CROSSTALK BETWEEN CAMP AND RAS IN ENDOCRINE CELLS

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DESCRIPTION (Taken from the applicant's abstract) Because cells respond to a wide array of signals received at the cell surface, their ability to integrate and respond appropriately to these signals is essential. One of the best studied examples of signaling crosstalk is the interplay between Ras- and cAMP-mediated pathways. Cyclic AMP inhibits proliferation in many cells, in part through uncoupling Ras from one of its downstream effectors, Raf-1. Elevations in cAMP, however, are not universally growth inhibitory. In thyrocytes, pituitary somatotrophs and other cells, cAMP stimulates proliferation. We discovered that thyrotropin (TSH) stimulates proliferation through a novel pathway involving both Ras and the cAMP-dependent protein kinase, but not the well described downstream effectors of Ras, Raf-1 and the mitogen-activated protein kinase cascade. The effectors used by Ras in the presence of TSH and elevated cAMP levels remain to be elucidated, but may include phosphatidylinositol 3-kinase, the isoform of protein kinase C, and other small G proteins. It is the goal of these studies to elucidate those molecules which transduce TSH-stimulated mitogenic signals, including the identification of novel Ras effectors, and the sites of interaction between Ras- and cAMP-mediated pathways. Using microinjection of purified signaling molecules and highly specific inhibitors, the biological activity of a panel of Ras mutants defective in various effector interactions will be assessed. Ras-overexpressing thyroid cells will be screened for novel proteins that bind to GTP-bound Ras. Other molecules proposed to function as Ras effectors will be assessed for growth-stimulating activity in the thyrocyte. Together these approaches will provide insight into novel Ras-mediated signaling pathways active in thyroid cells. Once these molecules are identified, we will test their activity in growth stimulation in other epithelial cells, including pituitary somatotrophs and mammary epithelial cells. Ras gene activation and the constitutive activation of cAMP-mediated signaling pathways have been identified in human thyroid and pituitary cancer. The elucidation of the molecular components of, and interactions between these pathways may identify novel targets for therapeutic intervention. My long term career goal is to identify the molecular components of growth-signaling pathways in a variety of epithelial cell types in a challenging academic environment. The University of Pennsylvania provides an exciting and rich research environment in which to pursue these studies.