Pseudomonas syringae pv. tomato DC3000 Effector HopG1 is a multi-faceted protein that Triggers Necrotic Cell Death that is attenuated by the Nonhost Resistance 2B (AtNHR2B) Protein

The plant pathogenic bacterium Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) has become a paradigm in plant-bacteria interactions due to its ability to cause disease in the model plant Arabidopsis thaliana. Pst DC3000 uses the type III secretion system to deliver type III secreted effectors (T3SEs) directly into the plant cytoplasm. Pst DC3000 T3SEs contribute to pathogenicity by suppressing plant defense responses and targeting plant’s physiological processes. Although the complete repertoire of effectors encoded in the Pst DC3000 genome have been identified, the specific function for most of them remains to be elucidated. The mitochondrial-localized T3E HopG1, suppresses plant defense responses and promotes the development of disease symptoms. Here, we show that HopG1 triggers necrotic cell death that enables the growth of non-adapted pathogens. We further showed that HopG1 interacts with the plant immunity-related protein AtNHR2B and that AtNHR2B attenuates HopG1- virulence functions.

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Jasmonic Acid at the Crossroads of Plant Immunity and Pseudomonas syringae Virulence

International Journal of Molecular Sciences ◽

10.3390/ijms21207482 ◽

2020 ◽

Vol 21 (20) ◽

pp. 7482

Author(s):

Aarti Gupta ◽

Mamta Bhardwaj ◽

Lam-Son Phan Tran

Keyword(s):

Jasmonic Acid ◽

Plant Defense ◽

Pseudomonas Syringae ◽

Regulatory Networks ◽

Genetic Manipulation ◽

Plant Immunity ◽

Stomatal Closure ◽

Defense Responses ◽

Plant Defense Responses ◽

Callose Deposition

Sensing of pathogen infection by plants elicits early signals that are transduced to affect defense mechanisms, such as effective blockage of pathogen entry by regulation of stomatal closure, cuticle, or callose deposition, change in water potential, and resource acquisition among many others. Pathogens, on the other hand, interfere with plant physiology and protein functioning to counteract plant defense responses. In plants, hormonal homeostasis and signaling are tightly regulated; thus, the phytohormones are qualified as a major group of signaling molecules controlling the most widely tinkered regulatory networks of defense and counter-defense strategies. Notably, the phytohormone jasmonic acid mediates plant defense responses to a wide array of pathogens. In this review, we present the synopsis on the jasmonic acid metabolism and signaling, and the regulatory roles of this hormone in plant defense against the hemibiotrophic bacterial pathogen Pseudomonas syringae. We also elaborate on how this pathogen releases virulence factors and effectors to gain control over plant jasmonic acid signaling to effectively cause disease. The findings discussed in this review may lead to ideas for the development of crop cultivars with enhanced disease resistance by genetic manipulation.

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Allelic Variants of the Pseudomonas syringae Type III Effector HopZ1 Are Differentially Recognized by Plant Resistance Systems

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-22-2-0176 ◽

2009 ◽

Vol 22 (2) ◽

pp. 176-189 ◽

Cited By ~ 39

Author(s):

Huanbin Zhou ◽

Robyn L. Morgan ◽

David S. Guttman ◽

Wenbo Ma

Keyword(s):

