Guanylyl cyclase receptors: Targets for medical intervention

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We will define the functions of GCB (NPR-B), a membrane guanylyl cyclase receptor activated by the local paracrine/autocrine ligand, C-type natriuretic peptide (CNP). GCB is highly expressed in chondrocytes, adipocytes and fibroblasts; GCB activation usually leads to an inhibition of cell proliferation and hypertrophy. We recently identified sphingosine-1 -phosphate (S1P) as a potent, effective and highly specific GCB desensitization factor. The first specific aim concentrates on the molecular pathway by which S1P specifically induces GCB desensitization. These studies are important in that CNP, which acts as an antagonist to S1P, is strongly implicated as having an important role in tissue remodeling, including atherosclerosis, restenosis following angioplasty, and even liver disease. Mouse genetic models, siRNA and interaction trap screens (immunoprecipitation, yeast two hybrid, Sos-Ras) will define proteins that mediate the S1P-induced desensitization of GCB. In preliminary research we have already identified various candidate GCB-associated proteins. These proteins will themselves be used in interaction trap screens to define other molecules within the S1P-GCB desensitization pathway. Desensitization of GCB is ultimately regulated by dephosphorylation of the receptor, and therefore the protein kinase/protein phosphatase responsible for GCB regulation will be purified, identified and mechanisms of regulation determined. We also have disrupted the GCB gene, and then corrected a severe dwarfism by expression of GCB specifically in chondrocytes in the null background. These GCB animals contained almost no fat compared to wild-type littermates. Therefore, the second specific aim concentrates on the functional roles of GCB in fibroblasts and adipocytes. We will compare the effects of various treatments (in vitro and in vivo) on fibroblast and adipocyte function, with particular emphasis on the role of GCB in tissue remodeling after wounding and in adipocyte differentiation. The proposed studies will have significant and substantial implications in human disease, since one of the largest health problems in America is obesity, often caused by an over-abundance of fat cells. Obesity is a high risk factor for non insulin-dependent diabetes and hypertension. Thus, the proposed research impacts on some of our most important medical issues, including wound repair, atherosclerosis, restenosis following angioplasty, and a host of diseases associated with obesity.