MOLECULAR-CLONING AND EXPRESSION OF PS20 GROWTH INHIBITOR - A NOVEL WAP-TYPE 4-DISULFIDE CORE DOMAIN PROTEIN EXPRESSED IN SMOOTH-MUSCLE

We previously reported the purification of ps20 (Rowley, D. R,, Dang, T, D., Larsen, IM,, Gerdes, M. J,, McBride, L., and Lu, B, (1995) J. B iol. Chem. 270, 22058-22065), a urogenital sinus mesenchymal cell, sec reted protein having growth-inhibitory properties, We report here clon ing of the 1.03-kilobase rat ps20 cDNA clone from the PS-1 (adult rat prostate smooth muscle) cDNA library. Partial clones were obtained by nested polymerase chain reaction with degenerate primers, and full-len gth ps20 cDNA clones were isolated by plaque hybridization. Sequence a nalysis revealed that ps20 protein contains a WAP-type ''four-disulfid e core'' motif and is a novel member of the WAP signature protein fami ly composed primarily of secreted serine protease inhibitors. Native p s20 immunoprecipitated from smooth muscle cells and recombinant ps20 b oth resolved on SDS-polyacrylamide gel electrophoresis with apparent m olecular mass of 27-29 kDa under reducing conditions and 21-23 kDa und er non-reducing conditions, respectively, Stable ps20-transfectant COS -7 cell lines secreted ps20 and were growth-inhibited relative to mock transfectants. In addition, COS-7 and prostate carcinoma PC-3 cells w ere growth-inhibited by bacterially expressed ps20, Northern analysis indicated differential expression by tissue with highest, expression i n the heart, Immunohistochemical localization of ps20 protein showed c ell-specific expression by both visceral and vascular smooth muscle in all tissues, including the prostate gland. These results indicate ps2 0 is a novel growth-regulatory member of the WAP signature family expr essed by smooth muscle cells.