A small peptide inhibits siRNA amplification in plants by mediating autophagic degradation of SGS3/RDR6 bodies.

Selective autophagy mediates specific degradation of unwanted cytoplasmic components to maintain cellular homeostasis. The suppressor of gene silencing 3 (SGS3) and RNA-dependent RNA polymerase 6 (RDR6)-formed bodies (SGS3/RDR6 bodies) are essential for siRNA amplification in planta. However, whether autophagy receptors regulate selective turnover of SGS3/RDR6 bodies is unknown. By analyzing the transcriptomic response to virus infection in Arabidopsis, we identified a virus-induced small peptide 1 (VISP1) composed of 71 amino acids, which harbor a ubiquitin-interacting motif that mediates interaction with autophagy-related protein 8. Overexpression of VISP1 induced selective autophagy and compromised antiviral immunity by inhibiting SGS3/RDR6-dependent viral siRNA amplification, whereas visp1 mutants exhibited opposite effects. Biochemistry assays demonstrate that VISP1 interacted with SGS3 and mediated autophagic degradation of SGS3/RDR6 bodies. Further analyses revealed that overexpression of VISP1, mimicking the sgs3 mutant, impaired biogenesis of endogenous trans-acting siRNAs and up-regulated their targets. Collectively, we propose that VISP1 is a small peptide receptor functioning in the crosstalk between selective autophagy and RNA silencing.

A putative nuclear copper chaperone promotes plant immunity in Arabidopsis.

Copper is essential for many metabolic processes but must be sequestrated by copper chaperones. It is well known that plant copper chaperones regulate various physiological processes. However, the functions of copper chaperones in the plant nucleus remain largely unknown. Here, we identified a putative copper chaperone induced by pathogens (CCP) in Arabidopsis thaliana. CCP harbors a classical MXCXXC copper-binding site (CBS) at its N-terminus and a nuclear localization signal (NLS) at its C-terminus. CCP mainly formed nuclear speckles in the plant nucleus, which requires the NLS and CBS domains. Overexpression of CCP induced PR1 expression and enhanced resistance against Pseudomonas syringae pv. tomato DC3000 compared with Col-0 plants. Conversely, two CRISPR/Cas9-mediated ccp mutants were impaired in plant immunity. Further biochemical analyses revealed that CCP interacted with the transcription factor TGA2 in vivo and in vitro. Moreover, CCP recruits TGA2 to the PR1 promoter sequences in vivo, which induces defense gene expression and plant immunity. Collectively, our results have identified a putative nuclear copper chaperone required for plant immunity and provided evidence for a potential function of copper in the salicylic pathway.

A selective autophagy receptor VISP1 induces symptom recovery by targeting viral silencing suppressors.

Selective autophagy is a double-edged sword in antiviral immunity and regulated by various autophagy receptors. However, it remains unclear how to balance the opposite roles by one autophagy receptor. We previously identified a virus-induced small peptide called VISP1 as a selective autophagy receptor that facilitates virus infections by targeting components of antiviral RNA silencing. However, we show here that VISP1 can also inhibit virus infections by mediating autophagic degradation of viral suppressors of RNA silencing (VSRs). VISP1 targets the cucumber mosaic virus (CMV) 2b protein for degradation and attenuates its suppression activity on RNA silencing. Knockout and overexpression of VISP1 exhibit compromised and enhanced resistance against late infection of CMV, respectively. Consequently, VISP1 induces symptom recovery from CMV infection by triggering 2b turnover. VISP1 also targets the C2/AC2 VSRs of two geminiviruses and enhances antiviral immunity. Together, VISP1 induces symptom recovery from severe infections of plant viruses through controlling VSR accumulation.

RNA In Situ Hybridization of Detecting Cucumber Mosaic Virus in Shoots of Nicotiana benthamiana Plants.

RNA in situ hybridization, a histological technique derived from Southern blotting and northern blotting, has been an important approach in biology studies for many years. In the studies of virus-plant interactions, RNA in situ hybridization provides a direct visualization of viral RNA in host plants. Here, we provide a detailed protocol for viral RNA in situ hybridization that has been successfully used to detect Cucumber mosaic virus genome (CMV) RNAs in shoots of N. benthamiana plants.

Genetic analysis of a Piezo-like protein suppressing systemic movement of plant viruses in Arabidopsis thaliana.

As obligate intracellular phytopathogens, plant viruses must take advantage of hosts plasmodesmata and phloem vasculature for their local and long-distance transports to establish systemic infection in plants. In contrast to well-studied virus local transports, molecular mechanisms and related host genes governing virus systemic trafficking are far from being understood. Here, we performed a forward genetic screening to identify Arabidopsis thaliana mutants with enhanced susceptibility to a 2b-deleted mutant of cucumber mosaic virus (CMV-2aT∆2b). We found that an uncharacterized Piezo protein (AtPiezo), an ortholog of animal Piezo proteins with mechanosensitive (MS) cation channel activities, was required for inhibiting systemic infection of CMV-2aT∆2b and turnip mosaic virus tagged a green fluorescent protein (GFP) (TuMV-GFP). AtPiezo is induced by virus infection, especially in the petioles of rosette leaves. Thus, we for the first time demonstrate the biological function of Piezo proteins in plants, which might represent a common antiviral strategy because many monocot and dicot plant species have a single Piezo ortholog.