The overall aim of these studies is to analyze peptides complexed to MHC molecules on different cells to determine immunodominant self peptides that may be involved in shaping the peripheral T cell receptor repertoire as well as those that may be targets for tissue specific autoimmunity. Our recently developed radiochemical methodology will be used to isolate and analyze the endogenously bound peptides. Immortalized cell lines of lymphatic origin, primary cultured cell lines and lines from different tissues established from a transgenic mouse bearing a conditional mutant of SV40 early region will be studied. Comparative peptide analysis will also be carried out on separated thymic cells involved in T cell maturation or lines derived from thymic cultures. From comparative HPLC analyses relevant self specific peptides will be characterized by large scale peptide isolation and amino acid sequence analysis. To complement our biochemical approach and broaden our understanding of specific peptides we will expand our panel of allogeneic antiKb CTL raised across H-2 (bm mutant anti parent) as well as non-H-2 differences to assess biological recognition of tissue specific peptides. We also propose to prepare tissue specific CTL by special immunization protocols. Thus, these approaches should be useful in analyzing not only peptides from peripheral tissues but also MHC bound peptides from presenting cells in the thymus (cortical epithelial cells, medullary and dendritic cells and macrophages) which could be responsible for shaping the T cell repertoire. The approach is of value for disease models specifically such as the autoimmune thyroiditis model in which there is an apparent involvement of MHC class I products (Kb) which can be studied for peptides relevant to the diseased state.
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