, Feller AC, et al. 1993. Ulcerative colitis-like disease in mice with a disrupted interleukin-2 gene. Cell 75:253–61 106. Suzuki H, Kundig TM, Furlonger C, Wakeham A, Timms E, et al. 1995. Deregulated T cell activation and autoimmunity in mice lacking interleukin-2 receptor β. Science 268:1472–76 107. Willerford DM, Chen J, Ferry JA, Davidson L, Ma A, et al. 1995. Interleukin-2 receptor alpha chain regulates the size and content of the peripheral lymphoid compartment. Immunity 3:521–30 108. Sharfe N, Dadi HK, Shahar M, Roifman CM. 1997. Human immune disorder arising from mutation of the alpha chain of the interleukin-2 receptor. Proc. Natl. Acad. Sci. USA 94:3168–71 109. Lenardo MJ. 1991. Interleukin-2 programs mouse αβ T lymphocytes for apoptosis. Nature 353:858–61 110. Khoruts A, Mondino A, Pape KA, Reiner SL, Jenkins MK. 1998. A natural immunological adjuvant enhances T cell clonal expansion through a CD28-dependent, interleukin (IL)2-independent mechanism. J. Exp. Med. 187:225–36 111. Deleted in proof 112. Kneitz B, Herrmann T, Yonehara S, Schimpl A. 1995. Normal clonal expansion but impaired Fas-mediated cell death and anergy induction in interleukin-2-deﬁcient mice. Eur. J. Immunol. 25:2572–77 113. Van Parijs L, Biuckians A, Ibragimov A, Alt FW, Willerford DM, et al. 1997. Functional responses and apoptosis of CD25 (IL-2R α)-deﬁcient T cells expressing a transgenic antigen receptor. J. Immunol. 158:3738–45 114. Suzuki H, Hayakawa A, Bouchard D, Nakashima I, Mak TW, et al. 1997. Normal thymic selection, superantigen-induced deletion and Fas-mediated apoptosis of T cells in IL-2 receptor beta chain-deﬁcient mice. Int. Immunol. 9:1367–74 115. Kovanen PE, Young L, Al-Shami A, Rovella V, Pise-Masison CA, et al. 2005. Global analysis of IL-2 target genes: identiﬁcation of chromosomal clusters of expressed genes. Int. Immunol. 17:1009–21 116. Rifa’i M, Kawamoto Y, Nakashima I, Suzuki H. 2004. Essential roles of CD8+CD122+ regulatory T cells in the maintenance of T cell homeostasis. J. Exp. Med. 200:1123–34 117. Suzuki H, Zhou YW, Kato M, Mak TW, Nakashima I. 1999. Normal regulatory α/β T cells effectively eliminate abnormally activated T cells lacking the interleukin 2 receptor β in vivo. J. Exp. Med. 190:1561–72 118. Malek TR, Yu A, Vincek V, Scibelli P, Kong L. 2002. CD4 regulatory T cells prevent lethal autoimmunity in IL-2Rβ-deﬁcient mice. Implications for the nonredundant function of IL-2. Immunity 17:167–78 119. Wolf M, Schimpl A, Hunig T. 2001. Control of T cell hyperactivation in IL-2-deﬁcient mice by CD4+ CD25− and CD4+ CD25+ T cells: evidence for two distinct regulatory mechanisms. Eur. J. Immunol. 31:1637–45 120. Kung JT, Beller D, Ju ST. 1998. Lymphokine regulation of activation-induced apoptosis in T cells of IL-2 and IL-2R β knockout mice. Cell. Immunol. 185:158–63 121. Cao X, Shores EW, Hu-Li J, Anver MR, Kelsall BL, et al. 1995. Defective lymphoid development in mice lacking expression of the common cytokine receptor gamma chain. Immunity 2:223–38 www.annualreviews.org • Genetic Disorders and Cell Death

349

ARI

23 February 2006

15:22

122. DiSanto JP, Rieux-Laucat F, Dautry-Varsat A, Fischer A, de Saint Basile G, et al. 1994. Defective human interleukin 2 receptor gamma chain in an atypical X chromosomelinked severe combined immunodeﬁciency with peripheral T cells. Proc. Natl. Acad. Sci. USA 91:9466–70 123. Nakajima H, Leonard WJ. 1997. Impaired peripheral deletion of activated T cells in mice lacking the common cytokine receptor gamma-chain: defective Fas ligand expression in gamma-chain-deﬁcient mice. J. Immunol. 159:4737–44 124. Borlado LR, Redondo C, Alvarez B, Jimenez C, Criado LM, et al. 2000. Increased phosphoinositide 3-kinase activity induces a lymphoproliferative disorder and contributes to tumor generation in vivo. FASEB J. 14:895–903 125. Jones RG, Parsons M, Bonnard M, Chan VS, Yeh WC, et al. 2000. Protein kinase B regulates T lymphocyte survival, nuclear factor κB activation, and Bcl-X(L) levels in vivo. J. Exp. Med. 191:1721–34 126. Parsons MJ, Jones RG, Tsao MS, Odermatt B, Ohashi PS, et al. 2001. Expression of active protein kinase B in T cells perturbs both T and B cell homeostasis and promotes inﬂammation. J. Immunol. 167:42–88 127. Rathmell JC, Elstrom RL, Cinalli RM, Thompson CB. 2003. Activated Akt promotes increased resting T cell size, CD28-independent T cell growth, and development of autoimmunity and lymphoma. Eur. J. Immunol. 33:2223–32 128. Di Cristofano A, Kotsi P, Peng YF, Cordon-Cardo C, Elkon KB, Pandolﬁ PP. 1999. Impaired Fas response and autoimmunity in Pten+ -mice. Science 285:2122–25 129. Podsypanina K, Ellenson LH, Nemes A, Gu J, Tamura M, et al. 1999. Mutation of Pten/Mmac1 in mice causes neoplasia in multiple organ systems. Proc. Natl. Acad. Sci. USA 96:1563–68 130. Suzuki A, de la Pompa JL, Stambolic V, Elia AJ, Sasaki T, et al. 1998. High cancer susceptibility and embryonic lethality associated with mutation of the PTEN tumor suppressor gene in mice. Curr. Biol. 8:1169–78 131. Suzuki A, Kaisho T, Ohishi M, Tsukio-Yamaguchi M, Tsubata T, et al. 2003. Critical roles of Pten in B cell homeostasis and immunoglobulin class switch recombination. J. Exp. Med. 197:657–67 132. Suzuki A, Yamaguchi MT, Ohteki T, Sasaki T, Kaisho T, et al. 2001. T cell-speciﬁc loss of Pten leads to defects in central and peripheral tolerance. Immunity 14:523–34 133. Bouillet P, Metcalf D, Huang DC, Tarlinton DM, Kay TW, et al. 1999. Proapoptotic Bcl-2 relative Bim required for certain apoptotic responses, leukocyte homeostasis, and to preclude autoimmunity. Science 286:1735–38 134. Knudson CM, Korsmeyer SJ. 1997. Bcl-2 and Bax function independently to regulate cell death. Nat. Genet. 16:358–63 135. Knudson CM, Tung KS, Tourtellotte WG, Brown GA, Korsmeyer SJ. 1995. Bax-deﬁcient mice with lymphoid hyperplasia and male germ cell death. Science 270:96–99 136. Lindsten T, Ross AJ, King A, Zong WX, Rathmell JC, et al. 2000. The combined functions of proapoptotic Bcl-2 family members bak and bax are essential for normal development of multiple tissues. Mol. Cell 6:1389–99 137. Strasser A, Harris AW, Cory S. 1993. E mu-bcl-2 transgene facilitates spontaneous transformation of early pre-B and immunoglobulin-secreting cells but not T cells. Oncogene 8:1–9 138. Strasser A, Whittingham S, Vaux DL, Bath ML, Adams JM, et al. 1991. Enforced BCL2 expression in B-lymphoid cells prolongs antibody responses and elicits autoimmune disease. Proc. Natl. Acad. Sci. USA 88:8661–65

Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

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· · Su

Lenardo

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ANRV270-IY24-11

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23 February 2006

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139. Hao Z, Duncan GS, Chang CC, Elia A, Fang M, et al. 2005. Speciﬁc ablation of the apoptotic functions of cytochrome C reveals a differential requirement for cytochrome C and Apaf-1 in apoptosis. Cell 121:579–91 140. Woo M, Hakem R, Furlonger C, Hakem A, Duncan GS, et al. 2003. Caspase-3 regulates cell cycle in B cells: a consequence of substrate speciﬁcity. Nat. Immunol. 4:1016–22 141. Woo M, Hakem R, Soengas MS, Duncan GS, Shahinian A, et al. 1998. Essential contribution of caspase 3/CPP32 to apoptosis and its associated nuclear changes. Genes Dev. 12:806–19 142. Dudley E, Hornung F, Zheng L, Scherer D, Ballard D, Lenardo M. 1999. NF-κB regulates Fas/APO-1/CD95- and TCR-mediated apoptosis of T lymphocytes. Eur. J. Immunol. 29:878–86 143. Lafuente MJ, Martin P, Garcia-Cao I, Diaz-Meco MT, Serrano M, Moscat J. 2003. Regulation of mature T lymphocyte proliferation and differentiation by Par-4. EMBO J. 22:4689–98 144. Gillette-Ferguson I, Sidman CL. 1994. A speciﬁc intercellular pathway of apoptotic cell death is defective in the mature peripheral T cells of autoimmune lpr and gld mice. Eur. J. Immunol. 24:1181–85 145. Matsuzawa A, Moriyama T, Kaneko T, Tanaka M, Kimura M, et al. 1990. A new allele of the lpr locus, lprcg, that complements the gld gene in induction of lymphadenopathy in the mouse. J. Exp. Med. 171:519–31 146. Watanabe-Fukunaga R, Brannan CI, Copeland NG, Jenkins NA, Nagata S, et al. 1992. Lymphoproliferation disorder in mice explained by defects in Fas antigen that mediates apoptosis. Nature 356:314–17 147. Roths JB, Murphy ED, Eicher EM. 1984. A new mutation, gld, that produces lymphoproliferation and autoimmunity in C3H/HeJ mice. J. Exp. Med. 159:1–20 148. Takahashi T, Tanaka M, Brannan CI, Jenkins NA, Copeland NG, et al. 1994. Generalized lymphoproliferative disease in mice, caused by a point mutation in the Fas ligand. Cell 76:969–76 149. Deleted in proof 150. Enders A, Bouillet P, Puthalakath H, Xu Y, Tarlinton DM, Strasser A. 2003. Loss of the pro-apoptotic BH3-only Bcl-2 family member Bim inhibits BCR stimulation-induced apoptosis and deletion of autoreactive B cells. J. Exp. Med. 198:1119–26 151. Hildeman DA, Zhu Y, Mitchell TC, Bouillet P, Strasser A, et al. 2002. Activated T cell death in vivo mediated by proapoptotic bcl-2 family member bim. Immunity 16:759– 67 152. Sun H, Lu B, Li RQ, Flavell RA, Taneja R. 2001. Defective T cell activation and autoimmune disorder in Stra13-deﬁcient mice. Nat. Immunol. 2:1040–47 153. Drappa J, Kamen LA, Chan E, Georgiev M, Ashany D, et al. 2003. Impaired T cell death and lupus-like autoimmunity in T cell-speciﬁc adapter protein-deﬁcient mice. J. Exp. Med. 198:809–21 154. Fanzo JC, Hu CM, Jang SY, Pernis AB. 2003. Regulation of lymphocyte apoptosis by interferon regulatory factor 4 (IRF-4). J. Exp. Med. 197:303–14 155. Field SJ, Tsai FY, Kuo F, Zubiaga AM, Kaelin WG Jr, et al. 1996. E2F-1 functions in mice to promote apoptosis and suppress proliferation. Cell 85:549–61 156. Lissy NA, Davis PK, Irwin M, Kaelin WG, Dowdy SF. 2000. A common E2F-1 and p73 pathway mediates cell death induced by TCR activation. Nature 407:642–45 157. Yamasaki L, Jacks T, Bronson R, Goillot E, Harlow E, Dyson NJ. 1996. Tumor induction and tissue atrophy in mice lacking E2F-1. Cell 85:537–48 www.annualreviews.org • Genetic Disorders and Cell Death

351

ANRV270-IY24-11

ARI

23 February 2006

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Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

158. Sommers CL, Park CS, Lee J, Feng C, Fuller CL, et al. 2002. A LAT mutation that inhibits T cell development yet induces lymphoproliferation. Science 296:2040–43 159. Hodge MR, Ranger AM, Charles de la Brousse F, Hoey T, Grusby MJ, et al. 1996. Hyperproliferation and dysregulation of IL-4 expression in NF-ATp-deﬁcient mice. Immunity 4:397–405 160. Peng SL, Gerth AJ, Ranger AM, Glimcher LH. 2001. NFATc1 and NFATc2 together control both T and B cell activation and differentiation. Immunity 14:13–20 161. Ranger AM, Oukka M, Rengarajan J, Glimcher LH. 1998. Inhibitory function of two NFAT family members in lymphoid homeostasis and Th2 development. Immunity 9:627– 35 162. Qian YC, Qin JZ, Cui G, Naramura M, Snow EC, et al. 2004. Act1, a negative regulator in CD40- and BAFF-mediated B cell survival. Immunity 21:575–87

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Contents

Annual Review of Immunology Volume 24, 2006

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Frontispiece Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p x The Tortuous Journey of a Biochemist to Immunoland and What He Found There Jack L. Strominger p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1 Osteoimmunology: Interplay Between the Immune System and Bone Metabolism Matthew C. Walsh, Nacksung Kim, Yuho Kadono, Jaerang Rho, Soo Young Lee, Joseph Lorenzo, and Yongwon Choi p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p33 A Molecular Perspective of CTLA-4 Function Wendy A. Teft, Mark G. Kirchhof, and Joaquín Madrenas p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p65 Transforming Growth Factor-β Regulation of Immune Responses Ming O. Li, Yisong Y. Wan, Shomyseh Sanjabi, Anna-Karin L. Robertson, and Richard A. Flavell p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p99 The Eosinophil Marc E. Rothenberg and Simon P. Hogan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 147 Human T Cell Responses Against Melanoma Thierry Boon, Pierre G. Coulie, Benoît J. Van den Eynde, and Pierre van der Bruggen p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 175 FOXP3: Of Mice and Men Steven F. Ziegler p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 209 HIV Vaccines Andrew J. McMichael p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 227 Natural Killer Cell Developmental Pathways: A Question of Balance James P. Di Santo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 257 Development of Human Lymphoid Cells Bianca Blom and Hergen Spits p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 287 Genetic Disorders of Programmed Cell Death in the Immune System Nicolas Bidère, Helen C. Su, and Michael J. Lenardo p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 321

