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Immunology and Cell Biology of Glycolipid Antigens
Summary: Albert Bendelac studies the cell biology and immunology of lipid antigens. His group focuses on the interactions between NKT cells, a conserved lineage of glycolipid-specific lymphocytes, and CD1, a family of major histocompatibility (MHC)-like molecules that bind various self and microbial lipids in the endosomal compartment for presentation at the plasma membrane.
Lipids and glycolipids perform specialized functions in a wide variety of biological processes. Fundamental aspects of their trafficking between subcellular compartments or recruitment into membrane microdomains, their interactions with protein receptors, their signaling properties, and other functions remain largely mysterious. In immune responses, where the detection of glycolipids of both self and microbial origin is an essential component of both innate and adaptive immunity, it is particularly important to understand these processes. The immune system also provides a unique model to study some of these basic unresolved questions.
Unlike microbial proteins, which mutate at high rate, lipids and glycolipids are relatively fixed targets for the immune system. Conserved families of membrane receptors have evolved to sense them. Some members of the Toll-like receptor (TLR) family, which is found from plants to insects and humans, directly activate key components of the inflammatory cascade upon interaction with microbial cell wall glycolipids. We study a family of membrane-bound major histocompatibility complex (MHC)-like glycoproteins, the CD1 family, which evolved in vertebrates to capture intracellular lipids and glycolipids of both self and foreign origin for presentation to T lymphocytes. With its lipid tail buried within hydrophobic channels in CD1, the lipid presents its polar head for specific recognition by the T cell antigen receptor (TCR).
We focus on a recently identified lymphoid lineage, the natural killer T (NKT) cell, expressing a conserved set of TCRs with specificity for CD1d. Connections between NKT cells and diseases of infectious, autoimmune, and malignant origin suggest that recognition of lipids of both self and microbial origin is a central immunological component of these processes. We are pursuing two main areas of investigation, one centered on the biochemistry and cell biology of lipid antigens involved in immunity and the other centered on the development and function of T cells that recognize these lipid antigens.
Cell Biology and Biochemistry of Self and Microbial Lipids Recognized by NKT Cells
After biosynthesis in the endoplasmic reticulum, CD1d traffics through the secretory pathway to reach the plasma membrane. It undergoes extensive recycling between the plasma membrane and endosomal compartments through interactions between its cytoplasmic tail and AP-2/AP-3 components of the clathrin-mediated endocytosis pathway. Along this pathway, CD1d acquires and exchanges various lipid cargoes. We have shown that lysosomal saposins perform central functions in this process and, in collaboration with Luc Teyton (Scripps Research Institute), have characterized some of their lipid-exchange function in cell-free assays. The more general question of how microbial and self lipids traffic inside the cell to load CD1 and the role of proteins such as lipid transfer proteins and processing enzymes in these processes remain, however, poorly characterized. We are developing new approaches and reagents to identify these proteins and to visualize the lipids as they traffic through different compartments to load CD1. To provide sensitive and specific functional readouts for the screening and characterization of such proteins, we are synthesizing, in collaboration with Paul Savage (Brigham Young University), a set of "designer lipids" that require specific cellular trafficking and processing steps before recognition by the NKT cell.
We have identified two families of related but structurally different glycosphingolipids (GSLs) that appear to be presented by CD1d to regulate major aspects of the development and function of NKT cells. The self-GSL isoglobotrihexosylceramide (iGb3), a mammalian ceramide with a β-branched Galα3Galβ4Glcβ1 oligosaccharide chain, is recognized by NKT cells and seems essential for their development and maturation. In response to many microbial infections, iGb3, rather than specific microbial lipids, is also responsible for NKT cell activation. In these conditions, TLR signaling of the antigen-presenting cell is essential for iGb3-mediated activation of NKT cells, perhaps through induction of iGb3.
