Molecular recognition in the immune system
My primary research interest is in the biochemical processes that underlie cellular recognition and signaling. Research in my laboratory has concentrated on the immune system, because of its intrinsic importance to human health and disease, and because of its treasure of biochemical mechanisms by which cells communicate with their environment and with each other. Our approach combines in vitro biophysical and biochemical studies of the proteins involved in these processes, with cellular studies of their functions and intermolecular interactions. We have focus on two related areas: antigen presentation by MHC proteins, and the molecular mechanisms of T cell activation.
Antigen presentation by MHC proteins
A key feature of immune recognition is the interaction of proteins encoded by the Major Histocompatibility Complex (MHC) with antibody-like receptors on T cells. MHC proteins bind peptide antigens within the cell, and display them at the cell surface for interaction with T cell receptors. MHC proteins are highly polymorphic, with allelic differences associated with differences in peptide binding preferences and in individual susceptibility to allergy, infection, and autoimmune disease. We study MHC proteins and their interactions with peptides through a combination of biophysical and cellular analysis. The crystal structure of an MHC-peptide complex revealed that peptides of completely different sequence can be accommodated in the site with almost no alteration of the MHC molecule, with binding specificity determined by subtle details of interactions in the side-chain binding pockets. Hydrodynamic, enzymatic, and spectroscopic studies have identified a peptide-dependent conformational change, in which the region ß58-69 of the MHC protein folds over the bound peptide to trap it in the site, in the rate determining step for the overall peptide binding reaction. In continuing studies, we are investigating the mechanism of peptide-exchange catalysis by DM, a major unsolved problem with implications to other systems in which structural homologues function as chaperones or conformational catalysis. In cellular studies, we have discovered a new extracellular pathway for antigen presentation particularly active in immature dendritic cells, in which peptide generation and MHC loading occur entirely outside of the cell. This pathway may help to explain the unique antigen presentation abilities of dendritic cells, particularly their role in maintaining peripheral tolerance to self-antigens. Recently we have observed that this process occurs also in microglia, enigmatic brain cells involved in immune tolerance and response to infection.
T cell activation
The interaction of MHC-peptide complexes on the surface of an antigen presenting cell with receptors on T cells induces characteristic T cell effector functions. Once activated, T cells play crucial roles in the initiation and control of the immune response to foreign material in the body, by killing infected cell and activating other immune system cells. The molecular events at the juxtaposed membranes of T cell and antigen presenting cell that trigger these cellular activation processes are unknown, but are believed to involve receptor aggregation or clustering. To approach the mechanism of aggregation-activated signaling in T cells, we developed a novel model system using chemically-defined oligomers of MHC-peptide complexes, and used them to determine the minimal requirements for T cell activation. For a given amount of receptor engagement, the extent of activation was equivalent for MHC dimers, trimers, tetramers, and octamers, but monomers were inactive, showing definitively that TCR dimerization was necessary and sufficient for activation. Using dimers with various topological constraints, we showed that that the activation trigger did not involve a ligand-induced allosteric change, as observed for many other dimerization-activated receptor systems, but rather a ligand-induced oligomerization. To investigate the molecular events by which oligomerization of ligand-binding domains communicates a signal into the cytoplasm, we have begun structure-function studies of TCR cytoplasmic domains implicated in signaling. We have found that the cytoplasmic domain of TCR zeta exhibits a lipid-dependent folding transition, which regulates accessibility to cytoplasmic protein kinases known to interact with engaged receptor. Based on these results we proposed a novel mechanism for coupling receptor clustering to signaling cascades through zeta subunit conformational changes. In addition to helping unravel the activation mechanism, MHC oligomers are proving to be very useful reagents to detect and identify specific CD4+ T cells present at low frequency in mixed populations in blood and other clinical samples. In continuing work we are probing the structure of receptor components, developing new methods to investigate the T cell activation trigger, and using MHC oligomers to detect antigen-specific T cells in malaria, influenza, and HIV infection.
Figure 1: Structure of an antigenic peptide from influenza virus bound to the class IIMHC protein HLA-DR1. The MHC peptide binding domain is shown as a cyan surface,the influenza peptide as a CPK model. Antigen receptors on T cells bind to thiscomplex as part of the process that triggers an immune response. The structureand function of MHC proteins, and the cellular pathways by which they are loaded,are a focus of study in the Stern laboraotry.
Figure 2. Model for the interaction of an MHC-peptide complexon one cell with a T cell receptor on another cell. Recognitionof foreign MHC-peptide complexes activates the T cells to killthe presenting cell or to recruit other immune cells to the vicinity.The triggering process involves clustering or aggregation ofTCR on the T cell surface. Determination of the molecularmechanism of such clustering-induced signaling is anotherfocus of research in the Stern Laboratory.