Cucumber mosaic virus (CMV) 2b suppresses RNA silencing primarily through the binding of double-stranded RNA (dsRNA) of varying sizes. However, the biologically active form of 2b remains elusive. Here, we demonstrate that the single and double alanine substitution mutants in the N-terminal 15th leucine and 18th methionine of CMV 2b exhibit drastically attenuated virulence in wild-type plants, but are efficiently rescued in mutant plants defective in RNA-dependent RNA polymerase 6 (RDR6) and Dicer-like 4 (DCL4). Moreover, the transgenic plants of 2b, but not 2blm (L15A/M18A), rescue the high infectivity of CMV-Δ2b through the suppression of antiviral silencing. L15A, M18A or both weaken 2b suppressor activity on local and systemic transgene silencing. In contrast with the high affinity of 2b to short and long dsRNAs, 2blm is significantly compromised in 21-bp duplex small interfering RNA (siRNA) binding ability, but maintains a strong affinity for long dsRNAs. In cross-linking assays, 2b can form dimers, tetramers and oligomers after treatment with glutaraldehyde, whereas 2blm only forms dimers, rather than tetramers and oligomers, in vitro. Together, these findings suggest that L15 and M18 of CMV 2b are required for high affinity to ds-siRNAs and oligomerization activity, which are essential for the suppression activity of 2b on antiviral silencing.

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.

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.

Controlled release urea (CRU) is considered to enhance crop yields while alleviating negative environmental problems caused by the hazardous gas emissions that are associated with high concentrations of ammonium (NH4+) and nitrate (NO3−) in black soils. Short-term effects of sulfur-coated urea (SCU) and polyurethane-coated urea (PCU), compared with conventional urea, on NO3− and NH4+ in black soils were studied through the buried bag experiment conducted in an artificial climate chamber. We also investigated nitrogen (N) release kinetics of CRU and correlations between the cumulative N release rate and concentrations of NO3− and NH4+. CRU can reduce concentrations of NO3− and NH4+, and PCU was more effective in maintaining lower soil NO3−/NH4+ ratios than SCU and U. Parabolic equation could describe the kinetics of NO3− and NH4+ treated with PCU. The Elovich equation could describe the kinetics of NO3− and NH4+ treated with SCU. The binary linear regression model was established to predict N release from PCU because of significant correlations between the cumulative N release rate and concentrations of NO3− and NH4+. These results provided a methodology and data support for characterizing and predicting the N release from PCU in black soils.

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, functions of copper chaperones in plant nucleus remain largely unknown. Here, we identified a putative Copper Chaperone induced by Pathogens (CCP) in Arabidopsis thaliana. CCP harbors a classical MXCXXC Cu binding site (CBS) at its N terminus and a nuclear localization signal (NLS) at the 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 (Pst 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 potential function of Cu in the SA pathway.

Signaling by the evolutionarily conserved mitogen-activated protein kinase or extracellular signal-regulated kinase (MAPK/ERK) plays critical roles in converting extracellular stimuli into immune responses. However, whether MAPK/ERK signaling induces virus immunity by directly phosphorylating viral effectors remains largely unknown. Barley yellow striate mosaic virus (BYSMV) is an economically important plant cytorhabdovirus that is transmitted by the small brown planthopper (SBPH, Laodelphax striatellus) in a propagative manner. Here, we found that the barley (Hordeum vulgare) MAPK MPK3 (HvMPK3) and the planthopper ERK (LsERK) proteins interact with the BYSMV nucleoprotein (N) and directly phosphorylate N protein primarily on serine 290. Overexpression of HvMPK3 inhibited BYSMV infection, whereas barley plants treated with the MAPK pathway inhibitor U0126 displayed greater susceptibility to BYSMV. Moreover, knockdown of LsERK promoted virus infection in SBPHs. A phosphomimetic mutant of the N Ser290 (S290D) completely abolished virus infection because of impaired self-interaction of BYSMV N and formation of unstable N–RNA complexes. Altogether, our results demonstrate that the conserved MAPK and ERK directly phosphorylate the viral nucleoprotein to trigger immunity against cross-kingdom infection of BYSMV in host plants and its insect vectors.

No abstract

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.