Viral silencing suppressors and cellular proteins partner with plant RRP6-like exoribonucleases.

RNA silencing and RNA decay are functionally interlaced, regulate gene expression and play a pivotal role in antiviral responses. As a counter-defensive strategy, many plant and mammalian viruses encode suppressors which interfere with both mechanisms. However, the protein interactions that connect these pathways remain elusive. Previous work reported that RNA silencing suppressors from different potyviruses, together with translation initiation factors EIF(iso)4E, interacted with the C-terminal region of the tobacco exoribonuclease RRP6-like 2, a component of the RNA decay exosome complex. Here, we investigate whether other viral silencing suppressors and cellular proteins might also bind RRP6-like exoribonucleases. A candidate search approach based on yeast two-hybrid protein interaction assays showed that three other unrelated viral suppressors, two from plant viruses and one from a mammalian virus, bound the C-terminus of the tobacco RRP6-like 2, the full-length of the Arabidopsis RRP6L1 protein and its C-terminal region. In addition, RRP6-like proteins were found to interact with members of the cellular double-stranded RNA-binding protein (DRB) family involved in RNA silencing. The C-terminal regions of RRP6L proteins are engaged in homotypic and heterotypic interactions and were predicted to be disordered. Collectively, these results suggest a protein interaction network that connects components of RNA decay and RNA silencing that is targeted by viral silencing suppressors.

TMV induces RNA decay pathways to modulate gene silencing and disease symptoms.

RNA decay pathways comprise a combination of RNA degradation mechanisms that are implicated in gene expression, development and defense responses in eukaryotes. These mechanisms are known as the RNA Quality Control or RQC pathways. In plants, another important RNA degradation mechanism is the post-transcriptional gene silencing (PTGS) mediated by small RNAs (siRNAs). Notably, the RQC pathway antagonizes PTGS by preventing the entry of dysfunctional mRNAs into the silencing pathway to avoid global degradation of mRNA by siRNAs. Viral transcripts must evade RNA degrading mechanisms, thus viruses encode PTGS suppressor proteins to counteract viral RNA silencing. Here, we demonstrate that tobacco plants infected with TMV and transgenic lines expressing TMV MP and CP (coat protein) proteins (which are not linked to the suppression of silencing) display increased transcriptional levels of RNA decay genes. These plants also showed accumulation of cytoplasmic RNA granules with altered structure, increased rates of RNA decay for transgenes and defective transgene PTGS amplification. Furthermore, knockdown of RRP41 or RRP43 RNA exosome components led to lower levels of TMV accumulation with milder symptoms after infection, several developmental defects and miRNA deregulation. Thus, we propose that TMV proteins induce RNA decay pathways (in particular exosome components) to impair antiviral PTGS and this defensive mechanism would constitute an additional counter-defense strategy that lead to disease symptoms.