More essential for host defense and perhaps more significant from an evolutionary perspective is our recent identification of novel families of α-branched microbial glycosylceramides, such as α-galacturonosylceramide and α-glucuronosylceramide, that strongly stimulate NKT cells during the course of some infections. These infectious agents include alphaproteobacteria such as Sphingomonas, a pathogen found in soil and perhaps also Ehrlichia, a tickborne pathogen. These bacteria are Gram-negative lipopolysaccharide (LPS)-negative organisms whose cell wall largely escapes recognition by TLR, suggesting that the NKT cell lineage might have evolved to recognize evolutionary alternatives to LPS. Our laboratory is studying the composition of these unusual bacterial cell walls and their effects on innate and adaptive immunity. We are also attempting to model these infections in mouse to study the modalities of microbial invasion, lipid presentation, the interplay of NKT cells with antigen-presenting cells and other components of innate and adaptive immunity, and their role in clearing infection. How these lipids and perhaps other ligands of NKT cells also account for the reported roles of NKT cells in various inflammatory, autoimmune, and cancer conditions is under investigation in our laboratory.
Synthetic microbial ligands of NKT cells are superior adjuvants of immunity because they induce the cross-activation of dendritic cells and NKT cells, which involves CD40L/CD40 interaction as well as the prompt release of an array of cytokines and chemokines. With Paul Savage and Luc Teyton, we are performing pharmacological and structural studies of these adjuvants. Ultimately, we would like to understand the network of molecular and cellular interactions underlying their effects and test their protective value in vaccine trials.
NKT-Lineage Development
The TCR autoreactivity for iGb3 combined with the expression of inhibitory NK-lineage receptors (NKRs) suggests a "signal calibration" model for NKT cell development and activation, similar to that of NK cells, based on a balance of activating (TCR) and inhibitory (NKR) signals. NKRs are usually restricted to another lymphoid lineage, the NK cell, and the lineage bifurcation between T cells and NK cells is controlled by the induction of mutually exclusive transcription factors. Thus, the induction of the NK program in T cells raises an interesting paradox. Our previous experiments demonstrated that the NKT precursors branch off the conventional T cell development pathway at the thymic double-positive (DP) stage. This bifurcation is instructed by the expression of their conserved Vα14-Jα18 TCR, and we have proposed that two types of signals may be involved in this differentiation. One is the signaling emanating from the TCR upon engagement by agonist CD1d-iGb3 ligands. The other category of signal may be provided through homotypic interactions between SLAM (signaling lymphocyte activation molecule)-family receptors whose expression is restricted to the hemopoietic compartment of the thymus. These receptors signal through the adapter SAP (SLAM-associated protein) and the kinase Fyn, and they activate the NFκB cascade, all of which are essential for NKT development. They may also down-regulate Ras signaling through RasGAP, possibly explaining how NKT cells escape negative selection after agonist signaling. We are testing this model by combining genetic and biochemical approaches. More generally, it is apparent that many other lymphocyte lineages, such as γδ T cells, B-1 B cells, and regulatory T cells also arise from instructive signals, depending on the specificity of their antigen receptor. How differential signaling is harnessed to create such a diverse range of lineages is a fascinating area where our studies may provide relevant insights.
We are also pursuing the identification of the molecular mechanisms downstream of cell signaling that establish NKT-lineage commitment and differentiation, and we are generating comprehensive mRNA expression profiles at sequential stages of the differentiation pathway to define the transcriptional elements governing these transitions. Our goal is to identify a putative master transcriptional regulator of the NKT cell lineage.
Our research on lipid immunity has applications to autoimmune, infectious, and cancer diseases and the generation of vaccines. Basic aspects of this research are at the frontier between immunology and the underexplored field of lipid biology, including cell biology of lipids, lipid biochemistry, and lipid signaling. Because T cell–mediated lipid recognition has co-opted the mechanisms involved in these processes, tools and knowledge developed in our field may readily apply to lipid biology at large, and vice versa. Diseases such as atherosclerosis, lipid storage and lipid inflammation, and drugs developed to treat them, are likely to have consequences on lipid immunity, and conversely, CD1-mediated lipid immunity may influence the course of these diseases, as already suggested for atherosclerosis. With our long-standing collaborators in structural biology and lipid chemistry, we hope to elucidate new aspects of lipid immunology, and in turn apply immunological tools to the dissection of basic unresolved issues in lipid biology.
This work is supported in part by grants from the National Institutes of Health.
Last updated: May 15, 2006
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