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 353 Multiplexed Protein Array Platforms for Analysis of Autoimmune Diseases Imelda Balboni, Steven M. Chan, Michael Kattah, Jessica D. Tenenbaum, Atul J. Butte, and Paul J. Utz p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 391 How TCRs Bind MHCs, Peptides, and Coreceptors Markus G. Rudolph, Robyn L. Stanﬁeld, and Ian A. Wilson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 419 Annu. Rev. Immunol. 2006.24:321-352. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

B Cell Immunobiology in Disease: Evolving Concepts from the Clinic Flavius Martin and Andrew C. Chan p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 467 The Evolution of Adaptive Immunity Zeev Pancer and Max D. Cooper p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 497 Cooperation Between CD4+ and CD8+ T Cells: When, Where, and How Flora Castellino and Ronald N. Germain p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 519 Mechanism and Control of V(D)J Recombination at the Immunoglobulin Heavy Chain Locus David Jung, Cosmas Giallourakis, Raul Mostoslavsky, and Frederick W. Alt p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 541 A Central Role for Central Tolerance Bruno Kyewski and Ludger Klein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 571 Regulation of Th2 Differentiation and Il4 Locus Accessibility K. Mark Ansel, Ivana Djuretic, Bogdan Tanasa, and Anjana Rao p p p p p p p p p p p p p p p p p p p p p p p 607 Diverse Functions of IL-2, IL-15, and IL-7 in Lymphoid Homeostasis Averil Ma, Rima Koka, and Patrick Burkett p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 657 Intestinal and Pulmonary Mucosal T Cells: Local Heroes Fight to Maintain the Status Quo Leo Lefrançois and Lynn Puddington p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 681 Determinants of Lymphoid-Myeloid Lineage Diversiﬁcation Catherine V. Laiosa, Matthias Stadtfeld, and Thomas Graf p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 705 GP120: Target for Neutralizing HIV-1 Antibodies Ralph Pantophlet and Dennis R. Burton p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 739 Compartmentalized Ras/MAPK Signaling Adam Mor and Mark R. Philips p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 771

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Genetic Analysis of Host Resistance: Toll-Like Receptor Signaling and Immunity at Large Bruce Beutler, Zhengfan Jiang, Philippe Georgel, Karine Crozat, Ben Croker, Sophie Rutschmann, Xin Du, and Kasper Hoebe Department of Immunology, Scripps Research Institute, La Jolla, California 92037; email: [email protected]

Annu. Rev. Immunol. 2006. 24:353–89 The Annual Review of Immunology is online at immunol.annualreviews.org This article’s doi: 10.1146/annurev.immunol.24.021605.090552 c 2006 by Copyright Annual Reviews. All rights reserved 0732-0582/06/0423-0353$20.00

Key Words mutagenesis, Mendelian genetics, infection, innate immunity

Abstract Classical genetic methods, driven by phenotype rather than hypotheses, generally permit the identiﬁcation of all proteins that serve nonredundant functions in a deﬁned biological process. Long before this goal is achieved, and sometimes at the very outset, genetics may cut to the heart of a biological puzzle. So it was in the ﬁeld of mammalian innate immunity. The positional cloning of a spontaneous mutation that caused lipopolysaccharide resistance and susceptibility to Gram-negative infection led directly to the understanding that Toll-like receptors (TLRs) are essential sensors of microbial infection. Other mutations, induced by the random germ line mutagen ENU (N-ethyl-N-nitrosourea), have disclosed key molecules in the TLR signaling pathways and helped us to construct a reasonably sophisticated portrait of the afferent innate immune response. A still broader genetic screen—one that detects all mutations that compromise survival during infection—is permitting fresh insight into the number and types of proteins that mammals use to defend themselves against microbes.

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INTRODUCTION

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Forward genetics: the genetic approach that begins with phenotype of unknown origin and ends with the identiﬁcation of culpable mutation(s)

Phenomena and Phenotypes: The Geneticist’s View of Resistance to Infectious Disease Few immunologists are content to forsake hypotheses and follow a classical genetic strategy. But perhaps more of them should. The forward genetic approach to biological problems is an unbiased one. It does not begin with broad speculation about how a biological system might operate, or even with more restricted hypotheses, such as guesses about the function of a particular protein or how cells interact. On the contrary, genetic exploration begins with no preconceptions at all. Hypotheses are the outcome of genetic investigation rather than the starting point (Figure 1). In genetics, as in biology at large, phenomena come ﬁrst. They are the ultimate source of all curiosity and all inquiry. Some phenom-

Figure 1 Summary of the classical (“forward”) genetic approach. Hypotheses (and experiments to test them) are the result of genetic inquiry rather than the starting point. 354

Beutler et al.

ena (the shapes of leaves, the presence of spots on the wings of moths, and the fact that dogs beget dogs whereas humans beget humans) are either so subtle or so commonplace that most people hardly think about them at all, although their importance may belie their banality. Other phenomena (aging, consciousness, and cancer) are uppermost in the popular mind and have impelled investigation on a broad scale, using all the tools technology can offer. However prosaic or glamorous the phenomena might seem, their analysis by classical genetic methods entails the identiﬁcation of phenotypes: alternative forms of phenomena. Where such phenotypes do not exist, they must be created. Once a monogenic phenotype is at hand, existing technology invariably permits identiﬁcation of the DNA sequence difference that is behind it. Ultimately, all proteins with nonredundant functions in a particular biological phenomenon may be identiﬁed in this manner. Even if it falls short of this idealized goal, classical genetic analysis creates a substrate upon which other methods may operate. The central phenomenon at issue in immunology is resistance to infection: the fact that we do not passively succumb to microbes when inoculated with them. It is a phenomenon that declares itself every day, and it would certainly fall into the “commonplace” category if not for the impressive consequences of its failure. Resistance to infection usually goes unnoticed, but it is essential to the survival of individuals and species. Genetic reasoning immediately tells us that the preponderance of resistance to infection is inherited rather than acquired, despite the quintessentially environmental status of microbes themselves. To argue in the most general way, different mammalian species show great variation in susceptibility to speciﬁc infectious agents. In fact, it is safe to say that there are no universal pathogens, i.e., microbes to which all multicellular organisms (or even all vertebrates) are susceptible. Never yet has a human died of mouse

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cytomegalovirus (MCMV) infection, at least as far as we know. Never yet has a mouse died of human immunodeﬁciency virus (HIV) infection. Admittedly, a vast number of mutational differences have set humans and mice forever apart. But a discrete and ultimately deﬁnable collection of mutations must specify ability to resist infection by these organisms and all others. Although far fewer mutational events distinguish individuals belonging to a single species, formal measurement of heritability reveals that here, too, susceptibility to infection is largely determined by genetic makeup. Premature death from infection in a biological parent confers a high probability of premature death in a child separated from the parent by adoption (1). Moreover, variation in susceptibility to a given infectious agent is frequently demonstrable among individuals of a single species and usually has a genetic basis. Among mice, polymorphisms at speciﬁc loci inﬂuence susceptibility to MCMV (discussed further below) (2–5), and among humans, polymorphisms at speciﬁc loci inﬂuence susceptibility to HIV (6, 7). What types of immunity are there, and which type confers the heritability of resistance to infection? The term “adaptive” is used to describe a particular type of immunity, represented only in vertebrates as far as we know and markedly inﬂuenced by the environment, i.e., by previous exposure to an infectious agent or antigen, which impels the expansion of lymphoid clones that are directed toward speciﬁc recognition of the same target. The term “innate” is used to describe all other forms of immunity. There is a tendency to regard innate immunity as heritable and adaptive immunity, with its remarkable plasticity, as acquired. But, of course, the fact that adaptive immunity can be eliminated by a well-placed mutation reveals that it, too, is inherited. To the geneticist, all heritable effects on immunity are of interest, and the distinction between innate and adaptive systems is of secondary importance, although often easy to make.

Adaptive immunity evolved at least twice (8). The exact selective pressures that drove the development of our own adaptive immune system are unknown and can only be imagined. Perhaps an anticipatory immune system offered species a means of coping with frequent and devastating plagues caused by microbes that escaped containment by the relatively static innate immune response. Whatever the reason, it may be assumed that the adaptive immune system arose in the context of an advanced and highly functional innate immune system, and it remains largely dependent on the innate system today. Subsequent to the rise of adaptive immunity, certain innate immune functions must have become redundant and as a consequence were lost; hence, innate immunity alone offers suboptimal protection to present-day vertebrates. The innate system has come to require certain molecules of adaptive immune origin (for example, IFNγ) to function properly. For these reasons, many of the mutations that affect innate immune function also affect adaptive immune function and vice versa.

MCMV: mouse cytomegalovirus Resistome: the set of all genes encoding proteins with nonredundant functions in host resistance

The Concept of the Resistome The resistome is deﬁned as the set of genes encoding proteins with nonredundant function in resistance to infection (9). It is possible to speak of a universal resistome, as well as of the resistome for speciﬁc organisms. Many components of the resistome are entirely conditional. If infectious organisms are not present, there is no need for these genes to exist. In effect, they are fully dedicated to immune function. Other components of the resistome fulﬁll separate and essential biological functions, having recently been appropriated by evolution to create resistance, or conversely, to serve a function unrelated to resistance. In some instances, it is difﬁcult to decide on the primary function of a protein that is known to participate in resistance. Toll, a transmembrane receptor with a dual function in host resistance (10) and in embryonic development www.annualreviews.org • Genetic Analysis of Host Resistance

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in Drosophila (11), probably was inherited as a mediator of immunity and co-opted to serve a developmental function, based on the use of the TIR (Toll/IL-1R/Resistance) domain for immunity in most species (12). But the premise is debatable, and some believe that the primordial function of Toll was developmental. On occasion, a protein that clearly evolved to serve a function that has nothing to do with immunity may offer strong resistance to infection in one allelic isoform, but it may fail to do so in another form (e.g., sickle hemoglobin, which confers resistance to Plasmodium falciparum, versus hemoglobin A, which does not). Examples of this sort may be viewed as evolutionary works in progress, and sufﬁcient selective pressure might drive the mutant isoform to homozygosity, whereupon, like Toll, the hemoglobin protein would necessarily be viewed as bifunctional, with a role in both oxygen transport and in immunity. Some resistome components (certainly most of them) confer resistance to a broad range of pathogens. In this sense, the resistome is degenerate. Where the innate immune system is concerned, low speciﬁcity is the general rule, to allow for effective resistance to many microbes within the constraints imposed by limitations in genomic size. Some components of the resistome, however, are highly speciﬁc in their effects, and although they serve to recognize or overcome a very restricted repertoire of microbes, they may be essential to survival of the species. Irrespective of functional category, a concerted and determined genetic approach will ultimately reveal every protein that plays a nonredundant role in resistance and, in principle, will even identify those with latent resistance functions, given that such functions do not yet exist. Such a genetic approach will do so even if such functions of the proteins in question are beyond the imagination, are without precedent, and are unpredictable by any exercise of logic. Therein lies the appeal of the genetic approach and the source of its power.

TIR: Toll/IL1R/Resistance

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TLR: Toll-like receptor

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The classical genetic approach is beholden to keen observation and is empowered by experimental designs that reveal phenotypes inapparent in the absence of special conditions. But the classical genetic approach does not depend on guesswork and may ultimately succeed where hypotheses fail. The classical genetic approach led to the identiﬁcation of the Toll-like receptors (TLRs) as the key proteins that allow us to recognize infection and, in large part, to the decipherment of signaling pathways that are activated by the TLRs. Genetic reasoning has also pointed to the existence of alternative pathways leading to activation of both the innate and adaptive immune systems.

THE PHENOMENON: HOST RESISTANCE Cell-Autonomous Immunity and the Evolution of Specialized Systems for Host Resistance Resistance to microbes must be measured with reference to a control rather than in absolute terms because deﬁning a state in which no resistance exists is presently impossible. For example, in the absence of the type I IFN receptor, mice are markedly compromised in their ability to resist MCMV infection (13). If the type II IFN receptor is also eliminated, susceptibility is still greater (13). Given that IFN-independent resistance mechanisms also may exist, susceptibility might be further enhanced by additional mutations. For the present, resistance must be assayed by measuring mean lethal inoculum, mean cytolytic inoculum, or the proliferation of the agent that is tested. Even in the absence of a functioning immune system, cultured cells offer resistance to the growth of microbes. For example, many cells, including ﬁbroblasts, are capable of mounting a type I IFN response. Cellautonomous immunity presumably operates in vivo as well as in vitro. However, advanced multicellular organisms have evolved

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additional defensive strategies that make use of specialized cells designed to sense microbes, alert the host, and destroy microbes. The innate immune system depends heavily on macrophages, several classes of dendritic cells (DCs), natural killer (NK) cells, NKT cells, and granulocytes for host defense. Interestingly, although lymphoid cells provide the cellular basis for adaptive immunity, they have also been used for purely innate immune functions. The responses of macrophages and myeloid DCs have been studied in greatest detail; NK cells have also received considerable attention.

