Scientific Report 2005

Immunology

Chairman’s Overview

The faculty of the Department of Immunology continues its long-standing commitments to the highest level of scientific achievement, the training of graduate and postgraduate students, and participation in numerous activities outside Scripps Research. But first an update on the demographics of the department. The department consists of 46 full-time faculty members; 37 of the members are less than 60 years old, and the majority are 50 years old or less. This age distribution helps ensure the long-time vitality and success of the department and is important to the overall excellence of Scripps Research. The scientific successes of our younger faculty members are well recognized as indicated by the receipt of highly competitive career development awards such as the Burroughs Wellcome Career Award in Biomedical Science received by Erica Ollmann Saphire and the highly coveted Established Investigator Award awarded to David Schlaepfer by the American Heart Association. Although a number of faculty members have retired, most recently Charles Cochrane, and more are planning to retire soon, an active recruitment program since 1996 has added 9 new faculty members.

Another interesting statistic is that 18% of the faculty are women, all of whom are laboratory heads and most of whom are tenured. In a recent letter in Nature Immunology, the importance of Immunology as a career for young female scientists was highlighted. Clearly, the Department of Immunology far exceeds all averages quoted in the letter. We are proud of our record.

Outside honors and appointments to review committees and top editorial boards accrue to all of the faculty. One important activity is serving on National Institutes of Health (NIH) review committees, and members of our department are well represented. Serving as a permanent member on an NIH study section is a major time commitment; currently 5 of our faculty serve in this capacity: Drs. Bokoch, Curtiss, Gascoigne, Kono, and Nemazee. Their service is critical to maintaining the integrity of the NIH review process, and their willingness to do so is appreciated by all of the members of the department.

Large-scale research grants have become a major focus at NIH. Consequently, it is critical that scientists in the department be able to respond to these new funding mechanisms. Today the department has several such grants. The most recent is a multimillion dollar grant to establish the Scripps Research Institute Molecular Screening Center. The goal of this pilot program is to discover small-molecule tools for translating basic biomedical discoveries more quickly into medically relevant applications. This grant also marks a key collaboration between Scripps Research in La Jolla and Scripps Florida. This funding is part of the NIH strategic funding plan, the Roadmap Initiative. Hugh Rosen is the principal investigator on the grant. His intention is to conduct high-throughput screens against various biological targets to uncover “proof of concept” molecules useful in studying human health and in developing new treatments for human diseases. As Dr. Rosen remarked, “With this grant, the NIH has recognized the unique capabilities of our established researchers in La Jolla with our newest investigators and equipment in Palm Beach County.”

During the past year, the output of seminal advances was high. Members of the department were authors of more than 30 articles that appeared in high-profile journals, including Science, Nature, Cell, Nature Immunology, Nature Cell Biology, and Nature Medicine. Most of the research described in these articles is given in detail in the reports of the individual laboratories, but the following are a few of the highlights

This department, long recognized for its continued contributions to studies of the adaptive immune system, is now recognized as one of the leaders in advancing our understanding of innate immunity. One reason for this recognition is the continued productivity of Bruce Beutler and his group. During the past year, they made several important advances. In an article published in Nature, they describe a key role for the transmembrane protein known as CD36 in sensing diacyglycerides. The researchers showed that a nonsense mutation of Cd36 causes a recessive immunodeficiency in which macrophages are insensitive to the R-enantiomer of MALP-2 (a diacylated bacterial lipopeptide) and to lipoteichoic acid. Mice homozygous for the mutation are hypersusceptible to Staphylococcus aureus infection. Studies on macrophages from mice with the mutation revealed that some, but not all, ligands for Toll-like receptor (TLR) 2 are CD36 dependent. Already known as a receptor for endogenous molecules, CD36 is also a selective and nonredundant sensor of microbial diacylglycerides that signal via the TLR2/6 heterodimer. This work provides new data on the role of innate immunity during infection and during chronic inflammatory diseases such as atherosclerosis in which endogenous diacyglycerides may be generated and trigger TLR2.

A second report by Dr. Beutler and colleagues published in Nature Immunology provides new information on CD14 and its role in TLR4 signaling. This research is particularly interesting because it was nearly 15 years ago that the studies of other members of the department published in Science provided the first molecular insights into the function of CD14 as a sensor for bacterial lipopolysaccharides.

