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Plant and Cell Physiology Advance Access originally published online on June 13, 2007
Plant and Cell Physiology 2007 48(7):1050-1060; doi:10.1093/pcp/pcm074
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© The Author 2007. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oxfordjournals.org

Characterization of Silencing Suppressor 2b of Cucumber Mosaic Virus Based on Examination of its Small RNA-Binding Abilities

Kazunori Goto1, Takashi Kobori2, Yoshitaka Kosaka2, Tomohide Natsuaki3 and Chikara Masuta1,*

1Cell Biology and Manipulation Laboratory, Graduate School of Agriculture, Hokkaido University, Sapporo, 060-8589 Japan
2Kyoto Prefectural Institute of Agricultural Biotechnology, Kyoto, 619-0244 Japan
3Laboratory of Plant Pathology, Faculty of Agriculture, Utsunomiya University, Utsunomiya, 321-8505 Japan

*Corresponding author: E-mail, masuta{at}res.agr.hokudai.ac.jp; Fax, +81-11-706-2483.


  Abstract

Double-stranded (ds) RNAs and imperfect hairpin RNAs of endogenous genes trigger post-transcriptional gene silencing (PTGS) and are cleaved by a Dicer-like nuclease into small interfering RNAs (siRNAs) and microRNs (miRNAs), respectively. Such small RNAs (siRNAs and miRNAs) then guide an RNA-induced silencing complex (RISC) for sequence-specific RNA degradation. While PTGS serves as an antiviral defense in plants, many plant viruses encode suppressors as a counter defense. Here we demonstrate that the PTGS suppressor (2b) of a severe strain (CM95R) of cucumber mosaic virus (CMV) can bind to in vitro synthesized siRNAs and even to long dsRNAs to a lesser extent. However, the 2b suppressor weakly bound to a miRNA (miR171) duplex in contrast to another small RNA-binding suppressor, p19 of tombusvirus that can effectively bind miRNAs. Because the 2b suppressor of an attenuated strain of CMV (CM95), which differs in a single amino acid from the 2b of CM95R, could barely bind siRNAs, we hypothesized that the weak suppressor activity of the attenuated strain resulted from a loss of the siRNA-binding property of 2b via a single amino acid change. Here we consider that 2b interferes with the PTGS pathway by directly binding siRNAs (or long dsRNA).

Keywords: Cucumber mosaic virus - PTGS - siRNA - Viral suppressor

Abbreviations: AGO1, Argonaute; ARR, arginine-rich region; CHS, chalcone synthase; CMV, cucumber mosaic virus; dpi, days post-infiltration; GFP, green fluorescent protein; GUS, ß-glucuronidase; IR, inverted repeat; LMW, low molecular weight; LUC, luciferase; miRNA, microRNA; NLS, nuclear localization signal; PTGS, post-transcriptional gene silencing; RISC, RNA-induced silencing complex; siRNA, small interfering RNA; TBSV, tomato bushy stunt virus.

(Received February 12, 2007; Accepted June 7, 2007)
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