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Journal of Virology, January 2003, p. 258-269, Vol. 77, No. 1
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.1.258-269.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

The RNA Replication Enhancer Element of Tombusviruses Contains Two Interchangeable Hairpins That Are Functional during Plus-Strand Synthesis{dagger}

T. Panavas and P. D. Nagy*

Department of Plant Pathology, University of Kentucky, Lexington, Kentucky 40546

Received 26 June 2002/ Accepted 24 September 2002

Replication of the RNA genomes of tombusviruses, which are small plus-sense RNA viruses of plants, may be regulated by cis-acting elements, including promoters and replication enhancers that are present in the RNA templates. Using a partially purified RNA-dependent RNA polymerase (RdRp) preparation (P. D. Nagy and J. Pogany, Virology 276:279-288, 2000), we demonstrate that the minus-strand templates of tombusviruses contain a replication enhancer, which can upregulate RNA synthesis initiating from the minimal plus-strand initiation promoter by 10- to 20-fold in an in vitro assay. Dissection of the sequence of the replication enhancer element revealed that the two stem-loop structures present within the ~80-nucleotide-long enhancer region have interchangeable roles in upregulating RNA synthesis. The single-stranded sequence located between the two stem-loops also plays an important role in stimulation of RNA synthesis. We also demonstrate that one of the two hairpins, both of which are similar to the hairpin of the minus-strand initiation promoter, can function as a promoter in vitro in the presence of short cytidylate-containing initiation sites. Overall, the in vitro data presented are consistent with previous in vivo results (D. Ray and K. A. White, Virology 256:162-171, 1999) and they firmly establish the presence of a replication enhancer on the minus-stranded RNA of tombusviruses.

* Corresponding author. Mailing address: Department of Plant Pathology, University of Kentucky, S-305, Ag. Sci. Bldg.-N, Lexington, KY 40546. Phone: (859) 257-2889. Fax: (859) 323-1961. E-mail: pdnagy2{at}uky.edu.

{dagger} Publication no. 02-12-115 of the Kentucky Agricultural Experiment Station.

Journal of Virology, January 2003, p. 258-269, Vol. 77, No. 1
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.1.258-269.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Ray, D., White, K. A. (2002). An Internally Located RNA Hairpin Enhances Replication of Tomato Bushy Stunt Virus RNAs. J. Virol. 77: 245-257 [Abstract] [Full Text]  


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