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

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

The Arabidopsis RLCK VI_A2 Kinase Controls Seedling and Plant Growth in Parallel with Gibberellin

The plant-specific receptor-like cytoplasmic kinases (RLCKs) form a large, poorly characterized family. Members of the RLCK VI_A class of dicots have a unique characteristic: their activity is regulated by Rho-of-plants (ROP) GTPases. The biological function of one of these kinases was investigated using a T-DNA insertion mutant and RNA interference. Loss of RLCK VI_A2 function resulted in restricted cell expansion and seedling growth. Although these phenotypes could be rescued by exogenous gibberellin, the mutant did not exhibit lower levels of active gibberellins nor decreased gibberellin sensitivity. Transcriptome analysis confirmed that gibberellin is not the direct target of the kinase; its absence rather affected the metabolism and signalling of other hormones such as auxin. It is hypothesized that gibberellins and the RLCK VI_A2 kinase act in parallel to regulate cell expansion and plant growth. Gene expression studies also indicated that the kinase might have an overlapping role with the transcription factor circuit (PIF4-BZR1-ARF6) controlling skotomorphogenesis-related hypocotyl/cotyledon elongation. Furthermore, the transcriptomic changes revealed that the loss of RLCK VI_A2 function alters cellular processes that are associated with cell membranes, take place at the cell periphery or in the apoplast, and are related to cellular transport and/or cell wall reorganisation

Argonaute 2 Controls Antiviral Activity against Sweet Potato Mild Mottle Virus in Nicotiana benthamiana

This article belongs to the Special Issue Plant Defense Responses against Viruses.RNA silencing is a sequence specific post-transcriptional mechanism regulating important biological processes including antiviral defense in plants. Argonaute (AGO) proteins, the catalytic subunits of the silencing complexes, are loaded with small RNAs to execute the sequence specific RNA cleavage or translational inhibition. Plants encode several AGO proteins and a few of them, especially AGO1 and AGO2, have been shown to be required for antiviral silencing. Previously, we have shown that the P1 protein of the sweet potato mild mottle virus (SPMMV) suppresses the primary RNA silencing response by inhibiting AGO1. To analyze the role of AGO2 in antiviral defense against the SPMMV, we performed a comparative study using a wild type and ago2−/− mutant Nicotiana benthamiana. Here we show that the AGO2 of N. benthamiana attenuates the symptoms of SPMMV infection. Upon SPMMV infection the levels of AGO2 mRNA and protein are greatly increased. Moreover, we found that AGO2 proteins are loaded with SPMMV derived viral small RNAs as well as with miRNAs. Our results indicate that AGO2 protein takes over the place of AGO1 to confer antiviral silencing. Finally, we provide a plausible explanation for the AGO2 mediated recovery of an SPMMV-infected sweet potato.This study was supported by grants of the Hungarian Ministry of Technology and Innovation by grant numbers GINOP-2.3.2-15-2016-00015 and GINOP-2.3.2-15-2016-00032). Additional financial support at CRAG was provided by Generalitat de Catalunya CERCA institution and by the Ministry of Science and Innovation-State Research Agency (AEI, Spain) through the “Severo Ochoa Programme for Centres of Excellence in R&D” SEV-2015-0533 and CEX2019-000902-S and the grant PID2019-105692RB-100 to JLM.Peer reviewe

The ROP2 GTPase Participates in Nitric Oxide (NO)-Induced Root Shortening in Arabidopsis

Nitric oxide (NO) is a versatile signal molecule that mediates environmental and hormonal signals orchestrating plant development. NO may act via reversible S-nitrosation of proteins during which an NO moiety is added to a cysteine thiol to form an S-nitrosothiol. In plants, several proteins implicated in hormonal signaling have been reported to undergo S-nitrosation. Here, we report that the Arabidopsis ROP2 GTPase is a further potential target of NO-mediated regulation. The ROP2 GTPase was found to be required for the root shortening effect of NO. NO inhibits primary root growth by altering the abundance and distribution of the PIN1 auxin efflux carrier protein and lowering the accumulation of auxin in the root meristem. In rop2-1 insertion mutants, however, wild-type-like root size of the NO-treated roots were maintained in agreement with wild-type-like PIN1 abundance in the meristem. The ROP2 GTPase was shown to be S-nitrosated in vitro, suggesting that NO might directly regulate the GTPase. The potential mechanisms of NO-mediated ROP2 GTPase regulation and ROP2-mediated NO signaling in the primary root meristem are discussed