Rous sarcoma virus-induced phosphorylation of a 50,000-molecular weight cellular protein.

Abstract

Transformation by Rous sarcoma virus (RSV) is mediated by a single virus-encoded polypeptide, pp60src (refs 1–3), which has protein kinase activity specific for tyrosine residues4–7. The identification of the proteins phosphorylated by pp60src is important in understanding the events involved in RSV-mediated transformation. One potential substrate is a 50,000-molecular weight (MW) phosphotyrosine-containing protein (pp50) which is associated in a complex with pp60src and a cellular 90,000-MW protein (pp90; ref. 8) and thus is immunoprecipitated by antibody to pp60src (refs 6, 9, 10). We have performed further studies to determine whether this 50,000-MW protein (pp50) is present in uninfected cells and whether its phosphorylation at tyrosine depends on RSV-induced transformation. Here we demonstrate that a phosphoserine-containing form of pp50 is present in uninfected chicken cells and that on transformation by RSV, a second form of pp50, detectable by two-dimensional analysis of 32P-labelled proteins, contains phosphotyrosine as well as phosphoserine. This phosphorylation of pp50 at tyrosine is not temperature sensitive (ts) in cells infected with mutant viruses containing a ts defect in the src gene. Previous results8 have suggested that the temperature stability of this phosphorylation might be a result of the strong interaction between the mutant pp60src protein and pp50.