Gary Bokoch and the members of his laboratory continue to provide new insights into the molecular physiology of phagocytic cells. During the past year, in an article in Nature Cell Biology, they reported the biochemical isolation of chronophin, a unique cofilin-activating phosphatase of the haloacid dehalogenase superfamily. Chronophin directly dephosphorylates cofilin with high specificity and colocalizes with cofilin in motile and dividing cells. Loss of chronophin activity blocks phosphocycling of cofilin, stabilizes F-actin structures, and causes massive defects in cell division. These findings identify a physiologic phosphoserine protein substrate for a mammalian haloacid dehalogenase–type phosphatase and indicate that chronophin is an important novel regulator of cofilin-mediated actin reorganization.

Researchers in Jiahuai Han’s group recently made a major contribution to understanding the mechanisms involved in stability of mRNA species that encode key molecules in innate immunity and inflammation. Adenosine-uridine–rich elements (AREs) in the 3´ untranslated region of unstable mRNAs dictate degradation of the mRNAs. An RNA interference–based screen in Drosophila S2 cells revealed that in TNF-α, components involved in microRNA processing and function are required for the rapid decay of mRNA that contains AREs. The requirement for the component Dicer in the instability of mRNA with AREs was confirmed in HeLa cells. The researchers further observed that miR16, a human microRNA containing a sequence complementary to the ARE sequence is required for the turnover of RNA that contains AREs. The role of miR16 in the decay of mRNA that contains AREs is sequence specific and requires the ARE-binding protein tristetraprolin. Tristetraprolin does not directly bind to miR16; rather it interacts through association with other components involved in microRNA processing to form a complex with miR16 and assists in the targeting of ARE. The targeting of ARE by microRNA therefore appears to be an essential step in ARE-mediated mRNA degradation. These findings provide an entirely new understanding of the function of microRNA and a new model for studies of AREs during a variety of host responses.

Other publications by members of the department include reports of the pioneering research of Luc Teyton and his group, in collaboration with A. Bendelac, University of Chicago, on natural killer T cells and the cells’ key membrane sensor CD1. Specifically, in articles published in Science, Nature, and Nature Immunology, the following was noted. Natural killer T cells are a distinct lineage of T cells that coexpress a conserved αβ T-cell receptor (TCR) and natural killer receptors. Although the TCR of natural killer T cells is characteristically autoreactive to CD1d, a lipid-presenting molecule, endogenous ligands for these cells have not been identified. Dr. Teyton and his group showed that isoglobotrihexosylceramide (iGb3), a lysosomal glycosphingolipid of previously unknown function, is recognized by both mouse and human natural killer T cells. Impaired generation of lysosomal iGb3 in mice lacking β-hexosaminidase B results in a severe deficiency in natural killer
T cells, suggesting that this lipid also mediates development of these T cells in mice. The findings suggest that expression of iGb3 in peripheral tissues may be involved in controlling the responses of natural killer cells to infections and malignant neoplasms and in autoimmunity.

Further, the scientists have shown microbial, antigen-specific activation of natural killer T cells against gram-negative, lipopolysaccharide-negative α-Proteobacteria such as Ehrlichia muris and Sphingomonas capsulata. Glycosylceramides from the cell wall of Sphingomonas act as direct targets for mouse and human natural killer T cells, controlling both septic shock and bacterial clearance in infected mice. In contrast, gram-negative, lipopolysaccharide-positive Salmonella typhimurium activates natural killer T cells through recognition of iGb3 presented by lipopolysaccharide-activated dendritic cells. These findings identify 2 novel antigenic targets of natural killer T cells in antimicrobial defense and show that glycosylceramides are an alternative to lipopolysaccharide for innate recognition of the gram-negative, lipopolysaccharide-negative bacterial cell wall.

Finally, in collaborative studies with Ian Wilson and his group, members of the Teyton group documented the crystal structure of CD1d in complex with a short-chain synthetic variant of α-galactosylceramide at a resolution of 2.2 Å. This structure indicates the basis for the high specificity of these microbial ligands and explains the restriction of the α-linkage as a unique pathogen-specific pattern recognition motif. Comparison of the binding of altered lipid ligands to CD1d and TCRs suggested that the differential helper T cell–like properties of natural killer T cells may originate largely from differences in the “loading” of the ligands in different cell types and hence in the tissue distribution of the ligands in vivo.

