The objective of this study is to continue to investigate the mechanisms by which extracellular stimulatory molecules such as hormones and growth factors turn on genes- an important example of signal transduction. This laboratory had earlier discovered that in Dictyostelium discoideum, extracellular cAMP induces several genes as would a peptide hormone, by binding to cell-surface receptors and activating signal transduction pathways. This observation and the unique features of the organism make Dictyostelium extremely advantageous for characterizing signal transduction pathways. Reversible protein phosphorylation appears to be important in the pathways between the cell-surface receptors and cAMP inducible genes. To identify the elements in the functioning of the signal transduction pathways, the proposed research will involve further cloning of genes encoding protein kinases and phosphoprotein phosphatases. The pattern of expression of the genes will help to focus on a subset of genes involved in the signal transduction pathways of interest. Such genes will be further analyzed by functional inactivation using antisense RNA and/or homologous recombinadon leading to gene disruption. Experiments are proposed to identify the potential substrates of these enzymes with the view to characterizing other steps in the cascades. The end of the reaction pathways probably involve interactions between DNA binding proteins and regulatory sites on inducible genes. The research will characterize the proteins involved in interacting with a previously defined cAMP """"""""responsibility"""""""" element (CRE) necessary for the induction of a well characterized gene by extracellular cAMP. These studies should help to identify the elements necessary for the induction of eukaryotic genes by extracellular molecules, and by analogy elucidate pathways involved in proviral induction in AIDS or abnormal proliferation in cancer.
