p33-independent activation of a truncated p92 RNA-dependent RNA polymerase of tomato bushy stunt virus in yeast cell-free extract

Judit Pogany, Peter D. Nagy

Research output: Contribution to journalArticlepeer-review

37 Scopus citations

Abstract

Plus-stranded RNA viruses replicate in membrane-bound structures containing the viral replicase complex (VRC). A key component of the VRC is the virally encoded RNA-dependent RNA polymerase (RdRp), which should be activated and incorporated into the VRC after its translation. To study the activation of the RdRp of Tomato bushy stunt virus (TBSV), a small tombusvirus of plants, we used N-terminal truncated recombinant RdRp, which supported RNA synthesis in a cell-free yeast extract-based assay. The truncated RdRp required a cis-acting RNA replication element and soluble host factors, while unlike the full-length TBSV RdRp, the truncated RdRp did not need the viral p33 replication cofactor or cellular membranes for RNA synthesis. Interestingly, the truncated RdRp used 3'-terminal extension for initiation and terminated prematurely at an internal cis-acting element.However, the truncated RdRp could perform de novo initiation on a TBSV plus-strand RNA template in the presenceof the p33 replication cofactor, cellular membranes, and soluble host proteins. Altogether, the data obtained with the truncated RdRp indicate that this RdRp still requires activation, but with the participation of fewer components than with the full-length RdRp, making it suitable for future studies on dissection of the RdRp activation mechanism.

Original languageEnglish
Pages (from-to)12025-12038
Number of pages14
JournalJournal of Virology
Volume86
Issue number22
DOIs
StatePublished - Nov 2012

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

Fingerprint

Dive into the research topics of 'p33-independent activation of a truncated p92 RNA-dependent RNA polymerase of tomato bushy stunt virus in yeast cell-free extract'. Together they form a unique fingerprint.

Cite this