Overall, this research provides a remarkably broad set of advances in understanding the functions of natural killer T cells and the structure of one of the key sensor molecules. In addition, it further illustrates the ability of members of the department to collaborate with other leaders in this field inside and outside of Scripps Research.

Members of the Department of Immunology have a long-standing interest in the functions of T cells in various biological models. Studies from Wendy Havran and her group are an example of this interest. A fine balance between rates of proliferation and apoptosis in the skin provides a defensive barrier and a mechanism for tissue repair after damage. Vγ3⁺ dendritic epidermal T cells are primary modulators of skin immune responses. Dr. Havran and her group showed that these cells both produce and respond to insulin-like growth factor 1 (IGF-1) after TCR stimulation. Mice deficient in the cells had a notable increase in epidermal apoptosis that was abrogated by the addition of Vγ3⁺ dendritic epidermal T cells or IGF-1. Furthermore, mice deficient in the cells had reduced activation of IGF-1 receptors at wound sites. These findings indicate critical functions for Vγ3⁺ dendritic epidermal T cell–mediated IGF-1 production in regulating skin homeostasis and repair.

Linda Sherman and members of her laboratory also provided important new insights into the role of T cells in tumor immunity. Studies published in Immunity, done in collaboration with M. Theobald, Johannes Gutenberg University, Mainz, Germany, revealed that efficient immune attack on malignant disease requires the concerted action of both CD8⁺ cytotoxic T lymphocytes (CTLs) and CD4⁺ T helper cells. The researchers used HLA-A*0201 (A2.1) transgenic mice, in which the mouse CD8 molecule cannot efficiently interact with the α3 domain of A2.1, to generate a high-affinity, CD8-independent TCR specific for a commonly expressed, tumor-associated CTL epitope derived from p53, a human tumor suppressor protein. Introduction of this TCR into human T cells resulted in CD8⁺ T lymphocytes with broad tumor-specific cytotoxic activity and CD4⁺ T cells with potent tumor-reactive, p53A2.1-specific helper activity. Both T-cell subsets interacted synergistically with dendritic cell intermediates and tumor targets. The intentional redirection of both CD4⁺ helper T cells and CD8⁺ CTLs by the same high-affinity, CD8-independent, tumor-specific TCRs could provide the basis for novel broad-spectrum cancer immunotherapeutic agents.

Other contributions of note during the past year include studies done by David Nemazee and members of his laboratory on B-cell development. In developing B cells, expression of immunoglobulin on the cell surface is an important signal to terminate expression of recombinase activator gene (RAG) and V(D)J recombination. However, autoreactive antigen receptors promote continued gene rearrangement and receptor editing. Regulation of RAG expression and editing by B-cell receptor signaling is poorly understood. Dr. Nemazee and his colleagues found that in editing-competent cells, RAG mRNA expression induced by B-cell receptor ligands is regulated at the level of RAG transcription. In immature B cells carrying innocuous receptors, RAG expression appears to be under rapidly reversible negative regulation. Research with transduction of a superrepressive IκBα protein indicated that NF-κB/Rel proteins promote RAG transcription. Interestingly, cells deficient in NF-κB overexpress RAG and undergo an exaggerated receptor editing response. These results implicate NF-κB transcription factors in the regulation of RAG transcription mediated by B-cell receptors. Rapidly activated NF-κB pathways may facilitate prompt antigen receptor–regulated changes in RAG expression important for editing and haplotype exclusion.

In closing, I can say without reservation that writing this report was a great pleasure, because it gave me a chance to review the accomplishments of the faculty of the Department of Immunology. These accomplishments span all aspects of 21st century science: cutting-edge contributions to knowledge of the immune system; education of undergraduate, graduate, and postgraduate students; and participation in important outside activities that include serving on NIH review committees and journal editorial boards and organizing and participating in important national and international scientific meetings. Each member of the faculty contributes in all these aspects of science and makes the Department of Immunology, to me, the model that many others aspire to emulate.