Sensing, Response, and Self-Tolerance: The Strategy of the Innate Immune System Immune responses are potentially injurious to the host and are also energy intensive. Although antimicrobial peptides are constitutively produced in some compartments, most responses are induced by infection. This requires that a system for recognition of microbes must exist, and in almost all biological systems recognition implies the presence of speciﬁc receptors. These receptors and the signals they elicit are part of the sensing apparatus of the host. In adaptive immune responses, recombination and clonal selection permit the generation of highly diverse, clonally expressed receptors that recognize microbial determinants, but not determinants of the host. However, this system is not called into play until several days after inoculation has occurred, and the host must initially rely on innate immune responses to contain an infection. The mammalian innate immune sensing apparatus has evolved to recognize broadly conserved molecules that are not found in the host but are represented in many microbial taxa. These include molecules such as lipopolysaccharide (LPS), bacterial lipopeptides, double-stranded RNA (dsRNA), and DNA bearing unmethylated CpG motifs. But occasionally, rather narrowly expressed deter-

minants are sensed as well. As discussed below, the G-glycoprotein of VSV is detected via the CD14/TLR4 apparatus of the host, a sensor that is principally devoted to LPS detection in mammals. Moreover, some vertebrates (ﬁsh, amphibians, reptiles) do not exhibit any response at all to LPS, and in some vertebrates (adult birds) the response is minimal. Yet representative birds and ﬁsh retain TLR4 orthologs. The simple model of receptors that are highly degenerate in their speciﬁcity may therefore need to be modiﬁed with time. An immune system must also create an environment that is inhospitable or lethal to microbes. This implies an effector arm of immune function. Processes that we associate with inﬂammation, including the generation of toxic radicals within phagocytic cells, the elaboration of hydrolytic enzymes, and mechanisms for containment of microbes (e.g., coagulation, granuloma formation), are components of the effector arm. It is reasonable to think that inﬂammation evolved chieﬂy to counter infection. Yet sterile inﬂammation, when it occurs, is a major cause of morbidity. In all likelihood, it depends on the same biochemical pathways as those used by the innate immune system. The ﬁnal requirement of the immune system is self-tolerance. A part of self-tolerance is self-/nonself-discrimination. This is accomplished by the innate immune system through an evolutionary process: The principal receptors of the innate immune system fail to recognize molecules of the host, at least under normal circumstances. There may be exceptions in that TLR3, -7, -8, and -9 recognize nucleic acids and, at least under some conditions, are able to recognize mammalian nucleic acids (14). Hence, one may speak of innate autoimmunity and adaptive autoimmunity (15) rather than autoimmunity per se. In addition to self-/nonself-discrimination, self-tolerance assumes that bystander effects should be minimized. As already mentioned, inﬂammation often damages healthy www.annualreviews.org • Genetic Analysis of Host Resistance

VSV: vesicular stomatitis virus LPS: lipopolysaccharide

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tissues. When infections are widespread, the process may be lethal. This appears to be a necessary imperfection in a system designed to enforce containment of small infections before they grow out of control. Small inocula occur far more commonly than inocula that are so large as to provoke life-threatening responses; hence, an aggressive system for dealing with microscale infections would be favored by evolution, even if this system is harmful when infections involve a substantial volume of tissue.

Figure 2 The Lps mutation held the key to understanding innate immune sensing. All LPS-induced phenomena were abolished by the mutation, which was widely assumed to affect the LPS receptor.

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Microbial Sensing, LPS, and the Link Between Pathology and Protection Molecules detected by the innate immune system began to be identiﬁed in the early part of the twentieth century, and structural analysis of some such molecules was completed more than 30 years ago (16), leaving open the question of which receptors must be responsible for their recognition. The prototypic microbial inducer, sensed strongly by mammals although not by all vertebrates (17), is LPS. The toxicity of LPS also depends on mononuclear phagocytic cells (18); most other cells of the host are essentially unresponsive to it. Ultimately, toxicity depends on speciﬁc cytokines elaborated by these cells, notably tumor necrosis factor (TNF) (19) and type I IFN (20). Widely used to induce local and systemic inﬂammation and known to reproduce most of the pathologic features of sepsis, LPS elicits responses at picomolar concentrations, a fact that spoke strongly in favor of the existence of a speciﬁc receptor. The dependence of LPS signaling on a single protein was revealed by the identiﬁcation, in 1965, of a remarkable phenotype in mice (21) seemingly caused by a spontaneous mutation and later traced to a single locus, termed Lps (22–24). The phenotype was ﬁrst observed in mice of the C3H/HeJ strain and was marked by a very speciﬁc and profound insensitivity to LPS. Neither the lethal effect of LPS nor any of the cellular effects of LPS occurred in these mice. This included the well-known adjuvant effect of LPS (25), i.e., its ability to provoke an adaptive immune response to coadministered protein antigens (26). Later, it was shown that mice of the C57BL/10ScCr strain had an allelic defect (27). The Lps mutation was widely believed to affect the LPS receptor or an essential component of the receptor, but formal proof awaited positional cloning data (Figure 2). Long in advance of the positional cloning of Lps, researchers showed that failure to sense LPS lowered the mean lethal inoculum

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of Salmonella typhimurium organisms by approximately four orders of magnitude (28, 29). Similar observations were later made for other Gram-negative microbes administered by other routes (30, 31). These studies provided solid proof that sensing LPS was important to an effective innate immune response to Gram-negative microbes or, conversely, that LPS was one of the key molecules recognized by the innate immune system. Finding the Lps locus assumed great practical importance because it seemed to offer the key to understanding innate immune sensing. Hidden in these studies was an important principle already mentioned above: The innate response evolved to be highly aggressive on a microscopic scale. The same response, when disseminated, can be lethal to the host, as in the example of endotoxin-induced shock. The innate response has evolved to balance the need for containment of microscale microbial infections, which are very common, against the risk of sepsis. Sepsis occurs as the result of a large infusion of microbes (an uncommon event) or because early containment of an infection has been unsuccessful. Sepsis may be seen, in most cases, as the result of innate immune failure.

The Basis of Innate Immune Sensing as Revealed by a Phenotype The positional cloning of the Lps locus was achieved by following a phenotype (failure of macrophages to produce TNF in response to LPS). This was deemed a biologically relevant phenotype because TNF had earlier been shown to be one of the central executors of the toxicity of LPS (19). TNF production was well known to depend on nuclear translocation of NF-κB (32). However, the primary signals elicited by LPS were unknown at that time. The mutation responsible for LPS unresponsiveness was found to be a missense error (P712H) altering the cytoplasmic domain of TLR4 (33). TLR4 was then known only as one of ﬁve mammalian paralogs of

Toll (34–37), a Drosophila protein with a dual role in development and in innate immunity. Speciﬁcally, ﬂies with mutations affecting Toll or components of the Toll signaling pathway failed to mount an adequate antimicrobial peptide response to fungi (10) [and, as was later shown, to Gram-positive bacteria as well (38)]. The Toll superfamily also included the IL-1 receptor (IL-1R) and IL-18R, which bear cytoplasmic domain homology to Toll, provoke inﬂammatory responses, and activate NF-κB (39–41). Because IL-1R and IL18R served immune functions, investigators speculated that TLR proteins in mammals might have either developmental functions (35) or immune function (36), or perhaps both. In fact, investigators had shown that ligation of TLR4, enforced by the creation of a chimeric protein in which a sequence encoding CD4 was substituted for the native TLR4 ectodomain-encoding sequence, could activate NF-κB in transfected cells (36), an observation consistent with the fact that Toll and IL-1R could activate NF-κB. However, this observation in itself did not actually address the function of the TLRs because NFκB activation has many consequences, some related to immunity and some not. Moreover, the experiment did not give any insight into the natural ligand for TLR4, nor did it reveal whether the ligand was endogenous (as in the case of Toll, IL-1R, and IL-18R) or exogenous. The fact that a point mutation in TLR4 entirely prevented mammalian responses to LPS and to Gram-negative bacteria, yet had no developmental consequences, strongly favored the interpretation that the TLRs had an immunologic function and speciﬁcally suggested that the TLRs acted as the long-sought sensors of molecules made by microbes. Although LPS was sensed speciﬁcally by TLR4, investigators immediately postulated that the other TLRs might sense other inﬂammatory molecules (e.g., dsRNA, unmethylated DNA, glucans, and lipopeptides), and that collectively the mammalian TLR paralogs www.annualreviews.org • Genetic Analysis of Host Resistance

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might alert the host to infection, triggering an immune response.

The Structure of the TLRs

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All TLRs are single-spanning transmembrane proteins with ectodomains largely composed of leucine-rich repeats (LRRs), and with a cytoplasmic domain largely composed of a TIR domain (Figure 3).

Figure 3 Schematic rendering of TLRs, based on the structure of the TLR3 ectodomain and the TLR2 TIR motif. TLR ectodomains are all likely to be dimerized, horseshoe-shaped, curved solenoids composed of numerous repeating LRRs. The cytoplasmic domains are compact and globular, consisting mostly of a TIR motif. In some cases (e.g., TLR2, TLR4), multiple accessory molecules may be required for ligand engagement. Adapter proteins, four of which are known to carry TLR signals, also have TIR domain structures. It is believed that ligands elicit a conformational change, allowing recruitment of speciﬁc adapter(s). The critical proline in the BB loop and the critical valine at the Pococurante site are rendered at the atomic level and colored red and yellow, respectively. These residues are essential for most, but not all, TIR motif interactions, as discussed in text. Ligands are presumed to engage TLRs near the point of interface between LRR subunits. (Rendering of ﬁgure was performed with PyMOL, 2005 DeLano Scientiﬁc LLC.) 360

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The structure of a representative TLR (TLR3) has recently been solved through Xray crystallography, revealing that TLR3 is a dimeric protein composed of two horseshoeshaped subunits that stack together side by side. A large quantity of carbohydrate projects from each horseshoe in one direction, so that only the carbohydrate-free surfaces can remain in contact with one another. At the point of interaction between subunits, a positively charged cluster of residues marks the point of presumed interaction between TLR3 and its ligand, dsRNA (42). The cytoplasmic domains of TLR1 and TLR2 have also been crystallized and their structures solved (43). Each is compact and globular, and there is no evidence of interaction between subunits in the unit cell. A surface structure known as the BB loop is present in each TIR domain and contains the residue corresponding to residue that is altered in the TLR4 protein of C3H/HeJ mice (P712H for TLR4; P681H for TLR2). The mutation does not prevent normal folding of the TIR domain, as shown by crystallization studies (43). However, it does abolish signal transduction from most of the TLRs (with some key exceptions, noted below), indicating that the residue in question is, in those cases, part of the signaling interface for interaction with adapter proteins, as discussed below. The similarity between TIR domains is such that threading programs can be used to model each TIR domain on those that have been directly analyzed by X-ray crystallography (44). It is widely believed that all the TLRs are dimeric, with some homodimeric and others (TLR2/TLR6 and TLR2/TLR1) heterodimeric. Dimeric structure is assumed because enforced dimerization triggers a response (36) and because membraneproximal modiﬁcations of cysteine residues in Toll cause dorsalization of the embryo in Drosophila (45). Moreover, IL-1R and IL-18R are heterodimers and signal via TIR domains, although the extracellular domain of each receptor contains immunoglobulin superfamily repeats. Dimeric structure must be taken

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into account in considering precisely how the TLRs signal. Some TLRs are clearly intracellular, residing predominantly or entirely within the endosomes. These include TLR3, -7, -8, and -9, which sense nucleic acids. TLR1, -2, -4, and -6 are at least largely expressed on the surface, although their presence within the phagosome and later components of the endocytic pathway is not excluded.

Other Components of the LPS Receptor At least some TLRs depend on other proteins to signal effectively (Figure 4). In the case of the TLR2/TLR6 heterodimer, CD36, a double-spanning plasma membrane protein of the class B scavenger receptor family, is known to participate in signaling events, as shown by the phenotype of mutant mice identiﬁed in a TLR signaling screen (46), described in detail below. The TLR2/TLR6 heterodimer also depends in part on CD14 to signal effectively, and mutations that eliminate CD14 partly (∼50%) impair sensing of all TLR2/TLR6 ligands.

The LPS receptor TLR4 depends even more strongly on CD14 than does the TLR2/TLR6 heterodimer, and without CD14, TLR4 cannot mobilize all of the adapter proteins that it requires for full signaling activity. This dependence suggests that TLR4 is organized into a supramolecular complex through interaction with CD14 and with its ligand, LPS. In fact, CD14 was ﬁrst shown to enhance LPS signals in 1990 and was the ﬁrst component of the LPS receptor complex to be identiﬁed (47). Only more recently was it shown, through a forward genetic approach, that CD14 is speciﬁcally required for the detection of smooth LPS (LPS with abundant O-glycosylation) rather than rough LPS or lipid A, and for signaling via the TRIF/TRAM pathway. CD14 recognizes other TLR4 ligands as well, such as glycoprotein G of VSV (P. Georgel, Z. Jiang, S. Kunz, K. Hoebe, E. Janssen, M. Oldstone, and B. Beutler, manuscript submitted). In addition, a small protein known as MD-2 associates with the TLR4 ectodomain and is required for LPS signal transduction. Mutations in MD-2 entirely prevent LPS signaling (48), matching the effect of mutations in TLR4.

