A glycine-rich nuclear effector VdCE51 of Verticillium dahliae suppresses plant immune responses by inhibiting the accumulation of GhTRXH2

Verticillium dahliae is an important soil-borne fungal pathogen that causes great yield losses in many cash crops.Effectors of this fungus are known to regulate plant immunity but the mechanism much remains unclear.A glycine-rich nuclear effector,VdCE51,was able to suppress immune responses in tobacco against Botrytis cinerea and Sclerotinia sclerotiorum.This effector was a required factor for full virulence of V.dahliae,and its nuclear localization was a requisite for suppressing plant immunity.The thioredoxin GhTRXH2,identified as a positive regulator of plant immunity,was a host target of VdCE51.Our findings show a virulence regulating mechanism whereby the secreted nuclear effector VdCE51 interferes with the transcription of PR genes,and the SA signaling pathway by inhibiting the accumulation of GhTRXH2,thus suppressing plant immunity.

The IDD Transcription Factors:Their Functions in Plant Development and Environmental Response

INDETERMINATE-DOMAIN proteins(IDDs)are a plant-specific transcription factor family characterized by a conserved ID domain with four zinc finger motifs.Previous studies have demonstrated that IDDs coordinate a diversity of physiological processes and functions in plant growth and development,including floral transition,plant architecture,seed and root development,and hormone signaling.In this review,we especially summarized the latest knowledge on the functions and working models of IDD members in Arabidopsis,rice,and maize,particularly focusing on their role in the regulatory network of biotic and abiotic environmental responses,such as gravity,temperature,water,and pathogens.Understanding these mechanisms underlying the function of IDD proteins in these processes is important for improving crop yields by manipulating their activity.Overall,the review offers valuable insights into the functions and mechanisms of IDD proteins in plants,providing a foundation for further research and potential applications in agriculture.

The receptor-like cytoplasmic kinase RIPK regulates broad-spectrum ROS signaling in multiple layers of plant immune system

Production of reactive oxygen species(ROS)via the activity of respiratory burst oxidase homologs(RBOHs)plays a vital role in multiple layers of the plant immune system,including pathogen-associated molecular pattern-triggered immunity(PTI),damage-associated molecular pattern-triggered immunity(DTI),effector-triggered immunity(ETI),and systemic acquired resistance(SAR).It is generally established that RBOHD is activated by different receptor-like cytoplasmic kinases(RLCKs)in response to various immune elicitors.In this study,we showed that RPM1-INDUCED PROTEIN KINASE(RIPK),an RLCK VII subfamily member,contributes to ROS production in multiple layers of plant immune system.The ripk mutants showed reduced ROS production in response to treatment with all examined immune elicitors that trigger PTI,DTI,ETI,and SAR.We found that RIPK can directly phosphorylate the N-terminal region of RBOHD in vitro,and the levels of phosphorylated S343/S347 residues of RBOHD are sigfniciantly lower in ripk mutants compared with the wild type upon treatment with all tested immune elicitors.We further demonstrated that phosphorylation of RIPK is required for its function in regulating RBOHD-mediated ROS production.Similar to rbohd,ripk mutants showed reduced stomatal closure and impaired SAR,and were susceptible to the necrotrophic bacterium Pectobacterium carotovorum.Collectively,our results indicate that RIPK regulates broad-spectrum RBOHD-mediated ROS signaling during PTI,DTI,ETI,and SAR,leading to subsequent RBOHD-dependent immune responses.

A Meloidogyne incognita effector Minc03329 suppresses plant immunity and promotes parasitism

