Studies from the previous two project periods have characterized properties of ps20, a novel protein secreted by prostate smooth muscle. We have reported the cloning of full length cDNA (rat and human) and genomic DNA (mouse and human). The ps20 protein is a new member of the whey acidic protein (WAP) family. This family exhibits growth and differentiation functions. We have made antibody probes to ps20, localized ps20 expression to smooth muscle cells (SMC) in vivo, produced recombinant biologically active ps20, localized ps20 gene to human chromosome 16q24.3, and identified CD9 as a putative membrane interacting/signaling protein. Expression of ps20 was preferential to smooth muscle in vivo, including prostate gland and vascular smooth muscle. Recombinant ps20 induced phenotypic modulation of prostate smooth muscle cells to the migratory, synthetic phenotype in vitro, characteristic of hyperplastic SMC in vascular wall (atherosclerosis) diseases and in culture. In addition, ps20 blocked TGF-beta1 induced phenotypic modulation of SMCs to the contractile phenotype and maintained cells in the synthetic phenotype. Expression of ps20 was specifically regulated by TGF-beta1 in smooth muscle, suggesting an autocrine negative feedback loop. Immunoprecipitation-Western experiments suggests an interaction between ps20 and the CD9 membrane protein in SMCs. These studies suggest ps20 may be a fundamental mediator of smooth muscle phenotype. Benign prostatic hyperplasia (BPH) is a stromal-based disease typified by hyperplastic smooth muscle and ps20-positive SMCs. The present proposal will address the hypothesis that ps20 functions as a key autocrine regulator of synthetic SMC phenotype and that ps20 and the synthetic SMC phenotype play roles in the genesis of BPH. To extend our studies and address this hypothesis in detail we propose: l.) To define the ps20-induced smooth muscle phenotype and identify specific ps20-interacting proteins. 2.) To determine whether there is a correlation between expression of synthetic smooth muscle markers, a nodular BPH phenotype, and ps20 expression in human BPH specimens; 3.) To determine the in vivo function of ps20 by targeted gene knockout in mice, and; 4.) To determine the inductive function of ps20 in vivo by targeted overexpression using prostate tissue-tissue recombination mouse models.
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