Suppression of post-transcriptional gene silencing by a plant viral protein localized in the nucleus

Abstract

A.P.Lucy and H.-S.Guo contributed equally to this work Post-transcriptional gene silencing (PTGS) is a homology-dependent RNA degradation process that may target RNA exclusively in the cytoplasm. In plants, PTGS functions as a natural defense mechanism against viruses. We reported previously that the 2b protein encoded by cucumber mosaic cucumovirus (CMV) is a virulence determinant and a suppressor of PTGS initiation in transgenic Nicotiana benthamiana. By fusion with the green fluorescent protein, we now show that the CMV 2b protein localizes to the nuclei of tobacco suspension cells and whole plants via an arginine-rich nuclear localization signal, 22 KRRRRR 27 . We further demonstrate that the nuclear targeting of the 2b protein is required for the efficient suppression of PTGS, indicating that PTGS may be blocked in the nucleus. In addition, our data indicate that the PTGS suppressor activity is important, but not sufficient, for virulence determination by the 2b protein.

Structural basis for RNA‐silencing suppression by Tomato aspermy virus protein 2b

“…It was speculated that residues near the nuclear localization sites are crucial for RNA suppression by CMV2b (Lucy et al, 2000). Given the sequence similarity between CMV2b and TAV2b, we wonder whether these conserved residues at TAV2b are also crucial for dsRNA binding.…”

“…Recently, CMV2b from the CM95R strain was reported to bind to in vitro-synthesized siRNAs, indicating that CMV2b might suppress RNA silencing by binding directly to siRNAs (Goto et al, 2007). Similar to CMV2b, Tomato aspermy virus (also a member of the cucumovirus family) protein 2b (TAV2b) was reported to suppress RNA silencing (Lucy et al, 2000). Although TAV2b and CMV2b are homologues in their sequence, they vary widely in RNA-silencing suppression and resulting phenotype.…”

Virus-Induced Necrosis Is a Consequence of Direct Protein-Protein Interaction between a Viral RNA-Silencing Suppressor and a Host Catalase

“…The CMV 2b protein is involved in local and systemic movement of CMV and inhibits host defense through RNA silencing (Ding et al, 1995;Ji and Ding, 2001;Soards et al, 2002;Shi et al, 2003) and also acts as a symptom determinant, which is apparently associated with its RNA-silencing suppressor (RSS) activity (Brigneti et al, 1998;Lucy et al, 2000). The 2b protein contains two nuclear localization signals (NLSs) in its N terminus (Lucy et al, 2000;Goto et al, 2007) and has been confirmed to be actually localized in the nucleus in infected tissues; the NLSs of the 2b protein are required for viral symptom induction and RSS activity (Lucy et al, 2000;Lewsey et al, 2009).…”

“…The 2b protein contains two nuclear localization signals (NLSs) in its N terminus (Lucy et al, 2000;Goto et al, 2007) and has been confirmed to be actually localized in the nucleus in infected tissues; the NLSs of the 2b protein are required for viral symptom induction and RSS activity (Lucy et al, 2000;Lewsey et al, 2009). Recently, 2b was found to bind to Argonaute1 (AGO1), necessary for the plant antiviral machinery, and to AGO4, necessary for the DNA methylation process that controls heterochromatin formation and transcription (Zhang et al, 2006;González et al, 2010).…”

Gene silencing: Double-stranded RNA mediated mRNA degradation and gene inactivation

“…The obligatory nuclear localization of the cucumber mosaic virus 2b protein [32] should help to identify steps of PTGS that occur in the nucleus. The host defense function of RNA-mediated silencing is demonstrated by the inc r e a s e d o f s e n s i t i v i t y o f Arabidopsis PTGS mutants to some viruses [21] and the mobilization of transposons in the Mut6 mutant Chlamydomonas [33].…”