The cucumber mosaic virus 1a protein regulates interactions between the 2b protein and ARGONAUTE 1 while maintaining the silencing suppressor activity of the 2b protein

Abstract

The cucumber mosaic virus (CMV) 2b viral suppressor of RNA silencing (VSR) is a potent counter-defense and pathogenicity factor that inhibits antiviral silencing by titration of short double-stranded RNAs. It also disrupts microRNA-mediated regulation of host gene expression by binding ARGONAUTE 1 (AGO1). But in Arabidopsis thaliana complete inhibition of AGO1 is counterproductive to CMV since this triggers another layer of antiviral silencing mediated by AGO2, de-represses strong resistance against aphids (the insect vectors of CMV), and exacerbates symptoms. Using confocal laser scanning microscopy, bimolecular fluorescence complementation, and co-immunoprecipitation assays we found that the CMV 1a protein, a component of the viral replicase complex, regulates the 2b-AGO1 interaction. By binding 2b protein molecules and sequestering them in P-bodies, the 1a protein limits the proportion of 2b protein molecules available to bind AGO1, which ameliorates 2b-induced disease symptoms, and moderates induction of resistance to CMV and to its aphid vector. However, the 1a protein-2b protein interaction does not inhibit the ability of the 2b protein to inhibit silencing of reporter gene expression in agroinfiltration assays. The interaction between the CMV 1a and 2b proteins represents a novel regulatory system in which specific functions of a VSR are selectively modulated by another viral protein. The finding also provides a mechanism that explains how CMV, and possibly other viruses, modulates symptom induction and manipulates host-vector interactions.

Cucumber Mosaic Virus 1a Protein Interacts with the Tobacco SHE1 Transcription Factor and Partitions between the Nucleus and the Tonoplast Membrane

“…The presence of the 1a protein in speckles in the cytoplasm ( Fig. 4D ) also is consistent with a recent report ( Watt et al, 2020 ), although the presence of the 1a protein in the nucleus ( Fig. 4A, C, and D ) was not observed in that study.…”

“…4F ). These speckles were observed by Watt et al (2020) , who described the speckles as P-bodies. However, those authors did not observe the 1a protein present in the nucleus, although this was not always present here ( Fig.…”

“…4C ) suggests that the 1a protein is able to relocalize some of the SHE1 from the nucleus to the tonoplast membrane. This is reminiscent to the 1a protein being able to relocalize the 2b from both the nucleus and diffused throughout the cytoplasm to the speckles designated P-bodies ( Watt et al, 2020 ). It is not known by what process these relocations take place.…”

“…However, this does not exclude the ability of the individual 1a and 2a proteins from interacting with other proteins and co-localizing to other subcellular structures. In fact, work by Kim et al (2002) showed that phosphorylation of the 2a protein occurred, preventing the interaction of the 1a and 2a proteins, providing a pool of 1a (and 2a) proteins that could interact with the various host proteins, such as those mentioned above, as well as other CMV-encodes proteins, such as the 2b protein ( Watt et al, 2020 ). The presence of the 1a protein in speckles in the cytoplasm ( Fig.…”

Infection of Arabidopsis by cucumber mosaic virus triggers jasmonate‐dependent resistance to aphids that relies partly on the pattern‐triggered immunity factor BAK1

“…The glucosinolate 4MI3M is a metabolite that aphids find distasteful (Kim et al, 2008 ). The 2a protein, one of five proteins encoded by CMV, is responsible for induction of feeding deterrence and concomitant growth inhibition, and this mechanism predominates during infection over a stronger anti‐aphid resistance mechanism due to direct and indirect interactions between two other viral proteins (the 1a and 2b proteins), and the plant Argonaute 1 protein (Rhee et al, 2020 ; Watt et al, 2020 ; Westwood et al, 2013 ). CMV infection induces expression of several genes known to be responsive to microbe‐associated molecular pattern (MAMP) molecules (Westwood et al, 2013 ).…”