Neutralization of mobile antiviral small RNA through peroxisomal import

Abstract

In animals, certain viral proteins are targeted to peroxisomes to dampen the antiviral immune response mediated by these organelles. In plants, RNA interference (RNAi) mediated by small interfering (si)RNA is the main antiviral defence mechanism. To protect themselves against the cell- and non-cell autonomous effects of RNAi, viruses produce viral suppressors of RNA silencing (VSR), whose study is crucial to properly understand the biological cycle of plant viruses and potentially find new solutions to control these pathogens. By combining biochemical approaches, cell-specific inhibition of RNAi movement and peroxisome isolation, we show here that one such VSR, the peanut clump virus (PCV)-encoded P15, isolates siRNA from the symplasm by delivering them into the peroxisomal matrix. Infection with PCV lacking this ability reveals that piggybacking of these VSR-bound nucleic acids into peroxisomes potentiates viral systemic movement by preventing the spread of antiviral siRNA. Collectively, these results highlight organellar confinement of antiviral molecules as a novel pathogenic strategy that may have its direct counterpart in other plant and animal viruses.

Artificial import substrates reveal an omnivorous peroxisomal importomer

“…The importomer is unique in its ability to act on folded oligomeric proteins, and a number of proteins are piggyback imported as interfering RNAs and employs a PTS1 signal to transport them into the peroxisome. 41 Together with data presented here, these findings suggest that the importomer is generally compatible with nucleic acids.…”

“…A better understanding of the molecular mechanisms involved during translocation will be a prerequisite for the development of such technologies. Recent work has identified a plant virus encoded protein that binds to antiviral small interfering RNAs and employs a PTS1 signal to transport them into the peroxisome . Together with data presented here, these findings suggest that the importomer is generally compatible with nucleic acids.…”

Immunocapture of dsRNA-bound proteins provides insight into tobacco rattle virus replication complexes and reveals Arabidopsis DRB2 to be a wide-spectrum antiviral effector

“…Similarly to the B2:FP N. benthamiana lines [13] and despite the clear expression of B2:GFP ( Supplementary Figure 1A) , the different lines showed little to moderate developmental phenotypes ( Supplementary Figure 1C ). These were very reminiscent of - but distinct from - those caused by ectopic expression of other RNA silencing suppressors such as P19 or HC-Pro [22, 23]. Such phenotypes may be determined (i) by inhibition of long dsRNA processing into siRNA or (ii) by disruption of miRNA function through their sequestration, or both.…”

“…Immunoprecipitations were performed as previously described [22], with minor modifications. 0.15 g of young rosette leaves were ground in liquid nitrogen, homogenized in a chilled mortar with 1 ml lysis buffer (50 mM Tris–HCl, pH 8, 50 mM NaCl, 1% Triton X-100) containing 1 tablet/50 ml of protease inhibitor cocktail (Roche), transferred to a tube and incubated for 15 min at 4°C on a wheel.…”

Viral suppressors: Combatting RNA silencing

“…RNA silencing suppressors from viruses that belong to different taxonomic groups are unrelated proteins that act with diverse mechanisms in different steps of the RNA silencing pathways, and they are often multifunctional proteins that act in several steps at the same time 2 . In this issue of Nature Plants, Incarbone et al show a completely novel RNA silencing suppression strategy that involves transportation of the mobile silencing signal into the peroxisomes to ruin plant defence 3 ( Fig. 1 Plants defend themselves from invading viruses using RNA silencing.…”