This project centers on the analysis of CD4 and CD8 T cells specific for beta cell autoantigens in the diabetic model of NOD mice. One goal is to document the spectrum of beta cells proteins and peptides responsible for triggering CD4 T cells and CD8 T cells. The plan is to expand the number # of CD4 T cell clones available to us in the search for the beta cell antigen; we also plan to induce T cells to known beta cell antigen/peptides and determine if these are diabetogenic. An effort will also be made to identify the proteins in the secretory granule from beta cells that stimulate some of our diabetogenic T cells. These efforts are coupled to biochemical studies on the Ag7 molecules and the peptides that associate with it. We want to examine for sequence motifs and the relationships between peptide binding specificity and affinity to Ag7 and Ag7 stability. Our ultimate goal will be to define diabetogenic peptides and to determine which are major (dominant) or minor and which are physiologically presented by islet APC. We also plan to examine the biology of CD8 T cells by creating a transgenic line bearing a T cell receptor gene for a CD8 islet cell specific T cell. If successful, this line may be useful for understanding the dynamics and the interrelationships between CD4 and CD8 T cells in islet cell injury. Finally, we will investigate peptides extracted from the Kd class I MHC molecules of insulinomas that stimulate islet specific CD8 T cells. A major issue of this proposal is the biochemical analysis of peptides associated with MHC molecules which will use technologies now in place to examine model protein antigens. The mass spectrometry resource will be used to i) identify peptides from beta cells that interact with Ag7; ii) identify peptides physiologically associated with Ag7 from APC of spleen or established lines and iii) identify peptides associated with Kd molecules responsible for stimulating CD8 T cells.
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