Meloidogyne incognita is a devastating plant-parasitic nematode.Effectors play important roles during the stages of nematodes infection and parasitism,but their molecular functions remain largely unknown.In this study,we characterized a new effector,Minc03329,which contains signal peptide for secretion and a C-type lectin domain.The yeast signal sequence trap experiments indicated that the signal peptide of Minc03329 is functional.In situ hybridization showed that Minc03329 was specifically expressed in the subventral esophageal gland.Real-time qPCR confirmed that the expression level of Minc03329 transcript was significantly increased in pre-parasitic and parasitic second-stage juveniles(pre-J2s and par-J2s).Tobacco rattle virus(TRV)-mediated gene silencing of Minc03329 in host plants largely reduced the pathogenicity of nematodes.On the contrary,ectopic expression of Minc03329 in Arabidopsis thaliana significantly increased plant susceptibility to nematodes.Transient expression of Minc03329 in Nicotiana benthamiana leaves suppressed the programmed cell death triggered by the pro-apoptotic protein BAX.Moreover,the transcriptome analysis of Minc03329-transgenic Arabidopsis and wild type revealed that many defense-related genes were significantly down-regulated.Interestingly,some different expressed genes were involved in the formation of nematode feeding sites.These results revealed that Minc03329 is an important effector for M.incognita,suppressing host defense response and promoting pathogenicity.

ORYZA SATIVA SPOTTED-LEAF 41(OsSPL41) Negatively Regulates Plant Immunity in Rice

Identification of immunity-associated leucine-rich repeat receptor-like protein kinases(LRR-RLK) is critical to elucidate the LRR-RLK mediated mechanism of plant immunity.Here,we reported the map-based cloning of a novel rice SPOTTED-LEAF 41(Os SPL41) encoding a putative LRR-RLK protein(Os LRR-RLK41/Os SPL41) that regulated disease responses to the bacterial blight pathogen Xanthomonas oryzae pv.oryzae(Xoo).An 8-bp insertion at position 865 bp in a mutant spotted-leaf 41(spl41) allele led to the formation of purple-brown lesions on leaves.Functional complementation by the wild type allele(Os SPL41) can rescue the mutant phenotype,and the complementary lines showed similar performance to wild type in a number of agronomic,physiological and molecular indices.Os SPL41 was constitutively expressed in all tissues tested,and Os SPL41 contains a typical transmembrane domain critical for its localization to the cell membrane.The mutant exhibited an enhanced level of resistance to Xoo in companion of markedly up-regulated expression of pathogenesis-related genes such as Os PR10a,Os PAL1 and Os NPR1,while the level of salicylic acid was significantly increased in spl41.In contrast,the over-expression lines exhibited a reduced level of H_(2)O_(2) and were much susceptible to Xoo with down-regulated expression of pathogenesis-related genes.These results suggested that Os SPL41 might negatively regulate plant immunity through the salicylic acid signaling pathway in rice.

Eureka lemon zinc finger protein ClDOF3.4 interacts with citrus yellow vein clearing virus coat protein to inhibit viral infection

Citrus yellow vein clearing virus(CYVCV)is a new citrus virus that has become an important factor restricting the development of China’s citrus industry,and the CYVCV coat protein(CP)is associated with viral pathogenicity.In this study,the Eureka lemon zinc finger protein(ZFP)ClDOF3.4 was shown to interact with CYVCV CP in vivo and in vitro.Transient expression of ClDOF3.4 in Eureka lemon induced the expression of salicylic acid(SA)-related and hypersensitive response marker genes,and triggered a reactive oxygen species burst,ion leakage necrosis,and the accumulation of free SA.Furthermore,the CYVCV titer in ClDOF3.4 transgenic Eureka lemon plants was approximately 69.4%that in control plants 6 mon after inoculation,with only mild leaf chlorotic spots observed in those transgenic plants.Taken together,the results indicate that ClDOF3.4 not only interacts with CP but also induces an immune response in Eureka lemon by inducing the SA pathways.This is the first report that ZFP is involved in the immune response of a citrus viral disease,which provides a basis for further study of the molecular mechanism of CYVCV infection.

