Molecular Genetic Analysis Of Lymphocyte Function

National Institute Of Allergy And Infectious Diseases

Investigators

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Abstract

Sections 1, 2, and part of 3, as listed above, deal with the MHC-I aspects of this project, and in general are directed to understand the molecular details of the loading of MHC-I molecules with self or antigenic peptides. Part 1 has resulted in a detailed understanding of the three-dimensional structure and structural changes that accompany MHC-I interaction with its chaperone, TAPBPR, and with the release of the chaperone from the MHC-I molecule on interaction with peptide. In additional studies (part 2) of the studies of MHC-I/peptide interactions, we have explored the role that the anti-retroviral drug, abacavir, plays in binding to MHC-I and distorting the self-peptide repertoire bound by susceptible MHC-I alleles. In particular, we examined the biological effects of abacavir binding to HLA-B*57:01 in a model animal system that we have developed. Thus, abacavir alone, when bound by HLA-B*57:01 in a transgenic animal, can elicit a neoantigen T cell response, dependent on regulation of CD4 T cells. This animal model system provides an explanation for the severe hypersensitivity reactions that are observed in a high proportion of HLA-B*57:01 individuals who receive the drug. The third part of this project is focused on structural and functional studies of T cell receptor recognition of antigens, how this leads to T cell signaling, and how this leads to autoimmune disease. Finally, in part 3, to provide a baseline for understanding antigen-specific structural changes in the T cell receptor (TCR), we have determined the X-ray structure of a virus specific, MHC-I-restricted TCR, as well as its complex with its MHC-I/viral antigen ligand. Remarkably, although the MHC/peptide complex has a relatively rigid structure, the TCR shows great movement of its CDR3 alpha and beta loops, indicative of a fly-casting mechanism for ligand engagement. Further characterization of this fly-casting mechanism is reflected in other projects. We have collaboratively explored the use of NMR to study conformational changes in the TCR that accompany binding of a high affinity peptide/MHC complex. These studies offer new insight into allosteric mechanisms that contribute to T cell activation. Other approaches to understanding the TCR mediated aspects of autoimmunity include: 1) the characterization of antigenic peptides recognized by the autoimmune T cells in transgenic mouse models of autoimmune gastritis; and 2) the structural determination of MHC-II molecules bound to their antigenic peptides. We have determined the high resolution X-ray structure of IAd in complex with the Th2 peptide known as PLL, as well as the structures of two other IAd/peptide complexes in which the peptides are related to PLL, but are of higher intrinsic affinity. These structures, determined at high resolution reveal a previously unrecognized binding motif (exploiting residues 1,4,6,7, and 9 of the peptide) for IAd, particularly with respect to the preference of glutamic acid at position 9 of the peptide. This provides a framework for understanding interactions of autoimmune TCR with self MHC-II/peptide complexes. These experimental structural studies permitted the modeling of another gastritis-inducing peptide, known as PIT, bound to IAd, and provide further insight into the molecular basis of autoimmune gastritis. Similarities in this theme of the relevant anchors and the topology of the peptide bound to the MHC are consistent with other MHC-II/autoantigen complexes and suggest some common features that may be specifically relevant to autoimmune antigens.