Showing papers by "Robert D. Fleischmann published in 2016"

Evidence that a 90-kDa phosphoprc specific phosphatase are involved i) melanoma cell proliferation by insu

Abstract: Proliferation of wild-type Cloudman S91 melanoma cells is inhibited when insulin is included in the cul- ture medium. Using growth inhibition as a selective marker, we isolated variant cell lines that are either resistant to insulin or dependent upon insulin for growth. We have studied the effects of insulin on proliferation by using combined genetic and biochemical approaches. Through a series of genetic hy- bridization analyses, we have identified three complementa- tion groups and determined that, in general, insulin-sensitivity is dominant to insulin-resistance. Through analyses of in vitro protein phosphorylation reactions, we have identified a pro- tein of 990 kDa (pp90) whose phosphorylation is a function of at least one of the complementation groups. Although pp90 is not phosphorylated in extracts of insulin-resistant variants, it is phosphorylated in extracts of insulin-sensitive hybrids formed between complementing resistant variants. Insulin it- self exhibits little or no regulation over the phosphorylation of pp90; rather, the ability to phosphorylate pp90 correlates with the ability of cells to respond to insulin. Migration in NaDod- SO4/polyacrylamide gels, solubility characteristics, and diva- lent cation requirements indicate that pp90 is distinct from the 95-kDa P-subunit of the insulin receptor. Both pp90 and its associated phosphoprotein kinase are found in 30,000 x g pel- lets of sonicated cell lysates, whereas a specific pp90 phospho- protein phosphatase activity is found in 30,000 x g superna- tant fractions. Phosphorylation of pp90 occurs at tyrosine and serine residues. Our evidence indicates that the state of phos- phorylation of pp90 is an important determinant in the regula- tion of cellular proliferation by insulin.