Egg-associated secretions from the brown planthopper(Nilaparvata lugens)activate rice immune responses

The brown planthopper(BPH,Nilaparvata lugens)is a notorious sap-sucking insect pest that damages rice(Oryza sativa)plants throughout Asia.During BPH feeding,saliva enters rice plant tissues,whereas during oviposition egg-associated secretions(EAS)are deposited in damaged plant tissue.Dynamic changes in rice to planthopper salivary effectors have been widely reported.However,the effects of EAS from planthopper on rice immunity remains largely unexplored.In this study,we found that both infestation of rice by gravid BPH female adults and treatment with the EAS elicited a strong and rapid accumulation of jasmonic acid(JA),JA-isoleucine,and hydrogen peroxide in rice.EAS enhanced plant defenses not only in rice but also in tobacco,and these impaired the performance of BPH on rice,as well as the performance of aphids and whiteflies on tobacco.High-throughput proteome sequencing of EAS led to 110 proteins being identified and 53 proteins with 2 or more unique peptides being detected.Some proteins from BPH EAS were also found in the salivary proteome from herbivores,suggesting potential evolutionary conservation of effector functions across feeding and oviposition;however,others were only identified in EAS,and these are likely specifically related to oviposition.These findings point to novel proteins affecting interactions between planthoppers and rice during oviposition,providing an additional source of information for effector studies.

Global characterization of OsPIP aquaporins reveals that the H_(2)O_(2)transporter OsPIP2;6 increases resistance to rice blast

Plasma membrane intrinsic proteins(PIPs)are conserved plant aquaporins that transport small molecules across the plasma membrane to trigger instant stress responses and maintain cellular homeostasis under biotic and abiotic stress.To elucidate their roles in plant immunity to pathogen attack,we characterized the expression patterns,subcellular localizations,and H_(2)O_(2)-transport ability of 11 OsPIPs in rice(Oryza sativa),and identified OsPIP2;6 as necessary for rice disease resistance.OsPIP2;6 resides on the plasma membrane and facilitates cytoplasmic import of the immune signaling molecule H_(2)O_(2).Knockout of OsPIP2;6 increases rice susceptibility to Magnaporthe oryzae,indicating a positive function in plant immunity.OsPIP2;6 interacts with OsPIP2;2,which has been reported to increase rice resistance to pathogens via H_(2)O_(2)transport.Our findings suggest that OsPIP2;6 cooperates with OsPIP2;2 as a defense signal transporter complex during plant–pathogen interaction.

Recent advances in plant immunity with cell death:A review

Cell death is an important physiological phenomenon in life.It can be programmed or unprogrammed.Unprogrammed cell death is usually induced by abiotic or biotic stress.Recent studies have shown that many proteins regulate both cell death and immunity in plants.Here,we provide a review on the advances in plant immunity with cell death,especially the molecular regulation and underlying mechanisms of those proteins involved in both cell death and plant immunity.In addition,we discuss potential approaches toward improving plant immunity without compromising plant growth.

RALF22 promotes plant immunity and amplifies the Pep3 immune signal

Rapid alkalinization factors(RALFs)in plants have been reported to dampen pathogenassociated molecular pattern(PAMP)-triggered immunity via suppressing PAMP-induced complex formation between the pattern recognition receptor(PRR)and its co-receptor BAK1.However,the direct and positive role of RALFs in plant immunity remains largely unknown.Herein,we report the direct and positive roles of a typical RALF,RALF22,in plant immunity.RALF22alone directly elicited a variety of typical immune responses and triggered resistance against the devastating necrotrophic fungal pathogen Sclerotinia sclerotiorum in a FERONIA(FER)-dependent manner.LORELEI(LRE)-like glycosylphosphatidylinositol(GPI)-anchored protein 1(LLG1)and NADPH oxidase RBOHD were required for RALF22-elicited reactive oxygen species(ROS)generation.The mutation of cysteines conserved in the C terminus of RALFs abolished,while the constitutive formation of two disulfide bridges between these cysteines promoted the RALF22-elicited ROS production and resistance against S.sclerotiorum,demonstrating the requirement of these cysteines in the functions of RALF22 in plant immunity.Furthermore,RALF22 amplified the Pep3-induced immune signal by dramatically increasing the abundance of PROPEP3 transcript and protein.Supply with RALF22 induced resistance against S.sclerotiorum in Brassica crop plants.Collectively,our results reveal that RALF22 triggers immune responses and augments the Pep3-induced immune signal in a FER-dependent manner,and exhibits the potential to be exploited as an immune elicitor in crop protection.

