Project Summary It is well known that particular major histocompatibility complex (MHC) alleles promote the appearance of many autoimmune diseases. It is also known that certain MHC alleles protect against such illnesses, even in the presence of the disease-promoting allele. Studies over the last 20 years have suggested many explanations for the inhibitory effects of some MHC alleles, with explanations ranging from the fact that the protective alleles may delete or dilute out the potentially damaging T cells to the notion that the protective alleles induce regulatory T cells which prevent attack on the tissue target of autoimmunity. Any of these explanations suggest that MHC heterozygosity affects the T cell receptor (TCR) repertoire and function of the T cells in the animal. A funded R21 supports studies on the effects of MHC heterozygosity on the TCRs bourne by naïve CD4s. Because large scale methods to sequence and identify the TCRα and TCRβ pairs in individual T cells were not available the mice used expressed a single TCRβ. Comparison of the TCRα sequences on naïve CD4 T cells in mice with different MHC alleles showed that, surprisingly, the TCR repertoire of CD4 cells in healthy MHC heterozygous mice was certainly not as large as that of the sum of the two homozygous parents and, in some cases, was considerably lower. Here we would like to find out how the TCR repertoires of CD8 T cells differ between healthy MHC homozygous and heterozygous animals. Other proposed experiments will study the effects of MHC heterozygosity on TCR repertoires in autoimmune prone animals. In this case the mice will express fully functional TCRα and TCRβ loci. Current methods allow knowledge of the pairs of TCRα and TCRβ on relatively few cells (<4000) compared with the total number of TCRα TCRβ pairs (>million) that are present in normal mice or humans. In the experiments proposed here we will adapt current methods in order to develop procedures that will provide greater yields of such pairs. The results of our experiments will inform knowledge of the consequences, for the T cell receptor repertoire, of MHC heterozygosity versus homozygosity in animals with fully functional TCRα and TCRβ loci. The results of such sequencing experiments, combined with data from experiments with tetramers expressing autoantigen MHC/peptide ligands will suggest ideas about how some MHC alleles are protective against autoimmune diseases. The results will also provide more efficient methods of evaluating TCR repertoires in normal individuals.