Phosphorylation of Ser232 directly regulates the transcriptional activity of the P protein of human respiratory syncytial virus: phosphorylation of Ser237 may play an accessory role.
The phosphoprotein P of human respiratory syncytial virus (RSV) was expressed in eukaryotic cells in phosphorylated form. Site-directed mutagenesis of the recombinant protein established Ser232 as the major site of phosphorylation in vivo. Phosphorylation of bacterially made P protein in vitro by purified casein kinase II (CKII) resulted in the phosphorylation of Ser237, whereas mainly Ser232 was phosphorylated by a crude cell extract. The P kinase activity in the cell extract exhibited properties characteristic of CKII. While the Ser232,237 to Ala double mutant was nearly completely defective for phosphorylation and transcription, phosphorylation at Ser232, through the use of appropriate P mutant or kinase, activated P protein. Phosphorylation of Ser237 restored activity only to the extent it facilitated phosphorylation of Ser232. Phosphate groups of P protein in RSV-infected cells were highly stable; inhibitors of protein serine phosphatases had no effect on the intracellular turnover of the phosphates. Highly purified viral polymerase L was transcriptionally active but devoid of P protein kinase activity. Thus, CKII-mediated phosphorylation of Ser232 appears to be the primary regulator of P protein activity while phosphorylation of Ser237 may be involved in a modulatory role under certain conditions.
D001665:Binding Sites; D005057:Eukaryotic Cells; D015707:HN Protein; D006801:Humans; D010710:Phosphates / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012136:Respiratory Syncytial Viruses / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D015707:HN Protein; D010710:Phosphates; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C116204:attachment protein G; D012694:Serine; D011494:Protein Kinases