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Journal of Virology, June 2005, p. 7812-7818, Vol. 79, No. 12
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.12.7812-7818.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Plant Virus-Derived Small Interfering RNAs Originate Predominantly from Highly Structured Single-Stranded Viral RNAs{dagger}

Attila Molnár,1,{ddagger} Tibor Csorba,1 Lóránt Lakatos,1 Éva Várallyay,1 Christophe Lacomme,2 and József Burgyán1*

Agricultural Biotechnology Center, Plant Biology Institute, P. O. Box 411, H-2101 Gödöllö, Hungary,1 Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, United Kingdom2

Received 8 November 2004/ Accepted 29 January 2005

RNA silencing is conserved in a broad range of eukaryotes and includes the phenomena of RNA interference in animals and posttranscriptional gene silencing (PTGS) in plants. In plants, PTGS acts as an antiviral system; a successful virus infection requires suppression or evasion of the induced silencing response. Small interfering RNAs (siRNAs) accumulate in plants infected with positive-strand RNA viruses and provide specificity to this RNA-mediated defense. We present here the results of a survey of virus-specific siRNAs characterized by a sequence analysis of siRNAs from plants infected with Cymbidium ringspot tombusvirus (CymRSV). CymRSV siRNA sequences have a nonrandom distribution along the length of the viral genome, suggesting that there are hot spots for virus-derived siRNA generation. CymRSV siRNAs bound to the CymRSV p19 suppressor protein have the same asymmetry in strand polarity as the sequenced siRNAs and are imperfect double-stranded RNA duplexes. Moreover, an analysis of siRNAs derived from two other nonrelated positive-strand RNA viruses showed that they displayed the same asymmetry as CymRSV siRNAs. Finally, we show that Tobacco mosaic virus (TMV) carrying a short inverted repeat of the phytoene desaturase (PDS) gene triggered more accumulation of PDS siRNAs than the corresponding antisense PDS sequence. Taken together, these results suggest that virus-derived siRNAs originate predominantly by direct DICER cleavage of imperfect duplexes in the most folded regions of the positive strand of the viral RNA.

* Corresponding author. Mailing address: Agricultural Biotechnology Center, Plant Biology Institute, P. O. Box 411, H-2101 Gödöllö, Hungary. Phone: 36-28-526-155. Fax: 36-28-526-145. E-mail: burgyan{at}abc.hu.

{dagger} Supplemental material for this article may be found at http://jvi.asm.org/.

{ddagger} Present address: The Sainsbury Laboratory, John Innes Centre, Colney, Norwich NR4 7UH, United Kingdom.

Journal of Virology, June 2005, p. 7812-7818, Vol. 79, No. 12
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.12.7812-7818.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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