This application is directed toward the investigation of the role of Class II MHC genes in susceptibility and resistance to IDDM through the study of three candidate islet antigens, glutamic acid decarboxylase (GAD) 65, heat shock protein (HSP) 60, and insulin B chain. Under the first aim, T-cell clones specific for these three antigens will be derived from NOD mice and will be used to identify immunodominant epitopes presented by susceptible (I-A g7), and resistant (I-A g7 PD, I-Ad) Class II molecules. These I-A molecules will be affinity purified for measuring binding affinities of peptides identified by the T-cell clones. The final part of aim 1 will be to determine various parameters of these Class II molecules-cell surface expression, level, stability, etc. - to investigate the nature of MHC-peptide complexes. The underlying hypothesis is that susceptibility and resistance to diabetes correlate with the binding of peptides to MHC Class II, the number of complexes formed, and/or their relative stability. Under the second aim, effects of the candidate antigens on disease progression will be studied. Mechanisms of the in vivo tolerizing effects of these antigens will be investigated through an analysis of the T-cell responses in recipient and control mice.
Aim 3 will look at the effects on diabetes development of expression of the three candidate antigens in transgenic mice under control of the Class I MHC promoter and/or the beta actin promoter. These studies will include induction of embryonic and neonatal deletion of islet-specific T-cells to determine whether candidate autoantigens are primary targets.
Aim 4 is directed toward the identification of new beta cell antigens, using T-cell clones of unknown antigen specificity to screen an islet cell cDNA expression library.