Figure 4 Shared and unique components of TLR complexes. Germ line mutations have proven the participation of CD14, CD36, and MD-2 in signaling by TLR2 and TLR4 complexes. Coreceptors broaden the speciﬁcity of the receptor complexes and in some cases inﬂuence the choice of adapters that are recruited. MALP-2 and LTA require CD36 for full signaling efﬁcacy; zymosan and PAM2 CSK4 do not. CD14 is partly required by all TLR2/TLRX and TLR2/TLR6 agonists, is fully required by smooth LPS, and is partly required by rough LPS. TLRX indicates uncertainty concerning the partner for TLR2 in the PAM2 CSK4 receptor complex; it is most likely a second TLR2 subunit. (Abbreviations: VSV-G, vesicular stomatitis virus glycoprotein G; MMTV-G, mouse mammary tumor virus surface glycoprotein.) www.annualreviews.org • Genetic Analysis of Host Resistance

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The Sensing Function of Other TLRs TRIF: Toll/IL-1R domain-containing adapter inducing IFN-β

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TRAM: TRIF-related adapter molecule PAM3 CSK4 : triacylated lipopeptide (N-Palmitoyl-S[2,3bis(palmitoyloxy)(2RS)-propy1]-[R]cysteinyl-[S]-seryl[S]-lysyl-[S]-lysyl[S]-lysyl-[S]-lysine x 3 HCI) MALP-2: macrophage activating lipopeptide 2 LTA: lipoteichoic acid

Targeted gene deletion has revealed that TLR2/TLR1 heterodimers are required for detection of triacylated bacterial lipopeptides or synthetic peptides such as PAM3 CSK4 (49). Some diacylated lipopeptides, such as the Mycoplasma-derived macrophage activating lipopeptide 2 (MALP-2), signal via the TLR2/TLR6 heterodimer, and they do so in a stereospeciﬁc manner: Only the R enantiomer uses this signaling complex (50). The TLR2/TLR6 complex reportedly also recognizes lipoteichoic acid (LTA) and β-glucans such as zymosan, although some caution is needed in interpreting cellular signaling events induced by microbial fractions such as these, given that they are not always entirely pure. Other diacylated peptides (e.g., PAM2 CSK4 ), although TLR2 dependent, require neither TLR1 nor TLR6 (51). Either a TLR2 homodimer or TLR2 associated with TLRX (candidates include TLR11, -12, and -13) might serve recognition of these molecular species. TLR5 recognizes ﬂagellin, a protein represented in both Gram-positive and Gram-

negative microbes (52). TLR7 (in mice and humans) and TLR8 (in humans only) recognize nucleoside analogs such as resiquimod or imiquimod (53), drugs with antineoplastic and antiviral potential that are now believed to mimic the natural ligand ssRNA (54– 56). TLR9 senses DNA bearing unmethylated CpG-containing motifs (57) and does so in a species-speciﬁc fashion (58). Humans (but not mice) express TLR10. Mice (but not humans) express TLR11, -12, and -13. There is some confusion of nomenclature in that TLR11 has been called TLR12 by some authors, and vice versa (59, 60). The molecular speciﬁcity of TLR10 remains unknown, although because of its structural similarity to TLR1 and TLR6, TLR10 likely senses lipopeptides. Its absence in mice has prevented examination of the knockout phenotype. In mice, TLR11 recognizes a proﬁlin-like component of Toxoplasma gondii (61) (evidently distinguishable from the host molecule). The functions of mouse TLR12 and TLR13 have not yet been determined (Table 1). Particularly where nucleotide ligands are concerned, speciﬁcity is not absolute, and

Table 1 TLR ligand specificities, adapters used either alone or in combination with one another, and species representationa Ligands

TLR

Adapters

Species

PAM3 CSK4

1,2

MyD88, MAL

Human, Mouse

PAM2 CSK4

2,X

MyD88, MAL

Human, Mouse

MALP-2, LTA, Zym

2,6

MyD88, MAL

Human, Mouse

dsRNA

3

TRIF

Human, Mouse

LPS, VSV-G, MMTV-G

4

MyD88, MAL, TRIF, TRAM

Human, Mouse

Flagellin

5

MyD88

Human, Mouse

ssRNA, IAQ

7

MyD88

Human, Mouse

ssRNA, IAQ

8

MyD88

Humanb

CpG-ODN

9

MyD88

Human, Mouse

Unknown

10

Unknown

Human

Proﬁlin

11

MyD88

Mouse

Unknown

12

Unknown

Mouse

Unknown

13

Unknown

Mouse

a

IAQ, imidazoquinolines, including resiquimod and imiquimod; MAL, MyD88 adapter–like; CpG-ODN, synthetic oligodeoxynucleotides containing CpG motifs. Other abbreviations as described in text. b TLR8 is encoded in the mouse genome, but activating ligands, if any, are unknown. 362

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host molecules can also trigger inﬂammatory responses, although perhaps less intense responses than those initiated by microbes. TLR3, -7, -8, and -9 are largely (although not necessarily entirely) dedicated to the detection of viral infection. However, other TLRs, most notably TLR4, also play an important role in viral sensing.

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The Net Importance of TLRs in Resistance to Microbes TLRs signal via four adapter proteins (MyD88, Mal, TRIF, and TRAM), discussed in greater detail below. Without the adapter protein MyD88, which is required for all signal transduction by many of the TLRs and for full signaling by TLR4, mice frequently succumb to infections derived from their own oral ﬂora [mixed infections of the submandibular nodes with α-hemolytic streptococci and Pasteurella pneumotropica have been observed (Z. Jiang & P. Georgel, unpublished observations)]. When both MyD88 and the adapter TRIF are absent, mice are even more susceptible to infection, and very few survive to weaning age (B. Beutler & K. Hoebe, unpublished observation). Individual TLR mutations are also compromising. For example, TLR4 mutations cause enhanced susceptibility to Salmonella typhimurium (28) and Escherichia coli (29) infections; TLR2 deletion causes enhanced susceptibility to Staphylococcus aureus (62) and other organisms. TLR3 deﬁciency (59) and to an even greater extent TLR9 deﬁciency (59, 63) cause susceptibility to MCMV infection, a highly prevalent pathogen in wild mice. By implication, the same mutations may enhance susceptibility to β-herpesviruses in other species. These observations suggest that the TLRs make an essential contribution to the recognition of infectious organisms.

The Special Case of Viruses Viruses were the last pathogens found to be recognized by TLRs (59, 63–68), and at the

time, it came as something of a surprise to see that they were so recognized. Viruses are primarily, although not entirely, sensed by TLRs dedicated to the detection of unmethylated DNA, dsRNA, and ssRNA. As such, the challenge of self-/nonself-discrimination is greater because host nucleic acids are only slightly different than virally encoded nucleic acids. ssRNA is, of course, very abundant in host cells, and most RNA species have at least some dsRNA regions. Some CpG dinucleotides within the mammalian genome are unmethylated and should therefore be recognized by TLR9. These facts have led to the suggestion that autoimmune diseases are fueled in part by TLR signaling, stimulated by host nucleoprotein complexes (14). At the very least, viruses test the limits of self-/nonself-discrimination, and why TLRs are so effective in recognizing them is still not entirely clear. TLRs 3, 7, 8, and 9 not only detect structural differences between viral nucleic acids and host nucleic acids, but also must engage the nucleic acids within endosomes, a compartment from which host nucleic acids are normally excluded (69). There are numerous examples of viral protein detection by TLRs as well. The envelope glycoprotein of mouse mammary tumor virus (MMTV) (65, 66) and glycoprotein G of VSV (P. Georgel, manuscript in preparation) are both recognized via TLR4. It has also been reported that the F-glycoprotein of respiratory syncitial virus activates TLR4 (64), although this assertion has recently been challenged (70). The ability of TLR4 to detect viruses is interesting for several reasons. Like TLR3, TLR4 uses the adapter protein TRIF and is capable of stimulating type I IFN responses. But TLR4 is normally thought of as the LPS receptor, and it is surprising to ﬁnd that entirely different molecules are engaged by it. The fact that they are raises structural and evolutionary questions. The ability of receptors to recognize structurally disparate molecules is not without precedent. For example, opiates are plant alkaloid agonists that www.annualreviews.org • Genetic Analysis of Host Resistance

MyD88: myeloid differentiation factor 88 MMTV: mouse mammary tumor virus

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TIR-DEPENDENT AND TIR-INDEPENDENT ADAPTIVE IMMUNE ACTIVATION TLRs mediate adjuvant effects induced by microbes, known since the time of Freund. The adjuvant signals initiated by the TLRs are largely directed toward a CD4 response. But they are not required for adaptive immune activation per se. In the absence of TLR adapter proteins MyD88 and TRIF, TLR signaling is at least greatly diminished. However, normal lymphoid development, IgG production, and allograft rejection are observed. These ﬁndings point to the existence of TIR-independent pathways for adaptive immune activation. Certain types of cell death, including death induced by UVor γ-irradiation or by Fas ligation, are capable of triggering a strong immunoadjuvant pathway that is chieﬂy (although not exclusively) directed toward CTL activation, and entirely TLR independent. This pathway is known to depend on the integrity of the type I IFN receptor and UNC-93B (the 12spanning ER protein affected by the 3d mutation). To the extent that the pathway can (weakly) activate CD4 cells, it is additionally dependent on CD36 (CD4 activation is selectively impaired by the oblivious mutation). However, CD36 plays no part in the internalization of antigen molecules. The TIR-independent immunoadjuvant pathway, which presumably serves the recognition of cell death triggered by certain viral infections, is currently being analyzed using ENU mutagenesis.

IRF: IFN regulatory factor IRAK: IL-1R-associated kinase

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trigger a response from endorphin receptors. However, it is a different task for receptors to maintain contact with multiple targets that are driven to evade detection. In the case of VSV glycoprotein G, mutations might nullify recognition, permitting the virus to evade detection. In the case of the MMTV envelope glycoprotein, investigators believe that the virus uses TLR4 for a subversive purpose, stimulating IL-10 production rather than the full spectrum of TLR signals, and minimizing the immune response (66).

tity long known as something made by microbes that was sensed by the host (33). TLRs are the key sensors of microbial infections of many and perhaps all types. It follows that they are ultimately responsible for most (although not necessarily all) infection-related phenomena, no matter how complex those phenomena may be (15). One of these infection-related phenomena is the adjuvant effect of microbes. The gold standard of adjuvants is Freund’s complete adjuvant, which is made using mycobacteria suspended in an oil-in-water emulsion. Molecules of microbial origin must have adjuvant properties. In 1955, investigators showed that LPS is endowed with adjuvant properties (26), and in subsequent years other molecules of microbial origin were also shown to have adjuvant effects (71–76). Among the key molecular events leading to an adaptive immune response is upregulation of costimulatory molecules (including but not limited to CD80 and CD86), which must occur on the surface of antigen-presenting cells (APCs) to activate T cell mitogenesis (77). LPS elicits upregulation of these molecules on APCs. Medzhitov and colleagues (36) once speculated that this occurs through the ability of TLRs to activate NF-κB. However, it is now clear that the key event in upregulation of costimulatory molecules is the activation of a type I IFN response (78, 79). This, in turn, depends on LPS activation of the MyD88independent pathway and on activation of the adapters TRIF (67, 80), TRAM (81), and IRF3, discussed in more detail below. Other microbial adjuvants activate type I IFN synthesis through activation of the adapter MyD88, via IRAK-1 and IRF-7 (82).

The Connection Between Innate and Adaptive Immunity

TIR-Dependent and TIR-Independent Pathways: A Death-Driven Pathway for CTL Activation

As described so far, the function of TLRs in mammals was originally established by ﬁnding the receptor for LPS, a molecular en-

The fact that TLRs mediate microbial adjuvanticity led to early suggestions that they might mediate adaptive immune activation at

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Figure 5 An immunoadjuvant pathway driven by programmed cell death. The pathway does not depend on TLR signaling, and the triggering ligands induced by programmed cell death are unknown. The 3d mutation prevents the APC from presenting exogenous antigen, so that both priming and cross-priming are inhibited. The oblivious mutation prevents priming, but not cross-priming. The process also depends on the production of type I IFN and on its signaling via IFNAR on the APC.

large. However, in mice that lack two of the adapters required for TLR signaling (MyD88 and TRIF), there is little or no response to most microbial ligands. Although we have already noted that such mice are drastically immunocompromised (K. Hoebe & B. Beutler, unpublished observation), they exhibit normal concentrations of circulating antibodies, have normal lymphoid architecture, and can reject allografts. Although Freund’s adjuvant does not promote an immune response in such mice, alternative pathways for adaptive immune activation clearly exist and are presumably activated by nonmicrobial stimuli. The danger model of costimulation holds that cell death or injury may provide the essential signal for an adaptive immune response (83). It appears that some, but not all, forms of cell death trigger a strong and TIR-independent signal that leads to the induction of type I IFN synthesis, as well as to priming and cross-priming of T cells with speciﬁcity for antigens expressed by the dy-

ing cell (Figure 5). Programmed cell death induced by γ- or UV-irradiation or by Fas ligation will trigger such a response (E. Janssen, K. Tabeta, M. Barnes, S. McBride, S. Schoenberger, A. Theoﬁlopoulos, B. Beutler, and K. Hoebe, manuscript submitted). The cells that are capable of sensing programmed cell death are B220− and CD8low lymphoid elements, and they can be differentiated from bone marrow precursors using Flt-3 ligand. They acquire antigen by nibbling. Notably, the system is not represented in myeloid DCs or in macrophages; the macrophages probably dispose of many or most of the cells that die through senescence or other processes of attrition in vivo. The system for adaptive immune activation may be called into service when normal routes for removal of apoptotic cells are overwhelmed. Moreover, cell death that is induced by pathologic stimuli (viruses, irradiation) is likely qualitatively different than that occurring as a result of constitutive apoptosis, and it www.annualreviews.org • Genetic Analysis of Host Resistance

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leads to the expression of recognizable surface molecules that are as yet unknown. The death-driven pathway evokes a far more powerful cytolytic T lymphocyte (CTL) response than Freund’s adjuvant. Antigens are usually detected by the system when injected into mice in picogram to nanogram quantities, expressed by syngeneic cells that have been induced to undergo apoptosis. The system presumably evolved to detect cell death induced by viral infections and comprises a route to rapid adaptive immune activation in response to this stimulus. Because it is TIR independent, it would be effective when TLR sensing is not. This system of recognition conceivably participates in diverse immunological events, including allograft rejection and autoimmunity.

THE INTENTIONAL CREATION OF PHENOTYPE Although a spontaneous mutation led to the concept that TLRs detect molecules of microbial origin, investigators have had to induce mutations, either deliberately or at random, to dissect the TLR signaling pathways and to identify participating molecules that are structurally dissimilar from those that are known. The induction of mutations that cause phenotypes of interest, and subsequent identiﬁcation of those mutations, is the essence of the classical or forward genetic approach. The germ line mutagen used for this purpose is N-ethyl-N-nitrosourea (ENU).

