STRUCTURE AND FUNCTION OF CD6 IN THE MOUSE

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Anti-CD6 monoclonal antibodies and immunotoxins have been used as immunosuppressive therapies for the treatment of renal allograft rejection and graft-versus-host disease. In particular, purging of donor marrow with anti-CD6 reagents has prevented graft-versus-host disease without significantly lowering the incidence of engraftment in allogeneic bone marrow transplantation. Thus, such reagents may be of important therapeutic value in treatment of malignancies and other disorders by allogeneic bone marrow transplantation. There is thus far only limited knowledge about the biological function of CD6 except that it plays some role in T cell activation. Furthermore, a mouse homolog of CD6 has not been identified to date. The goal of this study is to analyze the structure and function of CD6 in the mouse system. This will be done by further characterization and sequencing of isolated cDNA and genomic clones encoding this protein. Monoclonal antibodies specific for mouse CD6 will be generated using the gene gun to immunize rats followed by fusion of the spleen cells, selection and screening for appropriate antibody-secreting hybridomas. These antibodies will be used to determine the tissue and cell type distribution of mouse CD6 expression as well as the pattern and level of expression during thymocyte development. They will also be used to determine their functional effects upon mouse T cells, in particular as costimulatory reagents for T cell activation. The pathway of signal transduction through the long cytoplasmic tail of this protein will be approached by using the yeast-two-hybrid system to isolate clones encoding proteins that interact with this cytoplasmic tail and by examining effects of progressive deletions in the cytoplasmic tail on the ability of CD6 to costimulate T cell activation. The physiological importance of this molecule will be further examined by the generation of mutant mice in which the CD6 gene has been knocked out by homologous recombination. The effects of lack of CD6 expression upon T cell development and activation will be examined.