A single amino acid residue change in the P protein of parainfluenza virus 5 elevates viral gene expression.
Parainfluenza virus 5 (PIV5) is a prototypical paramyxovirus. The V/P gene of PIV5 encodes two mRNA species through a process of pseudotemplated insertion of two G residues at a specific site during transcription, resulting in two viral proteins, V and P, whose N termini of 164 amino acid residues are identical. Previously it was reported that mutating six amino acid residues within this identical region results in a recombinant PIV5 (rPIV5-CPI-) that exhibits elevated viral protein expression and induces production of cytokines, such as beta interferon and interleukin 6. Because the six mutations correspond to the shared region of the V protein and the P protein, it is not clear whether the phenotypes associated with rPIV5-CPI- are due to mutations in the P protein and/or mutations in the V protein. To address this question, we used a minigenome system and recombinant viruses to study the effects of mutations on the functions of the P and V proteins. We found that the P protein with six amino acid residue changes (Pcpi-) was more efficient than wild-type P in facilitating replication of viral RNA, while the V protein with six amino acid residue changes (Vcpi-) still inhibits minigenome replication as does the wild-type V protein. These results indicate that elevated viral gene expression in rPIV5-CPI- virus-infected cells can be attributed to a P protein with an increased ability to facilitate viral RNA synthesis. Furthermore, we found that a single amino acid residue change at position 157 of the P protein from Ser (the residue in the wild-type P protein) to Phe (the residue in Pcpi-) is sufficient for elevated viral gene expression. Using mass spectrometry and (33)P labeling, we found that residue S157 of the P protein is phosphorylated. Based on these results, we propose that phosphorylation of the P protein at residue 157 plays an important role in regulating viral RNA replication.
Timani
Khalid A
KA
Center of Molecular Immunology and Infectious Disease, Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, PA 16802, USA.
Sun
Dengyun
D
Sun
Minghao
M
Keim
Celia
C
Lin
Yuan
Y
Schmitt
Phuong Tieu
PT
Schmitt
Anthony P
AP
He
Biao
B
D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D015967:Gene Expression Regulation, Viral; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009154:Mutation; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D018108:Respirovirus / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D014709:Vero Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
D010750:Phosphoproteins; D012333:RNA, Messenger; D012367:RNA, Viral; C502891:V protein, Paramyxovirus; D014764:Viral Proteins