Cell Death ◽

Pseudomonas Syringae ◽

Plant Resistance ◽

Type Iii Secretion ◽

Defense Responses ◽

Plant Defense Responses ◽

Type Iii Effectors ◽

Type Iii ◽

New Evidence ◽

Bacterial Plant Pathogen

The bacterial plant pathogen Pseudomonas syringae depends on the type III secretion system and type III-secreted effectors to cause disease in plants. HopZ is a diverse family of type III effectors widely distributed in P. syringae isolates. Among the HopZ homologs, HopZ1 is ancient to P. syringae and has been shown to be under strong positive selection driven by plant resistance-imposed selective pressure. Here, we characterized the virulence and avirulence functions of the three HopZ1 alleles in soybean and Nicotiana benthamiana. In soybean, HopZ1 alleles have distinct functions: HopZ1a triggers defense response, HopZ1b promotes bacterial growth, and HopZ1c has no observable effect. In N. benthamiana, HopZ1a and HopZ1b both induce plant defense responses. However, they appear to trigger different resistance pathways, evidenced by two major differences between HopZ1a- and HopZ1b-triggered hypersensitive response (HR): i) the putative N-acylation sites had no effect on HopZ1a-triggered cell death, whereas it greatly enhanced HopZ1b-triggered cell death; and ii) the HopZ1b-triggered HR, but not the HopZ1a-triggered HR, was suppressed by another HopZ homolog, HopZ3. We previously demonstrated that HopZ1a most resembled the ancestral allelic form of HopZ1; therefore, this new evidence suggested that differentiated resistance systems have evolved in plant hosts to adapt to HopZ1 diversification in P. syringae.

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An efficient direct screening system for microorganisms that activate plant immune responses based on plant–microbe interactions using cultured plant cells

Scientific Reports ◽

10.1038/s41598-021-86560-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Mari Kurokawa ◽

Masataka Nakano ◽

Nobutaka Kitahata ◽

Kazuyuki Kuchitsu ◽

Toshiki Furuya

Keyword(s):

Immune Responses ◽

Plant Defense ◽

Pseudomonas Syringae ◽

Large Scale ◽

Plant Cells ◽

Defense Responses ◽

Root Tip ◽

General Strategy ◽

Plant Defense Responses ◽

Microbe Interactions

AbstractMicroorganisms that activate plant immune responses have attracted considerable attention as potential biocontrol agents in agriculture because they could reduce agrochemical use. However, conventional methods to screen for such microorganisms using whole plants and pathogens are generally laborious and time consuming. Here, we describe a general strategy using cultured plant cells to identify microorganisms that activate plant defense responses based on plant–microbe interactions. Microbial cells were incubated with tobacco BY-2 cells, followed by treatment with cryptogein, a proteinaceous elicitor of tobacco immune responses secreted by an oomycete. Cryptogein-induced production of reactive oxygen species (ROS) in BY-2 cells served as a marker to evaluate the potential of microorganisms to activate plant defense responses. Twenty-nine bacterial strains isolated from the interior of Brassica rapa var. perviridis plants were screened, and 8 strains that enhanced cryptogein-induced ROS production in BY-2 cells were selected. Following application of these strains to the root tip of Arabidopsis seedlings, two strains, Delftia sp. BR1R-2 and Arthrobacter sp. BR2S-6, were found to induce whole-plant resistance to bacterial pathogens (Pseudomonas syringae pv. tomato DC3000 and Pectobacterium carotovora subsp. carotovora NBRC 14082). Pathogen-induced expression of plant defense-related genes (PR-1, PR-5, and PDF1.2) was enhanced by the pretreatment with strain BR1R-2. This cell–cell interaction-based platform is readily applicable to large-scale screening for microorganisms that enhance plant defense responses under various environmental conditions.

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Method for the Production and Purification of Plant Immuno-Active Xylanase from Trichoderma

International Journal of Molecular Sciences ◽

10.3390/ijms22084214 ◽

2021 ◽

Vol 22 (8) ◽

pp. 4214

Author(s):

Gautam Anand ◽

Meirav Leibman-Markus ◽

Dorin Elkabetz ◽

Maya Bar

Keyword(s):

