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Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing

Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free s...

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Detalles Bibliográficos
Autores principales: Iki, Taichiro, Tschopp, Marie-Aude, Voinnet, Olivier
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Cold Spring Harbor Laboratory Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5393175/
https://www.ncbi.nlm.nih.gov/pubmed/28148824
http://dx.doi.org/10.1261/rna.060434.116
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author Iki, Taichiro
Tschopp, Marie-Aude
Voinnet, Olivier
author_facet Iki, Taichiro
Tschopp, Marie-Aude
Voinnet, Olivier
author_sort Iki, Taichiro
collection PubMed
description Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies. Comparative biochemical analyses of distinct VSRs in the reconstituted assay showed that in all cases tested, VSR interactions with siRNA duplexes inhibited the loading, but not the activity, of antiviral AGO1 and AGO2. Turnip crinkle virus P38 displayed the additional and unique property to bind both synthetic and RNA-dependent-RNA-polymerase-generated long dsRNAs, and inhibited the processing into siRNAs. Single amino acid substitutions in P38 could dissociate dsRNA-processing from AGO-loading inhibition in vitro and in vivo, illustrating dual-inhibitory strategies discriminatively deployed within a single viral protein, which, we further show, are bona fide suppressor functions that evolved independently of the conserved coat protein function of P38.
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spelling pubmed-53931752017-05-02 Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing Iki, Taichiro Tschopp, Marie-Aude Voinnet, Olivier RNA Article Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies. Comparative biochemical analyses of distinct VSRs in the reconstituted assay showed that in all cases tested, VSR interactions with siRNA duplexes inhibited the loading, but not the activity, of antiviral AGO1 and AGO2. Turnip crinkle virus P38 displayed the additional and unique property to bind both synthetic and RNA-dependent-RNA-polymerase-generated long dsRNAs, and inhibited the processing into siRNAs. Single amino acid substitutions in P38 could dissociate dsRNA-processing from AGO-loading inhibition in vitro and in vivo, illustrating dual-inhibitory strategies discriminatively deployed within a single viral protein, which, we further show, are bona fide suppressor functions that evolved independently of the conserved coat protein function of P38. Cold Spring Harbor Laboratory Press 2017-05 /pmc/articles/PMC5393175/ /pubmed/28148824 http://dx.doi.org/10.1261/rna.060434.116 Text en © 2017 Iki et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society http://creativecommons.org/licenses/by/4.0/ This article, published in RNA, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Iki, Taichiro
Tschopp, Marie-Aude
Voinnet, Olivier
Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title_full Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title_fullStr Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title_full_unstemmed Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title_short Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing
title_sort biochemical and genetic functional dissection of the p38 viral suppressor of rna silencing
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5393175/
https://www.ncbi.nlm.nih.gov/pubmed/28148824
http://dx.doi.org/10.1261/rna.060434.116
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