In vivo and in vitro phosphorylation of murine mammary tumour virus proteins.

A comparative study of in vitro and in vivo phosphorylation of murine mammary tumour virus, a type Brna virus, is reported. The protein kinase activity associated with murine mammary tumour virus catalysed the in vitro phosphorylation of endogenous virus polypeptides. This kinase activity required a divalent metal cation, a non-ionic detergent, and was stimulated in the presence of dithiothreitol. Exogenous cyclic AMP was not required. The 32P-labelled products of the in vitro reaction were completely sensitive to pronase digestion and the phosphate was attached mainly by phosphomonoester linkage to serine residues. As determined by SDS-polyacrylamide gel electrophoresis, heterogeneous labelling of major and minor virus polypeptides was observed under in vitro conditions. In contrast, the in vivo labelling of type B virus produced by a continuous cell line (MuMT-73), established from pooled mammary adenocarcinomas of Balb/cfC3H mice, demonstrated specific phosphoproteins associated with murine mammary tumour virus. The major phosphorylated proteins were found to have mol. wt. of 18 000 and 12 000 (p18 and p12) after isolation by molecular sieving chromatography and analysis by gel electrophoresis.