Immune System ◽

Plant Defense ◽

Plant Immunity ◽

Xylanase Activity ◽

Hydrolytic Enzymes ◽

Adaptive Immune System ◽

Defense Responses ◽

Plant Defense Responses ◽

Xylanase Production ◽

Ideal System

Plants lack a circulating adaptive immune system to protect themselves against pathogens. Therefore, they have evolved an innate immune system based upon complicated and efficient defense mechanisms, either constitutive or inducible. Plant defense responses are triggered by elicitors such as microbe-associated molecular patterns (MAMPs). These components are recognized by pattern recognition receptors (PRRs) which include plant cell surface receptors. Upon recognition, PRRs trigger pattern-triggered immunity (PTI). Ethylene Inducing Xylanase (EIX) is a fungal MAMP protein from the plant-growth-promoting fungi (PGPF)–Trichoderma. It elicits plant defense responses in tobacco (Nicotiana tabacum) and tomato (Solanum lycopersicum), making it an excellent tool in the studies of plant immunity. Xylanases such as EIX are hydrolytic enzymes that act on xylan in hemicellulose. There are two types of xylanases: the endo-1, 4-β-xylanases that hydrolyze within the xylan structure, and the β-d-xylosidases that hydrolyze the ends of the xylan chain. Xylanases are mainly synthesized by fungi and bacteria. Filamentous fungi produce xylanases in high amounts and secrete them in liquid cultures, making them an ideal system for xylanase purification. Here, we describe a method for cost- and yield-effective xylanase production from Trichoderma using wheat bran as a growth substrate. Xylanase produced by this method possessed xylanase activity and immunogenic activity, effectively inducing a hypersensitive response, ethylene biosynthesis, and ROS burst.

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A Verticillium dahliae Extracellular Cutinase Modulates Plant Immune Responses

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-06-17-0136-r ◽

2018 ◽

Vol 31 (2) ◽

pp. 260-273 ◽

Cited By ~ 12

Author(s):

Yue-Jing Gui ◽

Wen-Qi Zhang ◽

Dan-Dan Zhang ◽

Lei Zhou ◽

Dylan P. G. Short ◽

...

Keyword(s):

Cell Death ◽

Plant Defense ◽

Verticillium Dahliae ◽

Fungal Pathogens ◽

Sequence Divergence ◽

Defense Responses ◽

Carbohydrate Binding ◽

Dependent Manner ◽

Virus Induced Gene Silencing ◽

Plant Defense Responses

Cutinases have been implicated as important enzymes during the process of fungal infection of aerial plant organs. The function of cutinases in the disease cycle of fungal pathogens that invade plants through the roots has been less studied. Here, functional analysis of 13 cutinase (carbohydrate esterase family 5 domain–containing) genes (VdCUTs) in the highly virulent vascular wilt pathogen Verticillium dahliae Vd991 was performed. Significant sequence divergence in cutinase family members was observed in the genome of V. dahliae Vd991. Functional analyses demonstrated that only VdCUT11, as purified protein, induced cell death and triggered defense responses in Nicotiana benthamiana, cotton, and tomato plants. Virus-induced gene silencing showed that VdCUT11 induces plant defense responses in Nicotiana benthamania in a BAK1 and SOBIR-dependent manner. Furthermore, coinfiltration assays revealed that the carbohydrate-binding module family 1 protein (VdCBM1) suppressed VdCUT11-induced cell death and other defense responses in N. benthamiana. Targeted deletion of VdCUT11 in V. dahliae significantly compromised virulence on cotton plants. The cutinase VdCUT11 is an important secreted enzyme and virulence factor that elicits plant defense responses in the absence of VdCBM1.

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Two Arabidopsis Homologs of Human Lysine-Specific Demethylase Function in Epigenetic Regulation of Plant Defense Responses

Frontiers in Plant Science ◽

10.3389/fpls.2021.688003 ◽

2021 ◽

Vol 12 ◽

Author(s):

Seong Woo Noh ◽

Ri-Ra Seo ◽

Hee Jin Park ◽

Ho Won Jung

Keyword(s):

