Phosphorylation of vesicular stomatitis virus M protein: Evidence for a second virion-associated protein serine kinase activity

Abstract

The vesicular stomatitis virus (VSV) NS and M proteins are not only phosphorylated in vivo but are also further modified by the virion-associated protein kinase(s) concomitantly with the in vitro transcription process. Although NS phosphorylation is necessary for this transcription, no function has yet been ascribed for M protein phosphorylation. We show here that all phosphates added to M protein in vitro mapped to the trypsin-sensitive N-terminal basic domain (residues 1–43). The major site(s) (∼93%) corresponded to one or more of three serine residues within the first 17 amino acids. Nearly 1 mol phosphate/mol protein was added in vitro under optimal conditions suggesting that only one of these three candidate serine residues corresponds to the major site. This same M protein domain is thought to play an important role in virus RNA synthesis by inhibiting transcription. We show here that in vitro phosphorylation did not appear to affect this function. Two critical serine residues in the VSV NS protein were previously reported to be phosphorylated during in vitro transcription ( D. Chattopadhyay and A. K. Banerjee, 1987, Cell 49, 407–414 ). The sequence flanking these NS serines is very acidic while that of all three candidate phosphoserines in the M protein is very basic. We therefore predict that at least two distinct serine-specific kinase activities are packaged in virions, one specific for M and one specific for NS.