Association of a M(r) 90,000 phosphoprotein with protein kinase PKR in cells exhibiting enhanced phosphorylation of translation initiation factor eIF-2 alpha and premature shutoff of protein synthesis after infection with gamma 134.5- mutants of herpes simplex virus 1.
The protein encoded by the gamma 134.5 gene of herpes simplex virus precludes premature shutoff of protein synthesis in human cells triggered by stress associated with onset of viral DNA synthesis. The carboxyl terminus of the protein is essential for this function. This report indicates that the shutoff of protein synthesis is not due to mRNA degration because mRNA from wild-type or gamma 134.5- virus-infected cells directs protein synthesis. Analyses of the posttranslational modifications of translation initiation factor eIF-2 showed the following: (i) eIF-2 alpha was selectively phosphorylated by a kinase present in ribosome-enriched fraction of cells infected with gamma 134.5- virus. (ii) Endogenous eIF-2 alpha was totally phosphorylated in cells infected with gamma 134.5- virus or a virus lacking the 3' coding domain of the gamma 134.5 gene but was not phosphorylated in mock-infected or wild-type virus-infected cells. (iii) Immune precipitates of the PKR kinase that is responsible for regulation of protein synthesis of some cells by phosphorylation of eIF-2 alpha yielded several phosphorylated polypeptides. Of particular significance were two observations. First, phosphorylation of PKR kinase was elevated in all infected cells relative to the levels in mock-infected cells. Second, the precipitates from lysates of cells infected with gamma 134.5- virus or a virus lacking the 3' coding domain of the gamma 134.5 gene contained an additional labeled phosphoprotein of M(r) 90,000 (p90). This phosphoprotein was present in only trace amounts in the immunoprecipitate from cells infected with wild-type virus or mutants lacking a portion of the 5' domain of gamma 134.5. We conclude that in the absence of gamma 134.5 protein, PKR kinase complexes with the p90 phosphoprotein and shuts off protein synthesis by phosphorylation of the alpha subunit of translation initiation factor eIF-2.
D000818:Animals; D002478:Cells, Cultured; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D005814:Genes, Viral; D018259:Herpesvirus 1, Human / Q000235:genetics*; D006801:Humans; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics*; D019892:eIF-2 Kinase
D015852:Eukaryotic Initiation Factor-2; D010750:Phosphoproteins; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins; C066469:gamma 34.5 protein, Human herpesvirus 1; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase