This project will identify MHC class I restricted beta-cell autoantigens which are the targets of CD8+ T cells involved in the initiation of the autoimmune response in the NOD mouse model of IDDM. Studies begun in collaboration with David Serreze and Derry Roopenian at Jackson Laboratories have shown that infiltrating lymphocytes from islets of NOD mice developing diabetes can be cultivated in vitro when fed on islet cells. A CD8+ clone (A1-4.4)has been established from bulk cultures which recognize and specifically kill beta-cells. Three other lines have been isolated to date and clones are being established. These clones appear early in the course of autoimmune insulitis in the NOD mouse and presumable are reactive to critical targets in initiating the disease. This detection system is independent of biases related to known autoantibodies. We propose to isolate and characterize peptides from the MHC class I molecules of transformed beta-cells which are the targets for these CTL clones. Determining the sequence of the peptides by microsequencing and tandem mass spectrometry will allow identification of the autoantigen from which the peptides are derived, thus allowing us to identify antigens against which the autoimmune reactivity is directed. The information on peptide targets of the CD8+ T cells and the autoantigens provide an approach to understanding the mechanism for T cell destruction of beta-cells, and may allow development of reagents useful for prevention of IDDM.
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