Summary: Albert Bendelac studies innate lymphocytes, with a focus on NKT cells and their recognition of self- and microbial glycolipids presented by major histocompatibility-like molecules of the CD1 family. This research has direct implications for disease conditions such as microbial and viral infections, cancer, allergic inflammation (asthma), inflammatory bowel disease, and autoimmunity. His lab also studies synthetic ligands of innate lymphocytes, as potential adjuvants of infectious and cancer vaccines.
The hallmark of adaptive immunity is the production of naïve B and T lymphocytes that traffic to lymph nodes and undergo clonal expansion upon exposure to antigen, followed by the acquisition of specialized effector functions and tissue migration properties. In contrast with this well-established scenario, many unconventional subsets of B and T cells undergo clonal expansion and acquire distinct effector differentiation and tissue-homing properties during development, following exposure to endogenous rather than foreign ligands. These innate lymphocytes, which collectively are a substantial fraction of total lymphocytes, are viewed as distinct lineages carrying out "hardwired" rather than adaptive strategies of immune defense. An understanding of their development and their role in health and disease has just begun to emerge.
NKT cells, one of the most prominent populations of innate lymphocytes in mouse and human, are the main focus of our laboratory. We seek an understanding of the cellular and molecular determinants of their lineage, of their lipid antigens and the CD1 family of glycoproteins that present them at the cell surface, and of their role in multiple diseases, including allergic inflammation, infection, autoimmunity, and cancer.
Innate Lymphocyte Lineage Development
The key determinant of innate lymphocyte lineages is the expression of particular T cell antigen receptors (TCRs) or BCRs. Transgenic expression of these antigen receptors invariably induces their lineage of origin. NKT cells express conserved TCRs whose recognition of CD1d/lipid complexes is mostly carried out by a single-germline gene segment, Jα18, in mouse and human. A second determinant of the NKT lineage is the recognition of these CD1d ligands on thymocytes rather than epithelial cells. Redirection of major histocompatibility complex (MHC) class II expression on thymocytes is sufficient to generate CD4 T cells with innate-like effector properties. We have identified two essential molecular pathways elicited by the recognition of ligands on thymocytes. SLAM (signaling lymphocyte activation molecule)-family members signaling through the adaptor SAP (SLAM-associated protein) and the kinase Fyn activate the NFκB cascade, which is essential for cell expansion and survival of innate T lymphocytes. These SLAM-family receptors are engaged during homotypic (T-T) but not T–epithelial cell interactions.
A master transcription factor is PLZF (promyelocytic leukemia zinc finger), which is necessary and sufficient to induce the effector and migratory properties of NKT cells. However, the induction of PLZF after TCR engagement is independent of the SLAM pathway, implying that additional signals, perhaps elicited through the recognition of agonist ligands, are required to induce PLZF. Both SLAM and PLZF are involved in several other innate-like lineages, such as MAIT (mucosal-associated invariant T) cells specific for the MHC-like molecule MR1 and NKT-like Vδ6 T cells.
The emerging picture is that the mirror distribution of MHC versus MHC-like ligands on epithelial versus hemopoietic cell types is the fundamental decision maker between adaptive and innate-like T cell lineages. The equivalent pathways governing the choice between adaptive and innate-like B cell development remain to be identified. Our current efforts focus on understanding the mechanisms responsible for induction of PLZF during NKT cell development and on identifying the set of genes that are the target of PLZF. A powerful approach uses ChIP-seq (chromatin immunoprecipitation combined with parallel sequencing) and RNA-seq at sequential stages of NKT cell development.
Lipid Recognition in Health and Disease
Unlike proteins, which mutate at a high rate, lipids and glycolipids provide relatively fixed targets for the immune system. With the fatty acid chains buried within hydrophobic channels of CD1 glycoproteins, the lipid presents its polar head for specific recognition by the TCR. We have identified several prominent and conserved ligands of NKT cells, including the self-glycosphingolipid isoglobotrihexosylceramide (iGb3), a mammalian ceramide with a β-branched Galα3Galβ4Glcβ1 oligosaccharide chain, and microbial cell wall glycosylceramides such as α-galacturonosylceramide and α-glucuronosylceramide.
