Inhibition of vesicular stomatitis virus RNA synthesis by protein hyperphosphorylation

Theresa Li Yun Chang, Carol S. Reiss, Alice S. Huang

Research output: Contribution to journalArticlepeer-review

Abstract

Vesicular stomatitis virus (VSV) RNA synthesis requires the template nucleocapsid, the polymerase (L) protein, and the cofactor phosphorylated (P/NS) protein. To determine whether the degree of phosphorylation regulated VSV RNA synthesis, infected Chinese hamster ovary cells were treated with okadaic acid (OKA), a serine/threonine phosphatase inhibitor. OKA reduced viral penetration and uncoating but had little or no effect on primary transcription or viral protein synthesis. However, approximately 80% of total viral RNA synthesis was inhibited when 2 μM or more OKA was added to infected cells after viral uncoating had taken place. Analysis of proteins and RNA species in infected cells labeled with 32P showed that OKA led to hyperphosphorylation of two viral phosphoproteins, the P/NS protein and matrix protein (M), resulting in inhibition of full-length RNA synthesis and subsequent secondary transcription. Pulse-chase experiments demonstrated that the hyperphosphorylated P/NS species was converted rapidly from the less phosphorylated form. Hyperphosphorylated P/NS as well as the less phosphorylated form, but not M, were found to be associated with nucleocapsids isolated from cytoplasmic extracts. These results suggest that phosphorylation played an important role in the regulation between viral transcription and viral RNA replication as well as the turning off of RNA replication. Thus, phosphatase inhibitors promise to be a valuable tool for dissecting the regulatory mechanisms involving phosphorylated viral proteins.

Original languageEnglish (US)
Pages (from-to)4980-4987
Number of pages8
JournalJournal of virology
Volume68
Issue number8
DOIs
StatePublished - Aug 1994

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

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