ENU and Its Efficiency as a Mutagen In using a germ line mutagen, one would ideally like to create as many mutations as possible, consistent with the production of monogenic (as opposed to polygenic) phenotype, because only monogenic phenotype is readily mapped and positionally cloned. Practically speaking, ENU dosing is limited by the fact that male mice become sterile if administered too high a dose. ENU has been tested in many different strains of mice, and the standard pro366

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tocol that has emerged, in C57BL/6 mice, is administration of 100 mg/kg three times at weekly intervals. A 12- to 15-week period of infertility follows. During this interval, the testis is repopulated by spermatogonia, arising from approximately 10 to 100 precursors. After this, mice may be bred to transmit germ line mutations to the G1 population. For studies of immunity, C57BL/6 females are best used in breeding to maintain a pure genetic background. Among G1 mice, dominant phenotype may be identiﬁed. To produce recessive phenotype, G1 mice are crossed again to C57BL/6 females, and then to their own daughters. Among G3 mice, recessive phenotype may be identiﬁed. The potency of a germ line mutagen is ultimately limited by the mutational load compatible with life. Even if ENU did not induce sterility, it clearly induces nearly the maximum number of mutations tolerated in the G3 population. Kile et al. (84) have analyzed the frequency of lethal hits induced by ENU within a restricted area of the genome using a balancer chromosome, a dominantly marked, inverted chromosome that carries a homozygous lethal mutation. Balancer chromosomes can be used to capture lethal mutations because only carriers of the balancer are evident in the G3 population, whereas homozygotes for the mutagenized chromosome and homozygotes for the balancer itself are absent. Approximately 1/13 G1 mice born to mutagenized sires carried lethal hits within the balancer region, with lethality deﬁned as a failure to survive to the age of weaning. Because the inverted region encompassed only about 5% of all genes in the genome, and because lethal hits occur elsewhere as well, it may be surmised that each G1 mouse is, on average, heterozygous for about 1.5 recessive lethal hits genome-wide. If a G1 mouse bears a lethal hit, the probability of its transmission to homozygosity in each G3 mouse is 1/8 (0.125), and the likelihood of nontransmission to homozygosity in each G3 mouse is 1–0.125, or 0.875. Hence, 1–0.8751.5 , or about 18%, of the G3 population is lost as

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a result of homozygosity for ENU-induced mutation. If the efﬁciency of ENU were tenfold higher (15 lethal hits per G1 mouse), 86% of the G3 population would be lost, an amount of attrition that might be considered undesirable because multiple litters would be required to produce a very small number of mice for screening. In effect, ENU provides mutagenic power approaching the maximum that can be tolerated.

The Concept of a Genomic Footprint For any phenotype that the investigator deﬁnes, a certain and very exact set of nucleotide changes across the genome are capable of producing that phenotype. These nucleotides comprise the genomic footprint of the phenotype in question. Estimates from direct sequencing of sentinel regions of G1 genomic DNA reveal that ENU changes about 1 base pair (bp) per million bp in the haploid genome (85), and 1/8 of these changes are transmitted to homozygosity in each G3 mouse. If we assume that all bp within a genomic footprint are equally at risk from ENU, it is possible to calculate the size of the genomic footprint based on the frequency of observed phenotypic change. If 100 bp are at risk, and if pedigree construction is such that 50% of G1 mutations are captured in the homozygous state (e.g., two G2 mice per G1 sire and three G3 mice per G2 dam), one would expect to see the phenotype once, on average, for every 120,000 G3 mice examined. If 100,000 bp are at risk, the phenotype should be apparent once in every 120 G3 mice examined. Probably the phenotype with the largest genomic footprint of all is lethality, which, as noted earlier, is realized in about 18% of G3 mice, suggesting a genomic footprint of about 1.6 million bp. The genomic footprint of a phenotype may be small because it is masked by other phenotypes (typically lethality). For example, a particular immunodeﬁciency phenotype might rarely be observed because mice do not survive

to the age required by the assay. Alternatively, the genes that support a particular immune function might also be required for development in utero, and only the exceptional nucleotide change will produce the phenotype yet be compatible with viability to term. Although viable hypomorphic alleles probably exist for all genes, the nucleotide substitutions that produce them are sometimes scarce. Rather, it is more common that a mutational change has no measurable effect on the function of a protein at all, or that it utterly destroys the protein. On other occasions, the genomic footprint of a phenotype may be small because the phenomenon under analysis is served by redundant pathways. Few genes are truly redundant because locus duplication most commonly leads to the formation of a degenerate pseudogene. Most commonly, the claim of redundancy belies a screen that is not sufﬁciently powerful to resolve genes with overlapping function, although the functions of the two genes are actually distinguishable. But some recently duplicated genes do exist in the genome, and their functions, although important, might not be revealed by ENU mutagenesis.

Genomic footprint (of a phenotype): the set of all nucleotides that can cause the phenotype in question when mutated

How the Footprint Is Parceled into Genes The genomic footprint of a phenotype is scattered among genes that support the phenomenon under study. The mathematical distribution of the target nucleotide population among genes has not been described and is not known from experiment because genome-wide saturation of a phenotype has not been approached. Gene size presumably inﬂuences the distribution; perhaps more inﬂuential still is variation in the resilience of encoded proteins to mutagenic change. In addition, it is possible (though uncertain) that nucleotides differ regionally in their susceptibility to ENU. The distribution of the genomic footprint among genes is a matter of importance in www.annualreviews.org • Genetic Analysis of Host Resistance

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considering the approach to saturation. Starting with the assumption that all nucleotides are equally vulnerable to change induced by ENU [which is assuredly untrue because the great majority of base changes are known to be A → T transversions or A → G transitions (86)], and with the fact that 1/8 of 1 millionth of the genome is altered in homozygous form in each G3 mouse, we might imagine that 5.9% of all nucleotides would be struck if one million G3 mice were examined, following the same pedigree parameters as those outlined above. One might thus assume that all genes relevant to the phenotype would be revealed. This assumption, however, depends entirely on the distribution of the target nucleotide population. A given phenotypic footprint might have 10,000 nucleotides (a typical number) parceled into 50 genes (perhaps also a typical number). The average gene would have 200 target nucleotides and would indeed be revealed. But a gene with 10 vulnerable nucleotides would have only a 46% chance of being revealed. And the footprint might include many such genes. At any rate, no ENU screen has yet approached a depth of one million mice. According to our best estimate, using a protocol in which half of all mutations are transmitted to homozygosity, ∼10% phenotypic saturation is achieved with the analysis of ∼10,000 G3 mice.

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Some Summary Statistics Almost all ENU-induced phenotypes result from coding change (missense errors, nonsense errors, splicing errors, or, more rarely, single base pair deletions), although exceptions have been reported (87, 88). The coding region of the mouse genome encompasses approximately 42 Mb of DNA (1.5% of the genome as a whole). It may therefore be calculated that if ENU creates 1 bp change per million bp, about 42 mutations fall within parts of the genome where a phenotypic effect can be exerted. About 76% of nucleotide changes 368

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that fall within a coding sequence create coding change. Therefore, 32 coding changes occur in each G1 mouse born to a mutagenized sire. About four of these changes are transmitted to homozygosity in each G3 mouse. About 18% of mice die as a result of four random homozygous changes in coding sequence, which suggests that each individual change has approximately a 4.8% chance of delivering the lethal blow. The number of genes that are targets in this process (i.e., the number of genes with recessive lethal alleles) remains unknown. But if we assume that 1/3 of all genes have lethal alleles (a low-end estimate), we could conclude that, on average, about 16% of random coding changes within genes that can cause lethality do cause lethality. If 2/3 of all genes have lethal alleles (a high-end estimate), the ﬁgure would be 8%. This reveals the average resilience of proteins with respect to a deﬁned qualitative phenotype.

The Design of Screens A good screen reﬂects the designer’s appreciation of a biological phenomenon and curiosity about its cause. It avoids phenomena that are well understood, avoids redundancy, and, where possible, probes a large genomic footprint. The screening assay must be sufﬁciently robust to avoid even rare false positives. It is best if the assay is qualitative or, better still, binomial. But whatever the screen, phenotypes selected for analysis must be strong enough and penetrant enough to permit meiotic mapping, without which no progress can be made.

THE GENETIC DISSECTION OF TLR SIGNALING PATHWAYS What Was Known from the Start Analysis of TLR signaling pathways began before the mammalian TLRs were recognized, with the discovery that IL-1R signaled by way of the adapter protein MyD88 (89) and

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Figure 6 Proteins known prior to beginning ENU mutagenesis to participate in signaling from seven of the TLRs to the level of TNF activity.

with recognition that the serine kinase IRAK transduces IL-1 signals (90, 91), depending on TRAF-6 (92) and leading to the activation of NF-κB. Similarities between the IL-1 signaling pathway and the Toll developmental pathway were noted early on (93), as was the similarity of the IL-1R cytoplasmic domain to the cytoplasmic domain of Toll (94). Early and rapid progress in understanding TLR signal transduction rested on this similarity. Homology searches led to the identiﬁcation of all the TLRs that are presently known, and gene targeting established the speciﬁcity of most of them once the LPS sensing function of TLR4 was determined by positional cloning (33). Homology searches have also disclosed the existence of at least four cytoplasmic TIR adapter proteins. Whereas MyD88 and Mal (also known as Tirap) were easily detected by using the Basic Linear Alignment Search Tool (BLAST), TRIF and TRAM were far more distant and were found belatedly using the Hidden Markov Model search algorithm (HMMER).

However, the participation of proteins structurally unrelated to the TLRs was much more difﬁcult to ascertain, and in that research a pure genetic approach has also played a valuable role. By 2000, a total of 22 proteins were known to participate in TLR signaling from the level of seven of the TLRs to the level of TNF bioactivity, and these proteins could be designated as potential targets (Figure 6). A genetic screen was applied to identify additional components of the pathway. The screen was performed by stimulating peritoneal macrophages with TLR agonists and then measuring TNF production by biological assay (killing of L-929 cells). To date, a total of 11 mutations have been identiﬁed by screening approximately 20,000 mice. These mutations alter a total of 10 genes. Five of the genes were known at the outset of screening. The other ﬁve, however, were unknown components of the TLR signaling apparatus. Three of these have been positionally cloned to date. www.annualreviews.org • Genetic Analysis of Host Resistance

TRAF-6: TNF receptor–associated factor 6

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Calculation of Pathway Complexity

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A forward genetic approach permits unbiased estimation of the total number of protein components that are required for signaling from TLRs to TNF because each serves a nonredundant function. Given that 5 of the genes identiﬁed fell within the category of known genes (22 of which exist in all) whereas 5 did not, it might be guessed that 44 targets exist in total. Taking an independent approach to estimate the number of nonredundant proteins, 23% (5/22) of the known protein population was identiﬁed in the screen, corresponding to the degree of saturation achieved in a study of 20,000 mice. If 23% saturation nets 10 genes, 43 targets may be estimated to exist in all.

Figure 7 The position of Lps2 on the signaling map. The phenotype imparted by the mutation was one in which TLR3 and TLR4 were both unable to activate IRF-3. Production of TNF was also minimal, presumably because the normal protein also contributes to the activation of NF-κB. 370

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Independently again, if one assumes that all targets have uniform size (each subsumes an equal fraction of the genomic footprint), the identiﬁcation of one instance of allelism among 11 mutations would suggest the presence of approximately 53 targets. Although these estimates are tentative and deeper saturation must be pursued to determine the number of targets with conﬁdence, one may conclude that a relatively small number of targets exist and that by this time most of them are probably known.

Lps2 and the Identification of TRIF MyD88-deﬁcient mice showed evidence of a MyD88-independent pathway, whereby LPS, acting via TLR4, could partially stimulate NF-κB translocation after a brief delay and activate type I IFN production without impediment (95). With the targeted deletion of both MyD88 and Mal genes, the same phenomenon was observed (96). TLR3 signaling also had no requirement for either adapter. The molecular basis of the MyD88independent pathway remained unclear. The hallmark of MyD88-independent signaling was not NF-κB activation, but rather IRF-3 activation and subsequent type I IFN synthesis. However, the MyD88-independent pathway is also capable of activating NF-κB and presumably does so by activating a component of the cascade that is distal to MyD88 itself, thereby bypassing the lesion imposed by mutations in MyD88 and/or Mal (Figure 7). Researchers now believe that the point of intersection between MyD88-independent and MyD88-dependent pathways is TRAF-6 (97, 98), a protein that coordinates the activation of numerous kinases, including TAK-1 [transforming growth factor (TGF)-β-activated kinase 1], which is required for the phosphorylation of IKKβ (99, 100), leading to NF-κB activation. A mutation termed Lps2 ﬁrst revealed the molecular basis of MyD88-independent signaling. The Lps2 mutation prevented IRF-3 phosphodimer formation and IFN synthesis

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in response to either LPS or dsRNA, indicating that it affected a protein required for signaling from both TLR4 and TLR3. Moreover, Lps2 rendered mice susceptible to infection by MCMV (101). Positional cloning of the Lps2 mutation revealed a frameshift error in a new adapter protein, independently identiﬁed by homology searching (102) and by two-hybrid system analysis (103), and respectively known as TRIF or TICAM (TIR-containing adapter molecule)-1. On the basis of transfection studies, investigators concluded that this protein stimulates type I IFN production; however, investigators initially disagreed as to whether the protein served all TLRs (102) or only TLR3 (103). The phenotype of the mutation revealed a speciﬁc role in signaling via TLR4 and TLR3 (104), a conclusion conﬁrmed by gene targeting (80). Compound mutants lacking MyD88 and homozygous for Lps2 show no detectable responses to LPS and, indeed, show no responses at all to most TLR ligands (104). However, researchers have recently observed that TLR4 can indeed signal by way of TRAM, even in the absence of both MyD88 and TRIF, when activated by the VSV glycoprotein G (P. Georgel, Z. Jiang, S. Kunz, K. Hoebe, E. Janssen, M. Oldstone, and B. Beutler, manuscript submitted).

Inferences Concerning the Function of TRAM BLAST analysis using the TRIF protein sequence as a query disclosed a fourth adapter, termed adapter X (104) and known elsewhere as TICAM-2 (105) or TRAM (81). A speciﬁc role for TRAM in TLR4 signaling was suggested by the ﬁnding that in cells homozygous for the TrifLps2 allele, a fraction of macrophages remained capable of producing TNF in response to LPS. This TRIFindependent population of cells must depend on another adapter, and that adapter could not be MyD88 because all cells (and not merely some of them) were MyD88-

dependent; hence, MyD88 must be expressed and used in all cells (67). Hoebe et al. proposed that adapter X was TRAM, and that TRAM must serve MyD88-independent LPS signaling alongside TRIF, but not dsRNA signaling (67). Gene targeting, performed independently (81), substantiated this proposal (Figures 8 and 9). It is now clear that TRAM (but not TRIF) mediates signals initiated by the VSV glycoprotein G, which depends on CD14 and TLR4. Hence, TRAM can function entirely by itself and is not necessarily codependent on TRIF (P. Georgel, Z. Jiang, S. Kunz, K. Hoebe, E. Janssen, M. Oldstone, and B. Beutler, manuscript submitted).

