The aim of this proposal is to gain a better understanding of the structure and binding properties of the murine T cell receptor and to investigate the role of thymic selection in its expression. Initially, we will focus on characterizing the recently isolated alpha chain of the receptor with respect to genomic organization and the pattern of framework and hypervariable residues in the variable region sequences of different cDNA clones. We will also examine the relative expression of alpha and beta chains in different thymic subpopulations using cell separation procedures and hybridoma technology. In particular, we will follow up preliminary indications that alpha chain genes are expressed after beta chain genes, perhaps enabling us to define a """"""""pre-T"""""""" cell phenotype analogous to that established in B lymphocytes. Subsequent work will involve efforts to transfer a given antigen-MHC specificity into T cell tumor lines using rearranged genomic alpha and beta chain genomic DNAs and standard methodologies. If successful, this will provide an easily manipulatable system for studying different components of the receptor binding site and other important structural features of the receptor complex. We will also use mice which are transgenic for alpha and beta chains of defined specificity to ask questions about expression and thymic selection in mice with compatible and incompatible MHC haplo-types. Lastly, we will try a number of expression systems and truncation of the trans-membrane portion of the two chains in an effort to produce large quantities of soluble heterodimer. This material will be used for binding studies to try and define what T cell receptors can """"""""see"""""""" and with what affinity. This will also provide the necessary starting material for protein crystallography and subsequent structural studies.
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