Other self-ligands such as lysophosphatidyl choline, abundantly released during inflammation, and foreign lipids such as the α-glycosyldiacylglycerols of Borrelia burgdorferi have been reported. We are studying the relative importance of these ligands in NKT cell development and in the response to various infectious, inflammatory, and cancerous conditions. For example, we recently reported that infection with Sphingomonas resulted in chronic granulomatous inflammation of the liver with the production of autoantibodies to mitochondria, closely reproducing human primary biliary disease. The autoimmune process was triggered through the activation of NKT cells by Sphingomonas cell wall α-glycuronosylceramides, breaking B cell tolerance to self-epitopes contained in the PDC-E2 (pyruvate dehydrogenase complex E2) enzyme of Alphaproteobacteria (which are evolutionarily related to mitochondria).
We are also exploring connections between asthma and NKT cells. Like other microbial cell wall components (e.g., lipopolysaccharide), microbial NKT ligands are found in airborne particles. Their inhalation induces NKT-mediated allergic airway inflammation in mouse through mechanisms under investigation.
Cell Biology of Lipid Presentation by CD1 Molecules and Development of Synthetic NKT Ligands as Vaccine Adjuvants
The recognition of self- and microbial lipids involves lipid transport proteins, lipid-processing enzymes, and lipid-exchange proteins, including proteins that exchange lipids between CD1 and cell membranes. We have identified and characterized saposins as essential lipid-exchange proteins for CD1 loading in endosomal compartments. The cell biology of these events turns out to have major consequences on the stimulatory properties of individual lipids. This is the focus of laboratory studies on the development of synthetic NKT ligands as a new class of vaccine adjuvants. NKT cell recognition of ligand induces rapid and reciprocal activation of NKT cells and their antigen-presenting cell (dendritic cell, B cell, or macrophage) through CD40/CD40L signaling and Th1/Th2 cytokines. These interactions recruit cellular networks that activate innate and adaptive immunity.
Changes in lipid structure that do not impact TCR recognition nevertheless drastically alter the stimulatory properties of lipids. We have shown that two classes of variants elicit different Th1 or Th2 adjuvant properties based solely on the presence of long and saturated versus short or unsaturated lipid chains, respectively. These properties correlate with the requirement for endosomal loading (and lipid transfer proteins) for long and saturated lipids, whereas short or unsaturated lipids can be loaded at the cell surface. Using mice bearing a conditional CD1d allele, we are exploring the hypothesis that dendritic cells are required for the presentation of the long, but not the short, lipids. Dendritic cells are a major source of interleukin-12; their recruitment may explain the Th1 property of some NKT ligands. We are developing antipolysaccharide vaccines based on NKT adjuvants, which rely on NKT cell help to B cells. Good polysaccharide vaccines are essential for the eradication of various bacterial diseases, including pneumonia caused by pneumococcus, but the limited efficiency of these vaccines has been a long-standing worldwide health problem.
Other Innate-like Lineages
Our laboratory also investigates other major innate-like lymphocyte lineage populations. For example, we are studying gut intraepithelial lymphocytes, a population whose development, function, and ligand recognition have nevertheless remained obscure. We have isolated multiple TCRs originating from CD8αα TCRαβ intraepithelial lymphocytes, and we are generating transgenic systems to model their developmental pathway and study their ligand recognition properties.
Our studies focus on a distinct, innate host defense strategy that evolved on the margins of the adaptive immune system. Antigen receptor gene segments of specific protective value were selected and conserved across multiple species; their expression was also used to induce complex genetic programs that define a variety of specialized lymphocyte sublineages. The study of these idiosyncratic lineages is a necessary step to develop a full understanding of immunity, particularly its innate tissue-specific components. These fascinating genetic, developmental, biochemical, and cellular investigations have great potential for the development of novel immunomodulatory compounds that harness the myriad of functions associated with innate lymphocytes.
This work is supported in part by grants from the National Institutes of Health.
As of May 30, 2012