Heedless Gives Fresh Insight into the Nature of the TLR4 Signaling Complex The Heedless mutation was identiﬁed because it prevented TNF production in response to smooth LPS, that is, LPS with abundant Oglycosylation. Oddly, however, unlike TLR4 mutants (which show no responses to LPS of any kind), Heedless mutants were able to make almost normal quantities of TNF in response to rough LPS or synthetic lipid A (106). Remarkably, lipid A could not activate IRF-3 phosphodimerization, nor could it trigger type I IFN production, suggesting a defect of MyD88-independent signaling (106). Macrophages from Heedless mice also showed diminished responses to all TLR2/TLR6 ligands and were less capable of coping with infection by VSV, as were mice with the P712H mutation of TLR4 (106). The positional cloning of Heedless revealed a premature stop codon in CD14, previously regarded as a coreceptor for LPS signaling in general. The new interpretation of CD14 function is therefore as follows: 1. Rough LPS or lipid A can directly engage TLR4/MD-2 complexes, causing recruitment and activation of www.annualreviews.org • Genetic Analysis of Host Resistance

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MyD88, whereas smooth LPS cannot (Figure 10a, left); CD14 nulliﬁes the distinction between rough and smooth isoforms, permitting both types of LPS to activate TLR4 (Figure 10a, right); CD14 is absolutely required for LPSinduced recruitment of TRIF and TRAM, which leads to the activation of IRF-3; CD14 is additionally required for full activation of the TLR2/TLR6 complex (which, as noted below, also requires CD36 for sensing of some ligands) (Figures 8, 9). CD14 and TLR4 mediate sensing of VSV (Figures 8, 9).

Because CD14 is a glycosylphosphoinositol-tethered protein that projects from the surface of the cell, it would seem incapable of directly interacting with the cytoplasmic domains of TLRs and must inﬂuence signaling through interaction with the ectodomains. We may conclude that CD14 coordinates the interaction of ectodomains so as to permit the recruitment of adapter molecules that would otherwise be excluded from that activated complex (Figure 10b). These observations establish that TLR4 is a switch with several possible “on” positions. It can signal in different modes depending on the inducing ligand and on the proﬁle of coreceptors and/or adapters present in the responding cell. Macrophage production of IFN-β, induced by UV-inactivated VSV, is abolished by the Heedless mutation, as well as by mutation of TLR4 or TRAM (or, to a much lesser extent, TRIF), and by the Feckless mutation (described

below). The inducing molecule present in the virion is glycoprotein G rather than a nucleic acid (P. Georgel, Z. Jiang, S. Kunz, K. Hoebe, E. Janssen, M. Oldstone, and B. Beutler, manuscript submitted). Glycoprotein G of VSV is a conserved molecule, broadly represented in Rhabdoviridae, encompassing vertebrate pathogens such as VSV, rabies, and hemorrhagic viral septicemia virus of ﬁsh. As already noted, LPS is chieﬂy a stimulus to mammalian cells. Yet TLR4 is represented in ﬁsh, which do not respond to LPS. We may speculate that its primordial function entailed the detection of viruses, which would explain its connection to the type I IFN induction pathway. This function has been retained in mammals as well, but only in mammals has the ability to sense LPS also been acquired.

CpG1 and the Role of TLRs in Resistance to Viral Infection TLR3, -7, -8, and -9 are at least mostly intracellular molecules, synthesized in the ER, and are eventually transferred to the endosomes, possibly via transit over the cell surface and endocytosis (107). Several mutations affected signaling via these TLRs. The ﬁrst of these to be identiﬁed was one that blocked CpG sensing (CpG1). It was traced to a missense error in the ectodomain of TLR9. The Lps2 mutation had earlier suggested the importance of the TLR3 → TRIF axis in sensing MCMV infection and responding to it (67). The CpG1 mutation was associated with even more severe susceptibility to MCMV (59), as was a MyD88 mutation (59), indicating that the TLR9 → MyD88 and TLR3 → TRIF

−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→ Figure 8 An overview of the pathways discussed in text. All TLRs, whether located at the cell surface (TLR1, -2, -4, -5, and -6) or within endosomes [TLR3, -7 (or -8), and -9], converge on four adapter proteins, which act in discrete combinations with one another. These adapters lead to activation of protein kinases, including IRAK, IRAK4, TBK1, and IKKi. These kinases ultimately lead to activation of transcription factors, including NF-κB, IRF-3, and IRF-7, which mediate many of the inﬂammatory effects of microbial inducer molecules. Among the thousands of genes modulated by TLR signaling, TNF and IFN-β may be taken as landmarks of the response and are themselves known to be of key importance in subsequent activation of innate and adaptive immune responses. 372

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axes played nonredundant, additive roles in MCMV resistance. Although both axes contribute to type I IFN production, they may have type I IFN–independent effects as well, some of which may be nonoverlapping and may contribute to resistance (e.g., the activation of IL-12 production, which leads to IFN-γ production). The importance of TLR9 and MyD88 in MCMV resistance was independently analyzed using knockout mice, with similar conclusions (63). Moreover, TLR9 was shown to contribute to the response to herpes simplex virus I (HSV-I) (68).

The Poc mutation causes pronounced susceptibility to MCMV. However, mice are spared the severe immunodeﬁciency disease caused by a MyD88-knockout mutation and are demonstrably more resistant to at least some Gram-positive infections, suggesting that limited signaling through a single TLR complex (TLR2/TLR6) is sufﬁcient to offer fairly strong protection against a wide variety of microbes. The Lkd mutation has little or no effect on susceptibility to MCMV, probably because some signaling via TLR9 is retained in Lkd homozygotes, compared with the situation in CpG1 homozygotes.

Pococurante and Lackadaisical, MyD88 Alleles with Receptor-Selective Function, and Their Interpretation

Pococurante and Insouciant Reveal Something Special about MyD88 Signaling from the TLR2/TLR6 Complex

The Pococurante (Poc) phenotype is one in which there is an absence of MyD88 signaling, with the exception of signals that emanate from the TLR2/TLR6 heterodimer (Z. Jiang, P. Georgel, C. Li, J. Choe, K. Crozat, S. Rutschmann, X. Du, T. Bigby, S. Mudd, S. Sovath, I. Wilson, A. Olson, and B. Beutler, manuscript in preparation). The Lackadaisical (Lkd) phenotype shows normal MyD88 signaling, except for signals from TLR7 and TLR9, which are markedly diminished. Both mutations are missense errors in MyD88, each distinguishable from the knockout allele (Figure 11). The Poc mutation (I179N) affects a surface residue within the TIR domain of MyD88, whereas the Lkd mutation (Y116C) affects a portion of the polypeptide chain between the death domain and the TIR domain. Because the modiﬁed adapter protein is, in each case, capable of transmitting signals from some of the TLRs, it may be inferred that the mutations do not destroy the protein. Rather, they must affect the signaling interfaces that unite the adapters and receptors.

Poc is a particularly interesting mutation because it does disrupt signaling initiated by some TLR2/TLR6 ligands, notably zymosan and LTA. However, it does not disrupt signaling by MALP-2 or PAM2 CSK4 , both diacylated lipopeptides that signal via the TLR2/TLR6 heterodimer and via TLR2 (alone or in conjunction with a still unknown TLR), respectively. Insouciant (Int), a point mutation in the ectodomain of TLR6 (V327A), abolishes zymosan, LTA, and MALP-2 signaling but not PAM2 CSK4 signaling. When engrafted onto TLR4 or TLR9, the Poc mutation abolishes MyD88-dependent signaling. However, when engrafted onto TLR2 or TLR6, the mutation does not affect signaling by MALP-2 or PAM2 CSK4 . Moreover, the BB loop mutation, if engrafted onto MyD88 (P200H), does not prevent downstream signal transduction. These observations suggest that certain TLR2/TLR6 and TLR2/TLRX ligands

−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→ Figure 9 An overview of the pathways discussed in text, with mutants shown with red X. Mutations shown were induced with ENU, and all but two (Spacey and Feckless) have been identiﬁed positionally. 374

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(MALP-2 and PAM2 CSK4 ) cause a conformational change in the receptor that favors a unique mode of MyD88 association with the receptor complex (Figure 12). Although most TLRs (and for some ligands, TLR2/TLR6 as well) associate with MyD88 in a manner that causes both the BB loop and the Poc site to be included in the signaling interface, diacylated lipopeptides trigger a different form of receptor activation, requiring the participation of neither residue. The TIR domains of MyD88 and the TLR2/TLR6 receptor complex likely engage one another in a reciprocal manner in both cases. However, MyD88 clearly has two different ways of propagating a signal.

Oblivious: A Co-Factor for TLR2/TLR6 Signaling

Figure 10 (a) The Heedless mutation reveals that TLR4 can signal in two modes. Without CD14, Lipid A activates TLR4, causing recruitment of MyD88 and MAL (MyD88 adapter–like) only, whereas smooth LPS (long polysaccharide chain) cannot activate TLR4 at all. In the presence of CD14 (bent solenoid), both smooth and rough LPS can recruit both MyD88/MAL and TRIF/TRAM pathways. (b) Surface view of TLR4 dimer, with or without MyD88. A supramolecular effect on TLR4 aggregation is envisioned, so that with CD14, the geometry of interaction between subunits is correct for recruitment of all adapters. Without CD14, smooth LPS is excluded from the activation complex and Lipid A is only able to stimulate MyD88/MAL recruitment.

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Still another point of complexity concerning TLR2/TLR6 signaling is that the diacylated lipopeptide MALP-2 (but not PAM2 CSK4 ) and LTA are largely dependent on CD36 for signaling, a fact revealed by a premature stop codon in CD36 that caused a phenotype called oblivious (46). Other TLR2/TLR6 ligands (zymosan, for example) are not. Hence, the TLR2/TLR6 complex uses CD36 as a coreceptor for some of its ligands. Perhaps for this reason, oblivious mice show exaggerated susceptibility to Gram-positive infections (46). The oblivious mutation also prevents CD4 priming by cells exposed to antigens in the context of apoptosis. Thus, oblivious impairs one endpoint of the deathdriven immunoadjuvant pathway described earlier in this review (E. Janssen, K. Tabeta, M. Barnes, S. McBride, S. Schoenberger, A. Theoﬁlopoulos, B. Beutler, and K. Hoebe, manuscript submitted). This impairment suggests that CD36 has other immunological functions yet to be found. CD36 is a double-spanning plasma membrane protein and is one of three members of the class B scavenger receptor family (46, 108). It is known as a receptor for thrombospondin (109), for a role in the translocation

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Figure 11 The Pococurante and Lackadaisical phenotypes. MyD88 is required for signaling via most of the TLRs, but not TLR3, and is only partly required for signaling via TLR4. Pococurante mimics a null allele except insofar as it permits full signaling by some of the ligands that use the TLR2/TLR6 and TLR2/TLRX complexes. Lackadaisical is similar to wild-type MyD88, except for diminished signaling via TLR7 and TLR9.

of fatty acids (110, 111), and for its ability to bind oxidized LDL (112). Some endogenous molecules with inﬂammatory effects may stimulate TLR2/TLR6 signaling by way of CD36, although speciﬁc examples of this have not yet been demonstrated.

PanR1: A Point Mutation in TNF on a Pure C57BL/6 Background The codominant PanR1 phenotype was so named for pan resistance to all TLR lig-

ands, where TNF activity was the endpoint of response. The mutation was mapped to the TNF locus itself, and PanR1 was found to be a missense allele (P138T) with dominant inhibitory effects, resulting from the fact that a single mutant subunit prevents engagement of the TNF trimer by the p55 TNF receptor (Figure 13). Little if any TNF activity is evident in homozygotes, which consequently show exaggerated susceptibility to Listeria monocytogenes (S. Rutschmann, K. Hoebe, J. Zalevsky, X. Du,

Figure 12 The Pococurante mutation (Poc) reveals that MyD88 can interact with TLR2 complexes in two conformationally distinct ways. Poc site and the BB loop site mutations inactivate most TLRs, and also inactivate MyD88, preventing interactions when engrafted onto either molecule. The TLR2/TLR6 heterodimer and the TLR2/TLRX heterodimer are unique in that they are capable of interacting with MyD88 in two ways: one that is disrupted by Poc or BB loop mutations in either molecule, and one that is not (special signaling mode). The latter mode of interaction is induced only by diacylated lipopeptides. www.annualreviews.org • Genetic Analysis of Host Resistance

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3d: A Defect of Signaling via TLR3, -7, and -9, Coupled with Defective Exogenous Antigen Presentation

Figure 13 The location of the PanR1 mutation, which affects a surface residue (P138T) on the TNF homotrimer (yellow). Subunits are each shown in different colors.

N. Mann, P. Steed, and B. Beutler, manuscript submitted). Mice with the mutation, unlike mice with a knockout allele of TNF, have entirely normal lymphoid development, including Peyer’s patches and marginal zone B cells in the spleen and lymph nodes (S. Rutschmann, K. Hoebe, J. Zalevsky, X. Du, N. Mann, P. Steed, and B. Beutler, manuscript submitted). This may suggest that the Tnf gene knockout affects expression of the neighboring lymphotoxin (LTa) gene, which is known to be required for lymphoid development, and that TNF itself has no developmental role. Alternatively, an exceedingly low level of TNF activity—present in the homozygous PanR1 mutants but not in homozygous knockout mice—may be sufﬁcient for lymphoid development.

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A combined defect of nucleic acid sensing via TLR3, -7, and -9 was designated Triple D (3d) to denote a triple defect of signaling (113). Probably because nucleic acid sensing via both TLR3 and TLR9 is impaired, the mice are highly susceptible to MCMV infection. However, they also show susceptibility to other organisms, such as Staphylococcus aureus, which is not effectively cleared and sometimes causes metastatic infections when inoculated intradermally. The 3d mutation abolishes exogenous antigen presentation, preventing both priming and cross-priming of T cells in response to ovalbumin, administered either as a particulate antigen (expressed by cells that were induced to undergo apoptosis) or as a soluble protein. However, normal levels of class I and class II MHC proteins are expressed on the surface of 3d APCs (113). The phenotype was shown to result from a missense error affecting the 12-spanning, ER-resident membrane protein UNC-93B, which had no previously recognized function in mammals. The effect of the mutation implies communication between the ER and the endosomal/lysosomal pathway. We speculate that 3d is required for the trafﬁcking of certain membrane proteins within the cell, including proteins required for signaling via TLR3, -7, and -9 and other proteins required for exogenous antigen presentation (Figure 14). A named homolog of UNC-93B (UNC-93A) is also encoded in the mouse genome, and still more distant homologs, as yet unnamed, are identiﬁable as well. These homologs are related to a Caenorhabditis elegans protein, known to be a component of a two-pore potassium channel (114). However, UNC-93B apparently does not fulﬁll this function in mammals (113).

