RCY1, an Arabidopsis thaliana RPP8/HRT family resistance gene,conferring resistance to cucumber mosaic virus requires salicylic acid,ethylene and a novel signal transduction mechanism

The dominant locus, RCY1, in the Arabidopsis thaliana ecotype C24 confers resistance to the yellow strain of cucumber mosaic virus (CMV-Y). The RCY1 locus was mapped to a 150-kb region on chromosome 5. Sequence comparison of this region from C24 and a CMV-Y-susceptible C24 mutant predicts that the RCY1 gene encodes a 104-kDa CC-NBS-LRR-type protein. The RCY1 gene from C24, when expressed in the susceptible ecotype Wassilewskija (Ws), restricted the systemic spread of virus. RCY1 is allelic to the resistance genes RPP8 from the ecotype Landsberg erecta and HRT from the ecotype Dijon-17, which confer resistance to Peronospora parasitica biotype Emco5 and turnip crinkle virus (TCV), respectively. Examination of RCY1 plants defective in salicylic acid (SA), jasmonic acid (JA) and ethylene signaling revealed a requirement for SA and ethylene signaling in mounting a resistance response to CMV-Y. The RCY1 nahG etr1 double mutants exhibited an intermediate level of susceptibility to CMV-Y, compared to the resistant ecotype C24 and the susceptible ecotypes Columbia and Nossen. This suggests that in addition to SA and ethylene, a novel signaling mechanism is associated with the induction of resistance in CMV-Y-infected C24 plants. Moreover, our results suggest that the signaling pathways downstream of the RPP8, HRT, and RCY1 have evolved independently.     

Nicotiana benthamiana asparagine synthetase associates with IP-L and confers resistance against tobacco mosaic virus via the asparagine-induced salicylic acid signalling pathway

Asparagine synthetase is a key enzyme that catalyses the conversion of amide groups from glutamine or ammonium to aspartate, which leads to the generation of asparagine. However, the role of asparagine synthetase in plant immunity remains largely unknown. Here, we identified a Nicotiana benthamiana asparagine synthetase B (NbAS-B) that associates with tomato mosaic virus coat protein-interacting protein L (IP-L) using the yeast two-hybrid assay and examined its role in tobacco mosaic virus (TMV) resistance. The association of IP-L with NbAS-B was further confirmed by in vivo co-immunoprecipitation, luciferase complementation imaging, and bimolecular fluorescence complementation assays. IP-L and NbAS-B interact in the nucleus and cytosol and IP-L apparently stabilizes NbAS-B, thus enhancing its accumulation. The expressions of IP-L and NbAS-B are continuously induced on TMV-green fluorescent protein (GFP) infection. Co-silencing of IP-L and NbAS-B facilitates TMV-GFP infection. Overexpression of NbAS-B in tobacco reduces TMV-GFP infection by significantly improving the synthesis of asparagine. Furthermore, the external application of asparagine significantly inhibits the infection of TMV-GFP by activating the salicylic acid signalling pathway. These findings hold the potential for the future application of asparagine in the control of TMV.     

Staphylococcus aureus pathogenicity on Arabidopsis thaliana is mediated either by a direct effect of salicylic acid on the pathogen or by SA-dependent, NPR1-independent host responses

Staphylococcus aureus is a ubiquitous gram-positive bacterium that can cause superficial to serious systemic infections in animals and humans. Here we report the development of a plant infection model to study the pathogenesis of this bacterium. Three global regulatory mutants, RN6911 (agr-), ALC 488 (sarA-) ALC 842 (sarA-/agr-) and an alpha-toxin mutant defective in biofilm formation (DU1090) which are attenuated in animal pathogenesis, were also attenuated in their ability to infect plants, suggesting that these regulators that mediate synthesis of virulence factors essential for animal pathogenesis are also required for plant pathogenesis. Further, using Arabidopsis plants altered in defense responses such as the transgenic lines NahG [defective in salicylic acid (SA) accumulation], and 35S-LOX2- (defective in jasmonic acid production and hyper-accumulator of SA), and mutants ics1 (depleted in SA accumulation), and npr1-1 (non-expressor of pathogenesis-related protein) we show that resistance of Arabidopsis to typical plant pathogens and the animal pathogen S. aureus is conserved and is mediated by SA. The data presented here suggest that Arabidopsis thaliana resistance to S. aureus is mediated either by a direct effect of SA on the pathogen, specifically one that affects the attachment/aggregate formation on the root surface and reduces the pathogen's virulence, or by SA-dependent, NPR1-independent host responses.     