Pseudomonas Syringae ◽

Epigenetic Regulation ◽

Plant Immunity ◽

Systemic Disease ◽

Defense Responses ◽

Plant Defense Responses ◽

Promoter Regions ◽

Lysine Specific Demethylase ◽

Genome Wide ◽

Histone H3k4

Epigenetic marks such as covalent histone modification and DNA methylation are crucial for mitotically and meiotically inherited cellular memory-based plant immunity. However, the roles of individual players in the epigenetic regulation of plant immunity are not fully understood. Here we reveal the functions of two Arabidopsis thaliana homologs of human lysine-specific demethylase1-like1, LDL1 and LDL2, in the maintenance of methyl groups at lysine 4 of histone H3 and in plant immunity to Pseudomonas syringae infection. The growth of virulent P. syringae strains was reduced in ldl1 and ldl2 single mutants compared to wild-type plants. Local and systemic disease resistance responses, which coincided with the rapid, robust transcription of defense-related genes, were more stably expressed in ldl1 ldl2 double mutants than in the single mutants. At the nucleosome level, mono-methylated histone H3K4 accumulated in ldl1 ldl2 plants genome-wide and in the mainly promoter regions of the defense-related genes examined in this study. Furthermore, in silico comparative analysis of RNA-sequencing and chromatin immunoprecipitation data suggested that several WRKY transcription factors, e.g., WRKY22/40/70, might be partly responsible for the enhanced immunity of ldl1 ldl2. These findings suggest that LDL1 and LDL2 control the transcriptional sensitivity of a group of defense-related genes to establish a primed defense response in Arabidopsis.

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molecular link from PAMP perception to a MAPK cascade associated with tomato disease resistance

10.32747/2012.7597918.bard ◽

2012 ◽

Author(s):

Guido Sessa ◽

Gregory B. Martin

Keyword(s):

Cell Death ◽

Disease Resistance ◽

Immune Responses ◽

Map Kinase ◽

Pseudomonas Syringae ◽

Molecular Mechanisms ◽

Plant Immunity ◽

Defense Responses ◽

Research Activities ◽

Mapk Cascades

The research problem: The detection of pathogen-associated molecular patterns (PAMPs) by plant pattern recognition receptors (PRRs) is a key mechanism by which plants activate an effective immune response against pathogen attack. MAPK cascades are important signaling components downstream of PRRs that transduce the PAMP signal to activate various defense responses. Preliminary experiments suggested that the receptor-like cytoplasmickinase (RLCK) Mai5 plays a positive role in pattern-triggered immunity (PTI) and interacts with the MAPKKK M3Kε. We thus hypothesized that Mai5, as other RLCKs, functions as a component PRR complexes and acts as a molecular link between PAMP perception and activation of MAPK cascades. Original goals: The central goal of this research was to investigate the molecular mechanisms by which Mai5 and M3Kε regulate plant immunity. Specific objectives were to: 1. Determine the spectrum of PAMPs whose perception is transmitted by M3Kε; 2. Identify plant proteins that act downstream of M3Kε to mediate PTI; 3. Investigate how and where Mai5 interacts with M3Kε in the plant cell; 4. Examine the mechanism by which Mai5 contributes to PTI. Changes in research directions: We did not find convincing evidence for the involvement of M3Kε in PTI signaling and substituted objectives 1 and 3 with research activities aimed at the analysis of transcriptomic profiles of tomato plants during the onset of plant immunity, isolation of the novel tomato PRR FLS3, and investigation of the involvement of the RLCKBSKs in PTI. Main achievements during this research program are in the following major areas: 1. Functional characterization of Mai5. The function of Mai5 in PTI signaling was demonstrated by testing the effect of silencing the Mai5 gene by virus-induced gene silencing (VIGS) experiments and in cell death assays. Domains of Mai5 that interact with MAPKKKs and subcellular localization of Mai5 were analyzed in detail. 2. Analysis of transcriptional profiles during the tomato immune responses to Pseudomonas syringae (Pombo et al., 2014). We identified tomato genes whose expression is induced specifically in PTI or in effector-triggered immunity (ETI). Thirty ETI-specific genes were examined by VIGS for their involvement in immunity and the MAPKKK EPK1, was found to be required for ETI. 3. Dissection of MAP kinase cascades downstream of M3Kε (Oh et al., 2013; Teper et al., 2015). We identified genes that encode positive (SGT and EDS1) and negative (WRKY1 and WRKY2) regulators of the ETI-associated cell death mediated by M3Kε. In addition, the MKK2 MAPKK, which acts downstream of M3Kε, was found to interact with the MPK3 MAPK and specific MPK3 amino acids involved interaction were identified and found to be required for induction of cell death. We also identified 5 type III effectors of the bacterial pathogen Xanthomonaseuvesicatoria that inhibited cell death induced by components of ETI-associated MAP kinase cascades. 4. Isolation of the tomato PRR FLS3 (Hind et al., submitted). FLS3, a novel PRR of the LRR-RLK family that specifically recognizes the flagellinepitope flgII-28 was isolated. FLS3 was shown to bind flgII-28, to require kinase activity for function, to act in concert with BAK1, and to enhance disease resistance to Pseudomonas syringae. 5. Functional analysis of RLCKs of the brassinosteroid signaling kinase (BSK) family.Arabidopsis and tomato BSKs were found to interact with PRRs. In addition, certain ArabidospsisBSK mutants were found to be impaired in PAMP-induced resistance to Pseudomonas syringae. Scientific and agricultural significance: Our research activities discovered and characterized new molecular components of signaling pathways mediating recognition of invading pathogens and activation of immune responses against them. Increased understanding of molecular mechanisms of immunity will allow them to be manipulated by both molecular breeding and genetic engineering to produce plants with enhanced natural defense against disease.