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Figure 14 Possible mode of action of UNC-93B. The trafﬁc of speciﬁc membrane proteins required for signaling by TLR3, -7, -8, and -9 is postulated to depend on the ER-resident protein UNC-93B. These co-factors for TLR activity, and probably other proteins required for presentation and cross-presentation of exogenous antigens, cannot traverse the normal route from ER to endosome. Note that the TLR-independent cell death–induced activation pathway is also endocytic and, like the TLR pathway, leads to the induction of IFN-β. However, IFN-β induction does not depend on UNC-93B.

Feckless: A Protein Required for TLR3 to Activate NF-κB and for the Response to VSV As previously noted, TRIF activates IRF-3, yielding activation of the IFN-β gene, but also activates NF-κB via an interaction with TRAF-6 (97, 98). The Feckless phenotype was characterized by failure to activate NF-κB in response to dsRNA, although dsRNA could initiate IFN-β production (Z. Jiang & B. Beutler, unpublished data). Macrophages from Feckless mice were, moreover, found to be highly susceptible to VSV, and UV-inactivated VSV was incapable of stimulating a type I IFN response in Feckless cells. The G-glycoprotein of VSV is known to activate a type I IFN response via CD14, TLR4, TRAM, and IRF-7 (P. Georgel, Z. Jiang, S. Kunz, K. Hoebe, E. Janssen, M. Oldstone, and B. Beutler, manuscript submitted), but does not require TRIF to do so. Clearly required to carry a signal from TLR3 and TRIF to activate NF-κB, Feckless may also in-

teract with TRAM in an entirely independent pathway, leading to activation of IRF-7.

Spacey Spacey homozygotes are runted animals with low fertility and were identiﬁed because of diminished ability to respond to TLR9. The mutation has been mapped to chromosome 10 and is unlinked to the TLR9 locus itself. No genes known to be required for TLR signaling reside within the Spacey critical region. However, the mutation remains to be found.

Reverse genetics: the genetic approach that begins with a gene of unknown function and ends with a phenotype TBK1: TANK-binding kinase 1 IKKi: IκB kinase i

REVERSE GENETIC INVESTIGATIONS TBK1 and IKKi, IRF-7 and IRF-5 Intercurrent investigations using reverse genetic methods revealed that TBK1 and IKKi are the protein kinases responsible for activation of IRF-3 (98, 116–118), which is chieﬂy www.annualreviews.org • Genetic Analysis of Host Resistance

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responsible for induction of IFN-β gene expression caused by LPS or dsRNA. TBK1 appears to be the more important of the two (119). IRF-7, which exists at minimal concentrations in resting cells, is induced (presumably autoinduced) to high concentrations in response to TLR7 or TLR9 stimulation. These TLRs do not signal by way of TRIF or TRAM, and they activate type I IFN production via IRF-7 rather than IRF-3 (120). IRF-5 is required for MyD88-dependent signaling initiated by diverse MyD88-dependent TLRs (121). Investigators have suggested, although not formally proved, that IRF-5 is activated by IRAK-4. Alternatively, IRF-5 may be required for the transcription of genes encoding certain components of the pathway itself.

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Inhibitors of Signaling IRAK-M (122) and SOCS (suppressor of cytokine signaling)-1 (123) function as inhibitors of signaling via TLRs, the former probably inhibiting IRAK/IRAK-2/IRAK-4 signaling and the latter preventing STAT (signal transducer and activator of transcription)1 signaling. The orphan TIR domain receptors SIGIRR (single Ig repeat–related protein) (124) and ST2 (125) exert inhibitory inﬂuences as well, probably by virtue of direct interactions with TLRs. Upregulation of the phosphatase SHIP (Src homology inducible phosphatase)-1, which depends on autocrine stimulation by TGF-β, also inhibits LPS signal transduction, possibly blocking both MyD88-dependent and TRIFdependent pathways (126). The phenomenon of endotoxin tolerance is thus partially explained (127) and appears to operate at several different levels.

Saturation: How Many Genes Serve the Pathways? We can now point to 30 genes that make positive (as opposed to inhibitory) contributions to TLR signal transduction. As outlined above, roughly 40 to 50 genes are expected 380

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to make nonredundant contributions to signaling, subject to fairly broad limits of error. These ﬁgures might be taken to indicate that 10 to 20 genes remain to be found. We should emphasize that these ﬁgures include only those genes that are required for the production of TNF activity in response to ligands that depend on 7 of the 12 mouse TLRs. If the screen for responses were less focused, and if all the TLRs were stimulated, more targets might be detected.

A BROADER LOOK: THE MCMV RESISTOME AND ITS SIZE Because 11 transmissible mutations were identiﬁed by screening about 20,000 G3 mice, an authentic phenotype is picked up in about 1 of every 1800 mice examined. A substantially higher hit rate is observed in a screen that tests the integrity of all essential systems for the containment of MCMV infection. Approximately 1 in 400 G3 mice score positive in this screen, in which 105 PFU of virus (an inoculum insufﬁcient to affect normal C57BL/6 animals) is administered by an intraperitoneal route, and sickness or death is recorded within the ﬁrst seven days to assure that only innate immunodeﬁciency phenotypes are detected. This corresponds to 1 recessive mutation in every 33 pedigrees examined, and to a genomic footprint that encompasses about 45,000 nucleotides, parceled among about 300 genes. The innate resistome for MCMV thus corresponds to about 1% of the genome. Some of the mutations identiﬁed in a screen for MCMV susceptibility cause cosusceptibility to other viral pathogens. For example, macrophage susceptibility to VSV is commonly observed. But some mutations have a speciﬁc effect on NK cell function, or rather broad effects on vesicle transport, leading to hypopigmentation coupled with immunodeﬁciency. As of this writing, approximately 30 MCMV mutations have been identiﬁed in screening, and 10 mutations are displayed in

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Table 2

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Nine MCMV susceptibility mutations and their phenotypic characteristics

Name

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Outcome of MCMV infection

VSV Susceptible?a

IFN-α/β Rescue?b

NK cell defect?

Cloned?

Domino (recessive)

death on day 4 Necrotic spleen

Yes

No

Yes

Yes (STAT1)

Goodnight (recessive)

death on day 2–3 Necrotic liver

n.d.

n.a.

n.d.

No

Solitaire (recessive)

death on day 2–3

n.d.

n.a.

n.d.

No

Slumber (recessive)

death on day 2–3

n.d.

n.a.

n.d.

No

Jinx (recessive)

Elevated viral load in spleen

Yes

Yes

Yes

No

Warmﬂash (co-dominant)

Elevated viral load in spleen

Yes

Yes

No

No

May Day (recessive)

death on day 3

n.d.

n.a.

n.d.

No

Paris (recessive)

Exanthem

n.d.

n.a.

n.d.

No

Moneypenny (recessive)

death on day 6

Yes

Yes

n.d.

No

Havelock (recessive)

Elevated viral load in spleen

No

n.a.

No

No

a

Refers to the susceptibility of macrophages ex vivo. Refers to rescue of the VSV phenotype. n.a., not applicable; n.d., not determined.

b

Table 2. One among these has been positionally cloned. This mutation, Domino, is a missense allele of STAT-1, modifying the DNA-binding domain of the protein and also preventing normal phosphorylation and expression of the protein in cells (K. Crozat, P. Georgel, S. Rutschmann, N. Mann, X. Du, K. Hoebe, and B. Beutler, manuscript submitted). Remarkably, many of the mutations identiﬁed to date cause far greater susceptibility to MCMV than the Domino mutation, which effectively disrupts both type I and type II IFN signaling, as well as IL-27 signaling. We must therefore conclude that some proteins are far more important to the host than the IFNs, given the circumstance of MCMV infection. Clearly, much remains to be learned about antiviral defense.

CONCLUSIONS A classical genetic inquiry led directly to our present understanding of how the immune system detects infection. But beyond pointing to the function of the TLRs and helping

to elucidate their signaling pathways, the genetic approach has done much more as well. Intracellular sensors of microbes also exist. The nucleotide-binding oligomerization domain (NOD)-1 and NOD2 proteins, for example, are believed to function as cytoplasmic sensors of peptidoglycans (128–130). Their general role in immunity was originally discovered by a pure genetic investigation, undertaken in humans rather than in mice (131, 132). The NK receptor Ly49H, which detects the MCMV-encoded protein m157 (133), was identiﬁed by positional cloning as well (2, 3). So, too, was the key transcription factor required for the development of regulatory T cells (134). Mutagenesis is being directly applied to the problem of autoimmunity and has already yielded impressive results (135, 136). What other large-scale phenomena interest us in immunology? This is for the reader to ponder. Sometimes, asking the question—or in this case, seeing the phenomenon, cleverly concealed as it is—may be the most difﬁcult part of the process. www.annualreviews.org • Genetic Analysis of Host Resistance

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APPENDIX: GLOSSARY OF MUTATIONS Domino (Dom): STAT-1 (missense; null) Feckless (fks): unknown (failure to activate NF-κB with preserved IFN production in response to polyI:C) Heedless (hdl): CD14 (nonsense; null) Insouciant (Int): TLR6 (missense; null) Lackadaisical (Lkd): MyD88 (missense; diminished signaling via TLRs 7 and 9) Lps: TLR4 (missense; null) Annu. Rev. Immunol. 2006.24:353-389. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

Lps2: TRIF (frameshift; null; codominant or haploinsufﬁcient) Oblivious (Obl): CD36 (nonsense; null) Pococurante (Poc): MyD88 (missense; retains signaling only via TLR2/TLR6 and TLR2/TLRX) PanR1: TNF (missense; extreme hypomorph; dominant) Triple D (3d): unc-93b (missense; null) Spacey (Spy): unknown (diminished TLR9 signaling)

LITERATURE CITED 1. This classic study of the heritability of premature death due to many causes, including infection, formally established that premature death due to infection is more heritable than premature death of any other cause.

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1. Sorensen TI, Nielsen GG, Andersen PK, Teasdale TW. 1988. Genetic and environmental influences on premature death in adult adoptees. N. Engl. J. Med. 318:727–32 2. Lee SH, Girard S, Macina D, Busa M, Zafer A, et al. 2001. Susceptibility to mouse cytomegalovirus is associated with deletion of an activating natural killer cell receptor of the C-type lectin superfamily. Nat. Genet. 28:42–45 3. Brown MG, Dokun AO, Heusel JW, Smith HR, Beckman DL, et al. 2001. Vital involvement of a natural killer cell activation receptor in resistance to viral infection. Science 292:934–37 4. Desrosiers MP, Kielczewska A, Loredo-Osti JC, Adam SG, Makrigiannis AP, et al. 2005. Epistasis between mouse Klra and major histocompatibility complex class I loci is associated with a new mechanism of natural killer cell-mediated innate resistance to cytomegalovirus infection. Nat. Genet. 37:593–99 5. Lee SH, Zafer A, De Repentigny Y, Kothary R, Tremblay ML, et al. 2003. Transgenic expression of the activating natural killer receptor Ly49H confers resistance to cytomegalovirus in genetically susceptible mice. J. Exp. Med. 197:515–26 6. Samson M, Libert F, Doranz BJ, Rucker J, Liesnard C, et al. 1996. Resistance to HIV1 infection in caucasian individuals bearing mutant alleles of the CCR-5 chemokine receptor gene. Nature 382:722–25 7. Martinson JJ, Chapman NH, Rees DC, Liu YT, Clegg JB. 1997. Global distribution of the CCR5 gene 32-basepair deletion. Nat. Genet. 16:100–3 8. Pancer Z, Amemiya CT, Ehrhardt GR, Ceitlin J, Gartland GL, Cooper MD. 2004. Somatic diversiﬁcation of variable lymphocyte receptors in the agnathan sea lamprey. Nature 430:174–80 Beutler et al.

