title: Identification of a phosphorylation site within the P protein important for mRNA transcription and growth of parainfluenza virus 5.
abstract: The viral RNA-dependent RNA polymerase (vRdRp) of paramyxovirus consists of the large (L) protein and the phosphoprotein (P). P is heavily phosphorylated, and it is thought that the phosphorylation of P plays a role in regulating viral RNA synthesis. However, no phosphorylation site within the P protein in paramyxovirus has been identified as playing a positive role in viral RNA synthesis in virus infection. Using mass spectrometry analysis, the threonine residue at position 286 of P of parainfluenza virus 5 (PIV5) was found phosphorylated. Mutation of T286 to alanine (T286A), aspartic acid (T286D), or glutamic acid (T286E) reduced minigenome activity. Recombinant virus containing a mutation at the T286 position (rPIV5-P-T286A) grew slower than wild-type virus; viral mRNA synthesis and protein expression of rPIV5-P-T286A were delayed. Biochemical studies showed that the binding of NP or L protein with the P mutants or tetramer formation by the mutant P proteins was unaltered from that for wild-type P. While we failed to rescue rPIV5-P-T286E virus, several revertant viruses were obtained. All non-wild-type revertants had mutations at T286 and showed defects in both minigenome activity and viral growth. This is the first time that a phosphorylation site within the P protein in paramyxovirus has been found to play a positive role in viral mRNA synthesis and virus growth.
authors.affiliation: Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
authors.forename: Dengyun
authors.initials: D
authors.lastname: Sun
authors.forename: Priya
authors.initials: P
authors.lastname: Luthra
authors.forename: Pei
authors.initials: P
authors.lastname: Xu
authors.forename: Haeyoung
authors.initials: H
authors.lastname: Yoon
authors.forename: Biao
authors.initials: B
authors.lastname: He
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002417:Cattle; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D015967:Gene Expression Regulation, Viral; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D013058:Mass Spectrometry; D009154:Mutation; D019251:Nucleocapsid / Q000378:metabolism; D010750:Phosphoproteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019213:Rubulavirus / Q000235:genetics* / Q000254:growth & development*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C055859:P protein, Sendai virus; D010750:Phosphoproteins; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase