The MHC represents a principal susceptibility locus for type 1 diabetes (T1D) in humans and NOD mice, but the mechanisms by which particular MHC genes confer susceptibility or dominant resistance to T1D are not well understood. The major hypothesis of this project is that MHC class II molecules which confer susceptibility to T1D promote efficient positive selection of islet-specific T cell population(s) in the thymus and later present islet-derived peptides to these CD4 T cells in the target organ. This hypothesis is supported by preliminary data which demonstrate that T cells specific for a peptide recognized by a panel of islet specific T cell clones from NOD mice are present in the thymus at a frequency (about 1:1,000) that is several order of magnitude higher than estimated for an average T cell specificity (<1 per million). These studies have been enabled by novel tools that the PI has developed for the generation of MHC class II tetramers and the detection of T cells present at low frequencies. CD4 T cells specific for several islet autoantigens will be examined in the thymus of young NOD mice with tetramers of I-Ag7/peptide complexes in order to determine which islet specific T cells undergo efficient positive selection. Expression of other MHC class II molecules has been shown to confer dominant protection from T1D, and the PI will determine whether the mechanism is based on negative selection of islet-specific T cells in the thymus or modulation of their differentiation into effector populations is pancreatic lymph nodes and islets. A second major hypothesis of the project is that similar antigen presenting events are relevant in T1D in the NOD mouse model and in humans. This hypothesis is based on substantial structural similarities between murine I-Ag7 and human DQ8, as well as functional studies demonstrating similar peptide binding properties. This hypothesis will be examined by analysis of CD4 T cell populations in patient with new-onset T1D and in prediabetics using tetramers of DQ8 loaded with islet peptides that bind to both the human DQ8 and murine I-Ag7.