Use of double-stranded RNA templates by the tombusvirus replicase in vitro: Implications for the mechanism of plus-strand initiation

Tadas Panavas, Jozsef Stork, Peter D. Nagy

Research output: Contribution to journalArticlepeer-review

14 Scopus citations


Plus-stranded RNA viruses replicate efficiently in infected hosts producing numerous copies of the viral RNA. One of the long-standing mysteries in RNA virus replication is the occurrence and possible role of the double-stranded (ds)RNA formed between minus- and plus-strands. Using the partially purified Cucumber necrosis virus (CNV) replicase from plants and the recombinant RNA-dependent RNA polymerase (RdRp) of Turnip crinkle virus (TCV), in this paper, we demonstrate that both CNV replicase and the related TCV RdRp can utilize dsRNA templates to produce viral plus-stranded RNA in vitro. Sequence and structure of the dsRNA around the plus-strand initiation site had a significant effect on initiation, suggesting that initiation on dsRNA templates is a rate-limiting step. In contrast, the CNV replicase could efficiently synthesize plus-strand RNA on partial dsRNAs that had the plus-strand initiation promoter "exposed", suggesting that the polymerase activity of CNV replicase is strong enough to unwind extended dsRNA regions in the template during RNA synthesis. Based on the in vitro data, we propose that dsRNA forms might have functional roles during tombus- and carmovirus replication and the AU-rich nature of the terminus could be important for opening the dsRNA structure around the plus-strand initiation promoter for tombus- and carmoviruses and possibly many other positive-strand RNA viruses.

Original languageEnglish
Pages (from-to)110-120
Number of pages11
Issue number1
StatePublished - Aug 15 2006

Bibliographical note

Funding Information:
We thank Drs. Andy White and Judit Pogany for critical reading of the manuscript and for very helpful suggestions. This work was supported by the National Institute of Allergy and Infectious Diseases (NIH-NIAID AI05767001A1) and by the Kentucky Tobacco Research and Development Center at the University of Kentucky, awarded to PDN.


  • Cucumber necrosis virus
  • Plus-strand initiation
  • RdRp assay
  • Replication
  • Turnip crinckle virus
  • dsRNA

ASJC Scopus subject areas

  • Virology


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