Phosphorylation of v-mos Ser 47 by the mitotic form of p34^cdc2

P85^gag-mos mos is hyperphosphorylated during mitosis in normal rat kidney (NRK) cells transformed by Moloney murine sarcoma virus ts11O. We now report that P85^gag-mos is phosphorylated in vitro by the mitotic form of the cdc2 kinase (p34^cdc2, known as M-phase kinase) derived from virus-transformed cells. The major site of P85^gag-mos phosphorylation by the M-phase kinase in vitro lies within the amino-terminal portion of the viral mos protein sequence spanning residues 45-53, as determined by tryptic peptide mapping. A synthetic peptide corresponding to amino acids 37-55 of v-mos was specifically phosphorylated by the M-phase kinase, whereas v-mos peptides either lacking Ser 47 or substituted with Ala at residue 47 were not phosphorylated. Protein sequencing analyses established that the M-phase kinase specifically phosphorylates Ser 47. Tryptic phosphopeptide mapping of the in vivo-phosphorylated gag-mos protein from mitotic cells indicated that the 45-53 v-mos region was also phosphorylated within mitotic cells. These findings demonstrate that the M-phase kinase phosphorylates the viral mos protein at Ser 47. These results were unexpected in view of earlier reports regarding cdc2 kinase activation/stabilization by the c-mos kinase in maturing oocytes.