Vimentin phosphorylation by p37^mos protein kinase in vitro and generation of a 50-kDa cleavage product in v-mos-transformed cells

Previous studies have shown that vimentin, an intermediate filament protein, is reduced in amount in cells acutely infected with Moloney mouse sarcoma virus (Mo-MuSV). In this report, we provide evidence for specific alteration of vimentin in Mo-MuSV-transformed cells and demonstrate specific phosphorylation of vimentin by the p3710^mos protein kinase in vitro. Specificity of the phosphorylation reaction was demonstrated by using viral mos proteins encoded by various isolates of Mo-MuSV and p37^mos produced in yeast. A phosphotransfer domain mutant lacking the ability to autophosphorylate p37^mos failed to phosphorylate vimentin. Similarly, vimentin was not phosphorylated by the temperature-sensitive P85^gag-mos kinase derived from infected cells maintained at the restrictive temperature. In tsl 10 MuSV-transformed NRK cells, vimentin was phosphorylated at both the permissive and nonpermissive temperatures for transformation. However, at the permissive temperature, an altered form of vimentin (about 50 kDa) with a more basic isoelectric point and lower apparent molecular weight was detected. This 50-kDa product was highly phosphorylated as compared to the bulk of the normal 55-kDa form of vimentin. On the basis of its mobility in two-dimensional gels, the 50-kDa form of vimentin should lack the carboxy terminus. This type of alteration could conceivably modulate the function of vimentin filaments in the transformed cell.