New insight into Ca^(2+)-permeable channel in plant immunity

Calcium ions(Ca^(2+)) are crucial intracellular second messengers in eukaryotic cells. Upon pathogen perception, plants generate a transient and rapid increase in cytoplasmic Ca^(2+)levels, which is subsequently decoded by Ca^(2+)sensors and effectors to activate downstream immune responses. The elevation of cytosolic Ca^(2+)is commonly attributed to Ca^(2+)influx mediated by plasma membranelocalized Ca^(2+)–permeable channels. However, the contribution of Ca^(2+)release triggered by intracellular Ca^(2+)-permeable channels in shaping Ca^(2+)signaling associated with plant immunity remains poorly understood. This review discusses recent advances in understanding the mechanism underlying the shaping of Ca^(2+)signatures upon the activation of immune receptors, with particular emphasis on the identification of intracellular immune receptors as non-canonical Ca^(2+)-permeable channels. We also discuss the involvement of Ca^(2+)release from the endoplasmic reticulum in generating Ca^(2+)signaling during plant immunity.

Innate Immunity of Phlomis purpurea against Phytophthora cinnamomi: A Transcriptomic Analysis

Phlomis purpurea L.grows spontaneously in dry and stony habitats from the south of Iberian Peninsula and in cork oak(Quercus suber L.)and holm oak(Q.ilex ssp.rotundifolia,Lam.)plantations infested with Phytophthora cinnamomi(Rands).The aim of this study is to understand the genetic basis of P.purpurea innate immunity to this pathogen.The transcriptome analysis of P.purpurea upon challenging with P.cinnamomi revealed a set of up-regulated genes,related to signaling,transcription factors and response to stress.Transcripts involved in the synthesis of a number of proteins,namely:ANKYRIN,AP2,AQUAPORIN,ARMADILLO,At1G69870-LIKE,BHLH,BON1,CALMODULIN,CALNEXIN,CALRETICULINE,CC-NBS-LRR,CHAPERONE,CYTOCHROME,DUF,GH3,GMP,G-TYPE,LIPOXYGENASE,MLO-LIKE,MYB,NAC,NBS-LRR,PENTATRICOPEPTIDE,SUBTILISIN,WAK,bZIP and hormones such as BRASSINOSTEROID,JASMONATE,SALICYLATE,ETHYLENE-RESPONSIVE were identified.P.purpurea ability to cope with P.cinnamomi attack is based on the expression of a set of transcription factors and signaling molecules targeted by the pathogen.The information gathered contributes to the elucidation of the overall response of P.purpurea to P.cinnamomi attempted infection which can be helpful for improving woody species resistance to pathogenic oomycetes.

The RECEPTOR-LIKE PROTEIN53 immune complex associates with LLG1 to positively regulate plant immunity

Pattern recognition receptors(PRRs)sense ligands in pattern-triggered immunity(PTI).Plant PRRs include numerous receptor-like proteins(RLPs),but many RLPs remain functionally uncharacterized.Here,we examine an Arabidopsis thaliana RLP,RLP53,which positively regulates immune signaling.Our forward genetic screen for suppressors of enhanced disease resistance1(edr1)identified a point mutation in RLP53 that fully suppresses disease resistance and mildewinduced cell death in edr1 mutants.The rlp53 mutants showed enhanced susceptibility to virulent pathogens,including fungi,oomycetes,and bacteria,indicating that RLP53 is important for plant immunity.The ectodomain of RLP53 contains leucine-rich repeat(LRR)motifs.RLP53 constitutively associates with the LRR receptorlike kinase SUPPRESSOR OF BRASSINOSTEROIDINSENSITIVE1-ASSOCIATEDKINASE(BAK1)-INTERACTINGRECEPTORKINASE1(SOBIR1)and interacts with the co-receptor BAK1 in a pathogen-induced manner.The double mutation sobir1-12 bak1-5 suppresses edr1-mediated disease resistance,suggesting that EDR1 negatively regulates PTI modulated by the RLP53–SOBIR1–BAK1 complex.Moreover,the glycosylphosphatidylinositol(GPI)-anchored protein LORELEI-LIKE GPI-ANCHORED PROTEIN1(LLG1)interacts with RLP53 and mediates RLP53 accumulation in the plasma membrane.We thus uncovered the role of a novel RLP and its associated immune complex in plant defense responses and revealed a potential new mechanism underlying regulation of RLP immune function by a GPI-anchored protein.

