REGULATION AND FUNCTION OF THE CD4 GENE

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Mature T lymphocytes can be divided into two subsets based upon their expression of either of two cell surface glycoproteins, CD4 or CD8. T cells that recognize class II major histocompatibility complex (MHC) proteins generally express CD4 and are helper or inducer, while those that recognize class I MHC proteins generally express CD8 and are cytotoxic (or possibly suppressor). CD4 has been shown to play important roles in enhancing T cell responses to antigen and in the selection of T cells during thymic development. The goal of this application is to define the mechanisms by which CD4 accomplishes these two functions. A CD4-CD8-, Kb responsive T cell hybridoma will be used to determine whether CD4 can enhance responses to class I MHC proteins, and if so, whether this occurs merely by adhesion or also by signal transduction. The role of the tyrosine kinase p56lck in CD4-mediated enhancement of antigen responses will be examined using transfection of wild-type and mutant forms of p56lck in a class II restricted, antigen dependent T cell hybridoma. Interactions between CD4 and a constitutively active form of p56lck will be examined to determine whether their effects are additive, whether the two associate, and the effects of cross-linking of CD4 and/or the T cell receptor (TCR) on protein associations, tyrosine phosphorylation of substrates and antigen responses. The role of the TCR-associated p59fyn tyrosine kinase in CD-4 mediated stimulation of antigen responses will also be assessed using mutant and wild-type p59fyn constructs. The mechanism(s) by which double- positive thymocytes become single positive for CD4 if they have a class II restricted TCR, or CD8 if they have a class I restricted TCR will be explored. this will be done both by mating transgenic mice expressing a chimeric CD8/CD4 construct to a class I specific TCR transgenic mouse and to a class II deficient mouse. Homologous recombination will also be used to generate mice in which the endogenous CD8alpha gene is replaced with CD8alpha having a CD4 transmembrane region and cytoplasmic tail, or the endogenous CD4 gene is replaced with the reverse construct.