Imprimatins A and B: Novel plant activators targeting salicylic acid metabolism in Arabidopsis thaliana

Plant activators are agrochemicals that protect plants from a broad range of pathogens by activating the plant immune system. Unlike pesticides, they do not target pathogens; therefore, plant activators provide durable effects that are not overcome by pathogenic microbes. Although certain plant activators have been applied to paddy fields for more than 30 years, the molecular basis of the underlying immune induction are unclear. From the screening of 10,000 diverse chemicals by a high-throughput screening procedure to identify compounds that specifically enhance pathogen-induced cell death in Arabidopsis cultured cells, we identified 7 compounds, which we designated as immune priming chemicals (Imprimatins). These compounds increased disease resistance against pathogenic Pseudomonas bacteria in Arabidopsis plants. Pretreatments increased the accumulation of endogenous salicylic acid (SA) but reduced its metabolite, SA-O-β-D-glucoside (SAG). Imprimatins inhibited the enzymatic activities of 2 SA glucosyltransferases (SAGTs) in vitro at concentrations effective for immune priming. Single and double knockout Arabidopsis plants for both SAGTs consistently exhibited enhanced disease resistance and SA accumulation. Our results demonstrate that the control of the free SA pool through SA-inactivating enzymes can be a useful methodology to confer disease resistance in plants. SAGTs can pave the way for target-based discovery of novel crop protectants.     

NPR1-dependent salicylic acid signaling is not involved in elevated CO2-induced heat stress tolerance in Arabidopsis thaliana

Elevated CO₂ can protect plants from heat stress (HS); however, the underlying mechanisms are largely unknown. Here, we used a set of Arabidopsis mutants such as salicylic acid (SA) signaling mutants nonexpressor of pathogenesis-related gene 1 (npr1-1 and npr1-5) and heat-shock proteins (HSPs) mutants (hsp21 and hsp70-1) to understand the requirement of SA signaling and HSPs in elevated CO₂-induced HS tolerance. Under ambient CO₂ (380 µmol mol⁻¹) conditions, HS (42°C, 24 h) drastically decreased maximum photochemical efficiency of PSII (Fv/Fm) in all studied plant groups. Enrichment of CO₂ (800 µmol mol⁻¹) with HS remarkably increased the Fv/Fm value in all plant groups except hsp70-1, indicating that NPR1-dependent SA signaling is not involved in the elevated CO₂-induced HS tolerance. These results also suggest an essentiality of HSP70-1, but not HSP21 in elevated CO₂-induced HS mitigation.     

外源茉莉酸和茉莉酸甲酯诱导植物抗虫作用及其机理

综述了茉莉酸(jasmonic acid, JA)和茉莉酸甲酯(methyl jasmo nate, MJA)的分子结构和应用其诱导的植物抗虫作用及其机制。植物受外源茉莉酸或茉莉酸甲酯刺激后,一条反应途径是由硬脂酸途径激活防御基因,另一条途径是直接激活防御基因。防御基因激活后导致代谢途径重新配置,并可能诱导植物产生下列4种效应：（1）直接防御,即植物产生对害虫有毒的物质、抗营养和抗消化的酶类,或具驱避性和妨碍行为作用的化合物;（2）间接防御,即产生吸引天敌的挥发物;（3）不防御,即无防御反应;（4）负防御,即产生吸引害虫的挥发物。     

cmv1 is a gate for Cucumber mosaic virus transport from bundle sheath cells to phloem in melon

Cucumber mosaic virus (CMV) has the broadest host range among plant viruses, causing enormous losses in agriculture. In melon, strains of subgroup II are unable to establish a systemic infection in the near‐isogenic line SC12‐1‐99, which carries the recessive resistance gene cmv1 from the accession PI 161375, cultivar ‘Songwhan Charmi’. Strains of subgroup I overcome cmv1 resistance in a manner dependent on the movement protein. We characterized the resistance conferred by cmv1 and established that CMV‐LS (subgroup II) can move from cell to cell up to the veins in the inoculated leaf, but cannot enter the phloem. Immunogold labelling at transmission electron microscopy level showed that CMV‐LS remains restricted to the bundle sheath (BS) cells in the resistant line, and does not invade vascular parenchyma or intermediary cells, whereas, in the susceptible line ‘Piel de Sapo’ (PS), the virus invades all vein cell types. These observations indicate that the resistant allele of cmv1 restricts systemic infection in a virus strain‐ and cell type‐specific manner by acting as an important gatekeeper for virus progression from BS cells to phloem cells. Graft inoculation experiments showed that CMV‐LS cannot move from the infected PS stock into the resistant cmv1 scion, thus suggesting an additional role for cmv1 related to CMV transport within or exit from the phloem. The characterization of this new form of recessive resistance, based on a restriction of virus systemic movement, opens up the possibility to design alternative approaches for breeding strategies in melon.     

