Heterologous expression of plant virus genes that suppress post-transcriptional gene silencing results in suppression of RNA interference in Drosophila cells

Brian Reavy,Tomas Canto,Stuart A Macfarlane,Sheila Dawson

doi:10.1186/1472-6750-4-18

Abstract

BackgroundRNA interference (RNAi) in animals and post-transcriptional gene silencing (PTGS) in plants are related phenomena whose functions include the developmental regulation of gene expression and protection from transposable elements and viruses. Plant viruses respond by expressing suppressor proteins that interfere with the PTGS system.ResultsHere we demonstrate that both transient and constitutive expression of the Tobacco etch virus HC-Pro silencing suppressor protein, which inhibits the maintenance of PTGS in plants, prevents dsRNA-induced RNAi of a lacZ gene in cultured Drosophila cells. Northern blot analysis of the RNA present in Drosophila cells showed that HC-Pro prevented degradation of lacZ RNA during RNAi but that there was accumulation of the short (23nt) RNA species associated with RNAi. A mutant HC-Pro that does not suppress PTGS in plants also does not affect RNAi in Drosophila. Similarly, the Cucumber mosaic virus 2b protein, which inhibits the systemic spread of PTGS in plants, does not suppress RNAi in Drosophila cells. In addition, we have used the Drosophila system to demonstrate that the 16K cysteine-rich protein of Tobacco rattle virus, which previously had no known function, is a silencing suppressor protein.ConclusionThese results indicate that at least part of the process of RNAi in Drosophila and PTGS in plants is conserved, and that plant virus silencing suppressor proteins may be useful tools to investigate the mechanism of RNAi.

Highlights

RNA interference (RNAi) in animals and post-transcriptional gene silencing (PTGS) in plants are related phenomena whose functions include the developmental regulation of gene expression and protection from transposable elements and viruses
Genetic studies have identified a number of the proteins that are involved in these processes in Neurospora crassa [10,11,12], Caenorhabditis elegans [13,14,15] and Arabidopsis thaliana [16,17,18]. dsRNA has been shown to induce RNAi in cultured Drosophila cells [19,20,21,22] and biochemical studies of this system have revealed the involvement of two distinct activities; an RNA-induced silencing
A survey of a small number of other different plant viruses showed that the comovirus Cowpea mosaic virus, the geminivirus African cassava mosaic virus, the potexvirus Narcissus mosaic virus, the tobamovirus TMV, the sobemovirus Rice yellow mottle virus (RYMV), the tombusvirus Tomato bushy stunt virus (TBSV) and the tobravirus Tobacco rattle virus (TRV) were able to suppress GFP silencing [24]

Summary

Introduction

RNA interference (RNAi) in animals and post-transcriptional gene silencing (PTGS) in plants are related phenomena whose functions include the developmental regulation of gene expression and protection from transposable elements and viruses. Plant viruses respond by expressing suppressor proteins that interfere with the PTGS system. RNA interference (RNAi) is a process in which the introduction of dsRNAs into cells leads to inactivation of expression of genes containing homologous sequences by sequence-specific degradation of mRNA (for reviews see [1,2]). DsRNA has been shown to induce RNAi in cultured Drosophila cells [19,20,21,22] and biochemical studies of this system have revealed the involvement of two distinct activities; an RNA-induced silencing (page number not for citation purposes). Plant virus-encoded silencing suppressors may target different components of the PTGS system. We have investigated the effects on RNAi in cultured Drosophila cells of expression of some silencing suppressors from plant viruses

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Conclusion