A viral suppressor of RNA silencing inhibits ARGONAUTE 1 function by precluding target RNA binding to pre-assembled RISC

Nucleic Acids Res. 2017 Jul 27;45(13):7736-7750. doi: 10.1093/nar/gkx379.

Abstract

In most eukaryotes, RNA silencing is an adaptive immune system regulating key biological processes including antiviral defense. To evade this response, viruses of plants, worms and insects have evolved viral suppressors of RNA silencing proteins (VSRs). Various VSRs, such as P1 from Sweet potato mild mottle virus (SPMMV), inhibit the activity of RNA-induced silencing complexes (RISCs) including an ARGONAUTE (AGO) protein loaded with a small RNA. However, the specific mechanisms explaining this class of inhibition are unknown. Here, we show that SPMMV P1 interacts with AGO1 and AGO2 from Arabidopsis thaliana, but solely interferes with AGO1 function. Moreover, a mutational analysis of a newly identified zinc finger domain in P1 revealed that this domain could represent an effector domain as it is required for P1 suppressor activity but not for AGO1 binding. Finally, a comparative analysis of the target RNA binding capacity of AGO1 in the presence of wild-type or suppressor-defective P1 forms revealed that P1 blocks target RNA binding to AGO1. Our results describe the negative regulation of RISC, the small RNA containing molecular machine.

MeSH terms

  • Arabidopsis / genetics*
  • Arabidopsis / metabolism*
  • Arabidopsis / virology
  • Arabidopsis Proteins / antagonists & inhibitors*
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism
  • Argonaute Proteins / antagonists & inhibitors*
  • Argonaute Proteins / genetics
  • Argonaute Proteins / metabolism
  • Models, Biological
  • Mutagenesis, Site-Directed
  • Plants, Genetically Modified
  • Potyviridae / genetics
  • Potyviridae / metabolism
  • Potyviridae / pathogenicity
  • RNA Interference
  • RNA, Plant / genetics
  • RNA, Plant / metabolism*
  • RNA-Induced Silencing Complex / genetics
  • RNA-Induced Silencing Complex / metabolism*
  • Tobacco / genetics
  • Tobacco / metabolism
  • Viral Proteins / chemistry
  • Viral Proteins / genetics
  • Viral Proteins / metabolism
  • Zinc Fingers / genetics

Substances

  • AGO1 protein, Arabidopsis
  • Arabidopsis Proteins
  • Argonaute Proteins
  • P1 protein, potyvirus
  • RNA, Plant
  • RNA-Induced Silencing Complex
  • Viral Proteins
  • argonaute 2 protein, Arabidopsis