Four sequence positions of the movement protein of Cucumber mosaic virus determine the virulence against cmv1‐mediated resistance in melon

The resistance to a set of strains of Cucumber mosaic virus (CMV) in the melon accession PI 161375, cultivar ‘Songwhan Charmi’, is dependent on one recessive gene, cmv1, which confers total resistance, whereas a second set of strains is able to overcome it. We tested 11 strains of CMV subgroups I and II in the melon line SC12‐1‐99, which carries the gene cmv1, and showed that this gene confers resistance to strains of subgroup II only and that restriction is not related to either viral replication or cell‐to‐cell movement. This is the first time that a resistant trait has been correlated with CMV subgroups. Using infectious clones of the CMV strains LS (subgroup II) and FNY (subgroup I), we generated rearrangements and viral chimaeras between both strains and established that the determinant of virulence against the gene cmv1 resides in the first 209 amino acids of the movement protein, as this region from FNY is sufficient to confer virulence to the LS clone in the line SC12‐1‐99. A comparison of the sequences of the strains of both subgroups in this region shows that there are five main positions shared by all strains of subgroup II, which are different from those of subgroup I. Site‐directed mutagenesis of the CMV‐LS clone to substitute these residues for those of CMV‐FNY revealed that a combination of four of these changes [the group 64–68 (SNNLL to HGRIA), and the point mutations R81C, G171T and A195I] was required for a complete gain of function of the LS MP in the resistant melon plant.     

Different mechanisms of Trichoderma virens‐mediated resistance in tomato against Fusarium wilt involve the jasmonic and salicylic acid pathways

In the present study, we investigated the role of Trichoderma virens (TriV_JSB100) spores or cell‐free culture filtrate in the regulation of growth and activation of the defence responses of tomato (Solanum lycopersicum) plants against Fusarium oxysporum f. sp. lycopersici by the development of a biocontrol–plant–pathogen interaction system. Two‐week‐old tomato seedlings primed with TriV_JSB100 spores cultured on barley grains (BGS) or with cell‐free culture filtrate (CF) were inoculated with Fusarium pathogen under glasshouse conditions; this resulted in significantly lower disease incidence in tomato Oogata‐Fukuju plants treated with BGS than in those treated with CF. To dissect the pathways associated with this response, jasmonic acid (JA) and salicylic acid (SA) signalling in BGS‐ and CF‐induced resistance was evaluated using JA‐ and SA‐impaired tomato lines. We observed that JA‐deficient mutant def1 plants were susceptible to Fusarium pathogen when they were treated with BGS. However, wild‐type (WT) BGS‐treated tomato plants showed a higher JA level and significantly lower disease incidence. SA‐deficient mutant NahG plants treated with CF were also found to be susceptible to Fusarium pathogen and displayed low SA levels, whereas WT CF‐treated tomato plants exhibited moderately lower disease levels and substantially higher SA levels. Expression of the JA‐responsive defensin gene PDF1 was induced in WT tomato plants treated with BGS, whereas the SA‐inducible pathogenesis‐related protein 1 acidic (PR1a) gene was up‐regulated in WT tomato plants treated with CF. These results suggest that TriV_JSB100 BGS and CF differentially induce JA and SA signalling cascades for the elicitation of Fusarium oxysporum resistance in tomato.     

Signal transduction downstream of salicylic and jasmonic acid in herbivory-induced parasitoid attraction by Arabidopsis is independent of JAR1 and NPR1

Plants can defend themselves indirectly against herbivores by emitting a volatile blend upon herbivory that attracts the natural enemies of these herbivores, either predators or parasitoids. Although signal transduction in plants from herbivory to induced volatile production depends on jasmonic acid (JA) and salicylic acid (SA), the pathways downstream of JA and SA are unknown. Use of Arabidopsis provides a unique possibility to study signal transduction by use of signalling mutants, which so far has not been exploited in studies on indirect plant defence. In the present study it was demonstrated that jar1‐1 and npr1‐1 mutants are not affected in caterpillar (Pieris rapae)‐induced attraction of the parasitoid Cotesia rubecula. Both JAR1 and NPR1 (also known as NIM1) are involved in signalling downstream of JA in induced defence against pathogens such as induced systemic resistance (ISR). NPR1 is also involved in signalling downstream of SA in defence against pathogens such as systemic acquired resistance (SAR). These results demonstrate that signalling downstream of JA and SA differs between induced indirect defence against herbivores and defence against pathogens such as SAR and ISR. Furthermore, it was demonstrated that herbivore‐derived elicitors are involved in induced attraction of the parasitoid Cotesia rubecula     