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9. Beutler B, Crozat K, Koziol JA, Georgel P. 2005. Genetic dissection of innate immunity to infection: the mouse cytomegalovirus model. Curr. Opin. Immunol. 17:36–43 10. Lemaitre B, Nicolas E, Michaut L, Reichhart JM, Hoffmann JA. 1996. The dorsoventral regulatory gene cassette spatzle/Toll/cactus controls the potent antifungal response in Drosophila adults. Cell 86:973–83 11. Anderson KV, Bokla L, Nusslein-Volhard C. 1985. Establishment of dorsal-ventral polarity in the Drosophila embryo: the induction of polarity by the Toll gene product. Cell 42:791–98 12. Beutler B. 2004. Innate immunity: an overview. Mol. Immunol. 40:845–59 13. Presti RM, Pollock JL, Dal Canto AJ, O’Guin AK, Virgin HW. 1998. Interferon γ regulates acute and latent murine cytomegalovirus infection and chronic disease of the great vessels. J. Exp. Med. 188:577–88 14. Leadbetter EA, Rifkin IR, Hohlbaum AM, Beaudette BC, Shlomchik MJ, MarshakRothstein A. 2002. Chromatin-IgG complexes activate B cells by dual engagement of IgM and Toll-like receptors. Nature 416:603–7 15. Beutler B. 2004. Inferences, questions and possibilities in Toll-like receptor signalling. Nature 430:257–63 16. Luderitz O, Galanos C, Lehmann V, Nurminen M, Rietschel ET, et al. 1973. Lipid A: chemical structure and biological activity. J. Infect. Dis. 128:29 17. Berczi I, Bertok L, Bereznai T. 1966. Comparative studies on the toxicity of Escherichia coli lipopolysaccharide endotoxin in various animal species. Can. J. Microbiol. 12:1070– 71 18. Michalek SM, Moore RN, McGhee JR, Rosenstreich DL, Mergenhagen SE. 1980. The primary role of lymphoreticular cells in the mediation of host responses to bacterial endotoxin. J. Infec. Dis. 141:55–63 19. Beutler B, Milsark IW, Cerami A. 1985. Passive immunization against cachectin/tumor necrosis factor (TNF) protects mice from the lethal effect of endotoxin. Science 229:869– 71 20. Karaghiosoff M, Steinborn R, Kovarik P, Kriegshauser G, Baccarini M, et al. 2003. Central role for type I interferons and Tyk2 in lipopolysaccharide-induced endotoxin shock. Nat. Immunol. 4:471–77 21. Heppner G, Weiss DW. 1965. High susceptibility of strain A mice to endotoxin and endotoxin-red blood cell mixtures. J. Bacteriol. 90:696–703 22. Watson J, Riblet R. 1974. Genetic control of responses to bacterial lipopolysaccharides in mice. I. Evidence for a single gene that inﬂuences mitogenic and immunogenic respones to lipopolysaccharides. J. Exp. Med. 140:1147–61 23. Watson J, Riblet R. 1975. Genetic control of responses to bacterial lipopolysaccharides in mice. II. A gene that inﬂuences a membrane component involved in the activation of bone marrow-derived lymphocytes by lipipolysaccharides. J. Immunol. 114:1462–68 24. Watson J, Riblet R, Taylor BA. 1977. The response of recombinant inbred strains of mice to bacterial lipopolysaccharides. J. Immunol. 118:2088–93 25. Skidmore BJ, Chiller JM, Weigle WO, Riblet R, Watson J. 1976. Immunologic properties of bacterial lipopolysaccharide (LPS). III. Genetic linkage between the in vitro mitogenic and in vivo adjuvant properties of LPS. J. Exp. Med. 143:143–50 26. Condie RM, Zak SJ, Good RA. 1955. Effect of meningococcal endotoxin on the immune response. Proc. Soc. Exp. Biol. Med. 90:355–60 27. Coutinho A, Meo T. 1978. Genetic basis for unresponsiveness to lipopolysaccharide in C57BL/10Cr mice. Immunogenetics 7:17–24 www.annualreviews.org • Genetic Analysis of Host Resistance

10. Demonstration that the Toll protein of Drosophila, and upstream and downstream signaling components, are essential for activation of Drosomycin synthesis.

21. The first demonstration that mice of the C3H/HeJ substrain are highly resistant to the lethal effect of LPS.

383

Annu. Rev. Immunol. 2006.24:353-389. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-12

ARI

15 February 2006

33. This work first revealed that mammalian TLRs act as specific sensors of microbial molecules and are required for recognition and response to infection.

384

4:15

28. O’Brien AD, Rosenstreich DL, Scher I, Campbell GH, MacDermott RP, Formal SB. 1980. Genetic control of susceptibility to Salmonella typhimurium in mice: role of the LPS gene. J. Immunol. 124:20–4 29. Hagberg L, Hull R, Hull S, McGhee JR, Michalek SM, Svanborg Eden C. 1984. Difference in susceptibility to Gram-negative urinary tract infection between C3H/HeJ and C3H/HeN mice. Infect. Immun. 46:839–44 30. Woods JP, Frelinger JA, Warrack G, Cannon JG. 1988. Mouse genetic locus Lps inﬂuences susceptibility to Neisseria meningitidis infection. Infect. Immun. 56:1950–55 31. Macela A, Stulik J, Hernychova L, Kroca M, Krocova Z, Kovarova H. 1996. The immune response against Francisella tularensis live vaccine strain in Lpsn and Lpsd mice. FEMS Immunol. Med. Microbiol. 13:235–38 32. Shakhov AN, Collart MA, Vassalli P, Nedospasov SA, Jongeneel CV. 1990. κB-type enhancers are involved in lipopolysaccharide-mediated transcriptional activation of the tumor necrosis factor α gene in primary macrophages. J. Exp. Med. 171:35–47 33. Poltorak A, He X, Smirnova I, Liu M-Y, Van Huffel C, et al. 1998. Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene. Science 282:2085–88 34. Nomura N, Miyajima N, Sazuka T, Tanaka A, Kawarabayasi Y, et al. 1994. Prediction of the coding sequences of unidentiﬁed human genes. I. The coding sequences of 40 new genes (KIAA0001-KIAA0040) deduced by analysis of randomly sampled cDNA clones from human immature myeloid cell line KG-1. DNA Res. 1:27–35 35. Taguchi T, Mitcham JL, Dower SK, Sims JE, Testa JR. 1996. Chromosomal localization of TIL, a gene encoding a protein related to the Drosophila transmembrane receptor Toll, to human chromosome 4p14. Genome 32:486–88 36. Medzhitov R, Preston-Hurlburt P, Janeway CA Jr. 1997. A human homologue of the Drosophila Toll protein signals activation of adaptive immunity. Nature 388:394–97 37. Rock FL, Hardiman G, Timans JC, Kastelein RA, Bazan JF. 1998. A family of human receptors structurally related to Drosophila Toll. Proc. Natl. Acad. Sci. USA 95:588– 93 38. Rutschmann S, Kilinc A, Ferrandon D. 2002. Cutting edge: the Toll pathway is required for resistance to Gram-positive bacterial infections in Drosophila. J. Immunol. 168:1542– 46 39. Chaturvedi MM, Higuchi M, Aggarwal BB. 1994. Effect of tumor necrosis factors, interferons, interleukins, and growth factors on the activation of NF-κB: evidence for lack of correlation with cell proliferation. Lymphokine Cytokine Res. 13:309–13 40. Matsumoto S, Tsuji-Takayama K, Aizawa Y, Koide K, Takeuchi M, et al. 1997. Interleukin-18 activates NF-κB in murine T helper type 1 cells. Biochem. Biophys. Res. Commun. 234:454–57 41. Robinson D, Shibuya K, Mui A, Zonin F, Murphy E, et al. 1997. IGIF does not drive Th1 development but synergizes with IL-12 for interferon-γ production and activates IRAK and NFκB. Immunity 7:571–81 42. Choe J, Kelker MS, Wilson IA. 2005. Crystal structure of human Toll-like receptor 3 (TLR3) ectodomain. Science 309:581–85 43. Xu Y, Tao X, Shen B, Horng T, Medzhitov R, et al. 2000. Structural basis for signal transduction by the Toll/interleukin-1 receptor domains. Nature 408:111–15 44. Dunne A, Ejdeback M, Ludidi PL, O’Neill LA, Gay NJ. 2003. Structural complementarity of Toll/interleukin-1 receptor identity regions in Toll-like receptors and the adaptors Mal and MyD88. J. Biol. Chem. 278:41443–51 Beutler et al.

Annu. Rev. Immunol. 2006.24:353-389. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-12

ARI

15 February 2006

4:15

45. Schneider DS, Hudson KL, Lin TY, Anderson KV. 1991. Dominant and recessive mutations deﬁne functional domains of Toll, a transmembrane protein required for dorsalventral polarity in the Drosophila embryo. Genes Dev. 5:797–807 46. Hoebe K, Georgel P, Rutschmann S, Du X, Mudd S, et al. 2005. CD36 is a sensor of diacylglycerides. Nature 433:523–27 47. Wright SD, Ramos RA, Tobias PS, Ulevitch RJ, Mathison JC. 1990. CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. Science 249:1431–33 48. Nagai Y, Akashi S, Nagafuku M, Ogata M, Iwakura Y, et al. 2002. Essential role of MD-2 in LPS responsiveness and TLR4 distribution. Nat. Immunol. 3:667–72 49. Takeuchi O, Sato S, Horiuchi T, Hoshino K, Takeda K, et al. 2002. Cutting edge: role of Toll-like receptor 1 in mediating immune response to microbial lipoproteins. J. Immunol. 169:10–14 50. Takeuchi O, Kawai T, Muhlradt PF, Morr M, Radolf JD, et al. 2001. Discrimination of bacterial lipoproteins by Toll-like receptor 6. Int. Immunol. 13:933–40 51. Buwitt-Beckmann U, Heine H, Wiesmuller KH, Jung G, Brock R, et al. 2005. Toll-like receptor 6-independent signaling by diacylated lipopeptides. Eur. J. Immunol. 35:282– 89 52. Hayashi F, Smith KD, Ozinsky A, Hawn TR, Yi EC, et al. 2001. The innate immune response to bacterial ﬂagellin is mediated by Toll-like receptor 5. Nature 410:1099–103 53. Hemmi H, Kaisho T, Takeuchi O, Sato S, Sanjo H, et al. 2002. Small anti-viral compounds activate immune cells via the TLR7 MyD88-dependent signaling pathway. Nat. Immunol. 3:196–200 54. Heil F, Hemmi H, Hochrein H, Ampenberger F, Kirschning C, et al. 2004. Speciesspeciﬁc recognition of single-stranded RNA via Toll-like receptor 7 and 8. Science 303:1526–29 55. Diebold SS, Kaisho T, Hemmi H, Akira S, Reis e Sousa C. 2004. Innate antiviral responses by means of TLR7-mediated recognition of single-stranded RNA. Science 303:1529–31 56. Lund JM, Alexopoulou L, Sato A, Karow M, Adams NC, et al. 2004. Recognition of single-stranded RNA viruses by Toll-like receptor 7. Proc. Natl. Acad. Sci. USA 101:5598– 603 57. Hemmi H, Takeuchi O, Kawai T, Kaisho T, Sato S, et al. 2000. A Toll-like receptor recognizes bacterial DNA. Nature 408:740–45 58. Bauer S, Kirschning CJ, Hacker H, Redecke V, Hausmann S, et al. 2001. Human TLR9 confers responsiveness to bacterial DNA via species-speciﬁc CpG motif recognition. Proc. Natl. Acad. Sci. USA 98:9237–42 59. Tabeta K, Georgel P, Janssen E, Du X, Hoebe K, et al. 2004. Toll-like receptors 9 and 3 as essential components of innate immune defense against mouse cytomegalovirus infection 3. Proc. Natl. Acad. Sci. USA 101:3516–21 60. Zhang D, Zhang G, Hayden MS, Greenblatt MB, Bussey C, et al. 2004. A Toll-like receptor that prevents infection by uropathogenic bacteria. Science 303:1522–26 61. Yarovinsky F, Zhang D, Andersen JF, Bannenberg GL, Serhan CN, et al. 2005. TLR11 activation of dendritic cells by a protozoan proﬁlin-like protein. Science 308:1626–29 62. Takeuchi O, Hoshino K, Akira S. 2000. Cutting edge: TLR2-deﬁcient and MyD88deﬁcient mice are highly susceptible to staphylococcus aureus infection. J. Immunol. 165:5392–96 63. Krug A, French AR, Barchet W, Fischer JA, Dzionek A, et al. 2004. TLR9-dependent recognition of MCMV by IPC and DC generates coordinated cytokine responses that activate antiviral NK cell function. Immunity 21:107–19 www.annualreviews.org • Genetic Analysis of Host Resistance

46. The forward genetic approach revealed an essential function of CD36, showing that it acts as a coreceptor for specific ligands of the TLR2/TLR6 complex.

385

Annu. Rev. Immunol. 2006.24:353-389. Downloaded from arjournals.annualreviews.org by HINARI on 08/26/07. For personal use only.

ANRV270-IY24-12

ARI

15 February 2006

67. An ENU-induced mutation (Lps2) was the first to reveal the function of the adapter protein TRIF, later confirmed by gene knockout.

79. The long-known adjuvant effect of LPS, biochemically defined by the upregulation of costimulatory molecules, was ascribed to the MyD88independent pathway.

386

4:15

64. Kurt-Jones EA, Popova L, Kwinn L, Haynes LM, Jones LP, et al. 2000. Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus. Nat. Immunol. 1:398–401 65. Rassa JC, Meyers JL, Zhang Y, Kudaravalli R, Ross SR. 2002. Murine retroviruses activate B cells via interaction with Toll-like receptor 4. Proc. Natl. Acad. Sci. USA 99:2281– 86 66. Jude BA, Pobezinskaya Y, Bishop J, Parke S, Medzhitov RM, et al. 2003. Subversion of the innate immune system by a retrovirus. Nat. Immunol. 4:573–78 67. Hoebe K, Du X, Georgel P, Janssen E, Tabeta K, et al. 2003. Identification of Lps2 as a key transducer of MyD88-independent TIR signaling. Nature 424:743–48 68. Lund J, Sato A, Akira S, Medzhitov R, Iwasaki A. 2003. Toll-like receptor 9-mediated recognition of herpes simplex virus-2 by plasmacytoid dendritic cells. J. Exp. Med. 198:513–20 69. Crozat K, Beutler B. 2004. TLR7: A new sensor of viral infection. Proc. Natl. Acad. Sci. USA 101:6835–36 70. Ehl S, Bischoff R, Ostler T, Vallbracht S, Schulte-Monting J, et al. 2004. The role of Toll-like receptor 4 versus interleukin-12 in immunity to respiratory syncytial virus. Eur. J. Immunol. 34:1146–53 71. Youmans AS, Youmans GP. 1969. Factors affecting immunogenic activity of mycobacterial ribosomal and ribonucleic acid preparations. J. Bacteriol. 99:42–50 72. Youmans GP, Youmans AS. 1969. Allergenicity of mycobacterial ribosomal and ribonucleic acid preparations in mice and guinea pigs. J. Bacteriol. 97:134–39 73. Schmidtke JR, Johnson AG. 1971. Regulation of the immune system by synthetic polynucleotides. I. Characteristics of adjuvant action on antibody synthesis. J. Immunol. 106:1191–200 74. Capanna SL, Kong YM. 1974. Further studies on the prevention of tolerance induction by poly A:U. Immunology 27:647–53 75. Nauciel C, Fleck J, Martin JP, Mock M, Nguyen-Huy H. 1974. Adjuvant activity of bacterial peptidoglycans on the production of delayed hypersensitivity and on antibody response. Eur. J. Immunol. 4:352–56 76. Bultmann B, Finger H, Heymer B, Schachenmayr W, Hof H, Haferkamp O. 1975. Adjuvancy of streptococcal nucleic acids Z. Immunitatsforsch. Exp. Klin. Immunol. 148:425– 30 77. Borriello F, Sethna MP, Boyd SD, Schweitzer AN, Tivol EA, et al. 1997. B7–1 and B7–2 have overlapping, critical roles in immunoglo