Structure, biochemical function, and signaling mechanism of plant NLRs

To counter pathogen invasion,plants have evolved a large number of immune receptors,including membrane-resident pattern recognition receptors(PRRs)and intracellular nucleotide-binding and leucine-rich repeat receptors(NLRs).Our knowledge about PRR and NLR signaling mechanisms has expanded significantly over the past few years.Plant NLRs form multi-protein complexes called resistosomes in response to pathogen effectors,and the signaling mediated by NLR resistosomes converges on Ca2+-permeable channels.Ca2+-permeable channels important for PRR signaling have also been identified.These findings highlight a crucial role of Ca2+in triggering plant immune signaling.In this review,we first discuss the structural and biochemical mechanisms of non-canonical NLR Ca2+channels and then summarize our knowledge about immune-related Ca2+-permeable channels and their roles in PRR and NLR signaling.We also discuss the potential role of Ca2+in the intricate interaction between PRR and NLR signaling.

Specialized metabolites as versatile tools in shaping plant-microbe associations

Plants are rich repository of a large number of chemical compounds collectively referred to as specialized metabolites.These compounds are of importance for adaptive processes including responses against changing abiotic conditions and interactions with various co-existing organisms.One of the strikingly affirmed functions of these specialized metabolites is their involvement in plants’life-long interactions with complex multi-kingdom microbiomes including both beneficial and harmful microorganisms.Recent developments in genomic and molecular biology tools not only help to generate well-curated information about regulatory and structural components of biosynthetic pathways of plant specialized metabolites but also to create and screen mutant lines defective in their synthesis.In this review,we have comprehensively surveyed the function of these specialized metabolites and discussed recent research findings demonstrating the responses of various microbes on tested mutant lines having defective biosynthesis of particular metabolites.In addition,we attempt to provide key clues about the impact of these metabolites on the assembly of the plant microbiome by summarizing the major findings of recent comparative metagenomic analyses of available mutant lines under customized and natural microbial niches.Subsequently,we delineate benchmark initiatives that aim to engineer or manipulate the biosynthetic pathways to produce specialized metabolites in heterologous systems but also to diversify their immune function.While denoting the function of these metabolites,we also discuss the critical bottlenecks associated with understanding and exploiting their function in improving plant adaptation to the environment.

Epigenetic regulation of plant immunity:from chromatin codes to plant disease resistance

Facing a deteriorating natural environment and an increasing serious food crisis,bioengineering-based breeding is increasing in importance.To defend against pathogen infection,plants have evolved multiple defense mechanisms,including pathogen-associated molecular pattern(PAMP)-triggered immunity(PTI)and effector-triggered immunity(ETI).A complex regulatory network acts downstream of these PTI and ETI pathways,including hormone signal transduction and transcriptional reprogramming.In recent years,increasing lines of evidence show that epigenetic factors act,as key regulators involved in the transcriptional reprogramming,to modulate plant immune responses.Here,we summarize current progress on the regulatory mechanism of DNA methylation and histone modifications in plant defense responses.In addition,we also discuss the application of epigenetic mechanism-based resistance strategies in plant disease breeding.

CRISPR/Cas9-Targeted Knockout of Rice Susceptibility Genes OsDjA2 and OsERF104 Reveals Alternative Sources of Resistance to Pyricularia oryzae

Rice genes OsDjA2 and OsERF104,encoding a chaperone protein and an APETELA2/ethylene-responsive factor,respectively,are strongly induced in a compatible interaction with blast fungus,and also have function in plant susceptibility validated through gene silencing.Here,we reported the CRISPR/Cas9 knockout of OsDjA2 and OsERF104 genes resulting in considerable improvement of blast resistance.A total of 15 OsDjA2(62.5%)and 17 OsERF104(70.8%)T_(0)transformed lines were identified from 24 regenerated plants for each target and used in downstream experiments.Phenotyping of homozygous T1 mutant lines revealed not only a significant decrease in the number of blast lesions but also a reduction in the percentage of diseased leaf area,compared with the infected control plants.Our results supported CRISPR/Cas9-mediated target mutation in rice susceptibility genes as a potential and alternative breeding strategy for building resistance to blast disease.

