Interaction between the replicase proteins of Tomato Bushy Stunt virus in vitro and in vivo

K. S. Rajendran, Peter D. Nagy

Research output: Contribution to journalArticlepeer-review

46 Scopus citations


Tomato bushy stunt virus (TBSV), a tombusvirus with a non-segmented, plus-stranded RNA genome, codes for p33 and p92 replicase proteins. The sequence of p33 overlaps with the N-terminal domain of p92, which also contains the signature motifs of RNA-dependent RNA polymerases (RdRps) in its non-overlapping C-terminal portion. In this research, we demonstrate in vitro interactions between p33:p33 and p33:p92 using surface plasmon resonance analysis with purified recombinant p33 and p92. The sequence in p33 involved in the above protein-protein interactions was mapped to the C-terminal region, which also contains an RNA-binding site. Using the yeast two-hybrid assay, we confirmed that two short regions within p33 could promote p33:p33 and p33:p92 interactions in vivo. Mutations in either p33 or p92 within the short regions involved in p33:p33 and p33:p92 interactions decreased the replication of a TBSV defective interfering RNA in yeast, a model host, supporting the significance of these protein interactions in tombusvirus replication.

Original languageEnglish
Pages (from-to)250-261
Number of pages12
Issue number2
StatePublished - Sep 1 2004

Bibliographical note

Funding Information:
We thank Drs. Judit Pogany, Saulius Serva and Jozsef Gal for critical reading of the manuscript and for very helpful suggestions. The authors also thank Drs. A. White and H. Scholthof for providing constructs T100 and pHS-175 and Dr. Serva for his help with using the Anti-His monoclonal antibodies. This work was supported by NSF (MCB0078152) and by the Kentucky Tobacco Research and Development Center at the University of Kentucky, awarded to PDN.


  • Protein-protein interactions
  • RdRp
  • Replication
  • Surface plasmon resonance
  • Two-hybrid assay
  • Yeast host

ASJC Scopus subject areas

  • Virology


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