NON-H-2 HISTOCOMPATIBILITY GENES AND ENCODED ANTIGENS

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The success or failure of tissue allografts is determined by the products of a large family of autosomal and sex-linked, polymorphic genes that are designated histocompatibility (H) genes. The strongest barriers to successful transplantation are determined by the alloantigenic products of class I and class II genes of the major histocompatibility complex (MHC). The matching of donors and recipients for MHC class I and class II alloantigens not only reduces or eliminates MHC-encoded barriers but, conversely, optimizes the presentation of donor minor H antigens that require MHC matching of donor and recipient for presentation. The demonstration that the antigenic components of single minor H antigens are peptides bound to the peptide binding site of MHC molecules provides not only a reason for why minor H genes have eluded cloning, but, also provides an experimental approach to identifying and sequencing these H antigen peptides. Using the knowledge of the class I molecule that is required for the presentation of a single minor H antigen to cytolytic T lymphocytes (CTL) and the fact that subsets of endogenous peptides bind to individual class I molecules, we have chromatographically resolved peptides from multiple minor H antigens. The amino acid sequences of these minor H peptides can be applied to cloning and sequencing the respective genes. The overall objectives of this research program are to clone and identify minor H genes in the mouse and to understand the specificity and genetic control of the T cell response to their encoded antigens. The first aim is the amino acid sequencing of minor H antigen peptides by mass spectrometry and application of the amino acid sequences to the synthesis of degenerate oligonucleotides for the cloning of minor H gene CDNAS. H gene CDNAS will be confirmed by sequencing and DNA-mediated gene transfer into eukaryotic cells with subsequent testing with specific CTL. The second objective is the estimation of diversity of minor H peptides expressed by inbred mouse strains by amplification and sequencing of DNA sequences that encode the homologous peptides. The third objective is the testing of our hypothesis that minor H peptides comprise a group of non-abundant endogenous peptides that exhibit relatively higher affinity for class I molecules than that exhibited by abundant, endogenous peptides. Peptides derived from minor H antigens, viral and soluble antigens, and abundant endogenous proteins will be tested for relative affinity in competitive binding assays and assays for stabilization of expression of class I:peptide complexes. The fourth objective is the sequencing of immunodominant minor H antigen peptides and the testing of the hypothesis that the ranking of these dominant antigens is based on the relative affinity of their peptides for class I molecules. These studies constitute the first analysis of the T cell response to minor H antigens that will both sequence minor H antigen peptides and genes as well as investigate the affinity and specificity of the binding of these peptides to MHC class I molecules.