WeiTsing: a new face of Ca^(2+)-permeable channels in plant immunity

Plants employ pattern-and effector-triggered immunity(PTI and ETI)to synergistically defend invading pathogens and insect herbivores.Both PTI and ETI can induce cytosolic Ca^(2+)spikes,despite in different spatiotemporal patterns,to activate downstream Ca^(2+)-dependent immune signaling cascades.While multiple families of Ca^(2+)-permeable channels at the plasma membrane have been uncovered,the counterparts responsible for Ca^(2+)release from intracellular stores remain poorly understood.In a groundbreaking paper published recently by Cell,the authors reported that WeiTsing,an Arabidopsis endoplasmic reticulum(ER)-resident protein that was specifically expressed in the pericycle upon Plasmodiophora brassicae(Pb)infection,could form resistosome-like Ca^(2+)-conducting channel and protect the stele of Brassica crops from Pb colonization.As the channel activity of WeiTsing was indispensable for its immune function,the findings highlight a previously underappreciated role of Ca^(2+)release from intracellular repertoire in promoting plant disease resistance.

Twisting mode of supercoil leucine-rich domain mediates peptide sensing in FLS2–flg22–BAK1 complex

Plants and animals recognize microbial invaders by detecting pathogen-associated molecular patterns (PAMPs) through pattern-recognition receptors (PRRs). This recognition plays a crucial role in plant immunity. The newly discovered protein in plants that responds to bacterial flagellin, i.e., flagellin-sensitive 2 (FLS2), is ubiquitously expressed and present in many plants. The association of FLS2 and BAK1, facilitated by a highly conserved epitope flg22 of flagellin, triggers such downstream immune responses as activated MAPK pathway and elevated reactive oxygen species (ROS) for bacterial defense and plant immunity. Here we study the intrinsic dynamics and conformational change of FLS2 upon the formation of the FLS2–flg22–BAK1 complex. The top intrinsic normal modes and principal structural fluctuation components are very similar, showing two bending modes and one twisting mode. The twisting mode alone, however, accounts for most of the conformational change of FLS2 induced by binding with flg22 and BAK1. This study indicates that flg22 binding suppresses FLS2 conformational fluctuation, especially on the twisting motion, thus facilitating FLS2–BAK1 interaction. A detailed analysis of this sensing mechanism may aid better design on both PRR and peptide mimetics for plant immunity.

Antibody-Like Phosphorylation Sites. Theme for Studies of Cancer, Aging and Evolution

Sequence similarities were found between protein and DNA sequences encoding certain part of conserved variable immunoglobulin domains (i.e. conserved IgV) and phosphorylation sites. Hypermutation motifs were then indicated in the majority of the corresponding non-IgV nucleotide sequences. According to database confirmations or double prediction of phosphorylation sites, 80% of the selected human and mouse IgV-related phosphorylation sites or their highly probable candidates exhibited substrate relationship to ataxia-telangiectasia-mutated kinase known as ATM. In accordance with literature data, inactivation of ATM by mutations can participate in the mechanisms of carcinogenesis, neurodegeneration and possibly also in aging. In agreement with this relationship, some of the selected IgV-/ATM-related segments formed molecules specifically involved in carcinogenesis. The selected IgV-related sequence segments were also similar to certain segments of higher plants containing immunoglobulin-like repeats and related regions. Bioinformatic analysis of some selected plant sequences then indicated the presence of catalytic domains composing serine/threonine/tyrosine receptor/receptor-like kinases, which are considered important structures for evolution of very early and part of later Ig-domain-related immunity. The analyzed conserved domain similarities also suggested certain interesting structural and phylogenic relationships, which need to be further investigated. This review in fact briefly summarizes the findings on the subject from the last twenty years.