Determination of a phosphorylation site in Nipah virus nucleoprotein and its involvement in virus transcription

Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639, Japan.
Journal of General Virology (Impact Factor: 3.18). 05/2011; 92(Pt 9):2133-41. DOI: 10.1099/vir.0.032342-0
Source: PubMed

ABSTRACT Many viruses use their host's cellular machinery to regulate the functions of viral proteins. The phosphorylation of viral proteins is known to play a role in genome transcription and replication in paramyxoviruses. The paramyxovirus nucleoprotein (N), the most abundant protein in infected cells, is a component of the N-RNA complex and supports the transcription and replication of virus mRNA and genomic RNA. Recently, we reported that the phosphorylation of measles virus N is involved in the regulation of viral RNA synthesis. In this study, we report a rapid turnover of phosphorylation in the Nipah virus N (NiV-N). The phosphorylated NiV-N was hardly detectable in steady-state cells, but was detected after inhibition of cellular protein phosphatases. We identified a phosphorylated serine residue at Ser451 of NiV-N by peptide mass fingerprinting by electrospray ionization-quadrupole time-of-flight mass spectrometry. In the NiV minigenome assay, using luciferase as a reporter gene, the substitution of Ser451 for alanine in NiV-N resulted in a reduction in luciferase activity of approximately 45 % compared with the wild-type protein. Furthermore, the substitution of Ser451 for glutamic acid, which mimics a phosphoserine, led to a more significant decrease in luciferase activity - approximately 81 %. Northern blot analysis showed that both virus transcription and replication were reduced by these mutations. These results suggest that a rapid turnover of the phosphorylation of NiV-N plays an important role in virus transcription and replication.

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Available from: Hiroko Kozuka-Hata, Sep 25, 2015
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    • "The N and the P proteins of NiV and HeV were shown to form homologous and heterologous N–P complexes, and have more than one P-binding site on the N protein (Chan et al. 2004). Huang et al. 2011 demonstrated transient phosphorylation at amino acid position 451 of the N protein, which when mutated to an alanine or glutamate significantly reduced NiV minigenome replication. "
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