Propyl gallate,a free radical scavenger,counteracts the benefits of exogenously applied salicylic acid and aggravates the deleterious effects of the southern bean mosaic virus in Rhizobium-nodulated Phaseolus vulgaris plants

Infection of Rhizobium-nodulated Phaseolus vulgaris by the southern bean mosaic virus (SBMV) markedly decreased the growth and nodulation of plants. Exogenous applications of salicylic acid (SA) at concentrations ≥10 μM further decreased growth and nodulation of virus-infected (V) plants. Only SA concentration of 5 μM in the solution improved the growth, nodulation, chlorophyll concentration and the catabolism of ureide in leaves of V plants. The spray of leaves with 2 mM propyl gallate (+Pg) decreased growth, nodulation as well as the chlorophyll and leaf ureide concentrations in V plants, regardless of the concentration of SA at which plants were grown. Ultrastructural damages in leaf cells caused by SBMV were also enhanced in V+Pg plants. The massive proliferation of virus particles and the presence of virus crystalline arrays within symbiosomes of nodules in V+Pg plants were attributed to proliferation of branched plasmodesmata in leaf vascular-parenchyma cell walls that facilitated virus movement. Virus particles were never observed in leaf and nodule tissues of V plants not sprayed with Pg. Exogenous applications of SA hindered while Pg increased the symbiotic performance of H plants, pointing out the complexity to be addressed in breeding for virus resistance in Rhizobium-nodulated beans.     

Glutathione contributes to resistance responses to TMV through a differential modulation of salicylic acid and reactive oxygen species

Systemic acquired resistance (SAR) is induced by pathogens and confers protection against a broad range of pathogens. Several SAR signals have been characterized, but the nature of the other unknown signalling by small metabolites in SAR remains unclear. Glutathione (GSH) has long been implicated in the defence reaction against biotic stress. However, the mechanism that GSH increases plant tolerance against virus infection is not entirely known. Here, a combination of a chemical, virus-induced gene-silencing-based genetics approach, and transgenic technology was undertaken to investigate the role of GSH in plant viral resistance in Nicotiana benthamiana. Tobacco mosaic virus (TMV) infection results in increasing the expression of GSH biosynthesis genes NbECS and NbGS, and GSH content. Silencing of NbECS or NbGS accelerated oxidative damage, increased accumulation of reactive oxygen species (ROS), compromised plant resistance to TMV, and suppressed the salicylic acid (SA)-mediated signalling pathway. Application of GSH or l -2-oxothiazolidine-4-carboxylic acid (a GSH activator) alleviated oxidative damage, decreased accumulation of ROS, elevated plant local and systemic resistance, enhanced the SA-mediated signalling pathway, and increased the expression of ROS scavenging-related genes. However, treatment with buthionine sulfoximine (a GSH inhibitor) accelerated oxidative damage, elevated ROS accumulation, compromised plant systemic resistance, suppressed the SA-mediated signalling pathway, and reduced the expression of ROS-regulating genes. Overexpression of NbECS reduced oxidative damage, decreased accumulation of ROS, increased resistance to TMV, activated the SA-mediated signalling pathway, and increased the expression of the ROS scavenging-related genes. We present molecular evidence suggesting GSH is essential for both local and systemic resistance of N. benthamiana to TMV through a differential modulation of SA and ROS.     

Altered growth and improved resistance of Arabidopsis against Pseudomonas syringae by overexpression of the basic amino acid transporter AtCAT1

Amino acid transporters in plants are crucial for distributing amino acids between plant organs and cellular compartments. The H⁺‐coupled plasma membrane transporter CAT1 (cationic amino acid transporter 1) facilitates the high‐affinity uptake of basic amino acids. The uptake of lysine (Lys) via the roots was not altered in loss‐of‐function mutants, in accordance with the minor expression of CAT1 in roots, but plants ectopically overexpressing CAT1 incorporated Lys at higher rates. Exogenous Lys inhibited the primary root of Arabidopsis, whereas lateral roots were stimulated. These effects were augmented by the presence or absence of CAT1. Furthermore, the total biomass of soil‐grown plants ectopically overexpressing CAT1 was reduced and the time to flowering was accelerated. These effects were accompanied by only minor changes in the overall amino acid profile. Interestingly, CAT1 belongs to a specific small cluster of nitrogen‐containing metabolite transporter genes that are rapidly up‐regulated upon infection with Pseudomonas syringae and that may participate in the systemic response of plants to pathogen attack. The overexpression of CAT1 indeed enhanced the resistance to the hemibiotrophic bacterial pathogen P. syringae via a constitutively activated salicylic acid (SA) pathway, which is consistent with the developmental defects and the resistance phenotype.