P7 and P8 proteins of High Plains wheat mosaic virus, a negative-strand RNA virus, employ distinct mechanisms of RNA silencing suppression

Abstract

High Plains wheat mosaic virus (genus Emaravirus), an octapartite negative-sense RNA virus, encodes two RNA silencing suppressors, P7 and P8. In this study, we found that P7 and P8 efficiently delayed the onset of dsRNA-induced transitive pathway of RNA silencing. Electrophoretic mobility shift assays (EMSA) revealed that only P7 protected long dsRNAs from dicing in vitro and bound weakly to 21- and 24-nt PTGS-like ds-siRNAs. In contrast, P8 bound strongly and relatively weakly to 21- and 24-nt ds-siRNAs, respectively, suggesting size-specific binding. In EMSA, neither protein bound to 180-nt and 21-nt ssRNAs at detectable levels. Sequence analysis revealed that P7 contains a conserved GW motif. Mutational disruption of this motif resulted in loss of suppression of RNA silencing and pathogenicity enhancement, and failure to complement the silencing suppression-deficient wheat streak mosaic virus. Collectively, these data suggest that P7 and P8 proteins utilize distinct mechanisms to overcome host RNA silencing for successful establishment of systemic infection in planta.