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The Majority of the Type III Effector Inventory of Pseudomonas syringae pv. tomato DC3000 Can Suppress Plant Immunity

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-22-9-1069 ◽

2009 ◽

Vol 22 (9) ◽

pp. 1069-1080 ◽

Cited By ~ 147

Author(s):

Ming Guo ◽

Fang Tian ◽

Yashitola Wamboldt ◽

James R. Alfano

Keyword(s):

Pseudomonas Syringae ◽

Plant Immunity ◽

In Planta ◽

Tomato Dc3000 ◽

Type Iii Effector ◽

Type Iii Effectors ◽

Type Iii ◽

Molecular Pattern ◽

Effector Triggered Immunity ◽

Type Iii Protein Secretion

The Pseudomonas syringae type III protein secretion system (T3SS) and the type III effectors it injects into plant cells are required for plant pathogenicity and the ability to elicit a hypersensitive response (HR). The HR is a programmed cell death that is associated with effector-triggered immunity (ETI). A primary function of P. syringae type III effectors appears to be the suppression of ETI and pathogen-associated molecular pattern–triggered immunity (PTI), which is induced by conserved molecules on microorganisms. We reported that seven type III effectors from P. syringae pv. tomato DC3000 were capable of suppressing an HR induced by P. fluorescens(pHIR11) and have now tested 35 DC3000 type III effectors in this assay, finding that the majority of them can suppress the HR induced by HopA1. One newly identified type III effector with particularly strong HR suppression activity was HopS2. We used the pHIR11 derivative pLN1965, which lacks hopA1, in related assays and found that a subset of the type III effectors that suppressed HopA1-induced ETI also suppressed an ETI response induced by AvrRpm1 in Arabidopsis thaliana. A. thaliana plants expressing either HopAO1 or HopF2, two type III effectors that suppressed the HopA1-induced HR, were reduced in the flagellin-induced PTI response as well as PTI induced by other PAMPs and allowed enhanced in planta growth of P. syringae. Collectively, our results suggest that the majority of DC3000 type III effectors can suppress plant immunity. Additionally, the construct pLN1965 will likely be a useful tool in determining whether other type III effectors or effectors from other types of pathogens can suppress either ETI, PTI, or both.

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A Survey of the Pseudomonas syringae pv. tomato DC3000 Type III Secretion System Effector Repertoire Reveals Several Effectors That Are Deleterious When Expressed in Saccharomyces cerevisiae

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-21-4-0490 ◽

2008 ◽

Vol 21 (4) ◽

pp. 490-502 ◽

Cited By ~ 34

Author(s):

Kathy R. Munkvold ◽

Michael E. Martin ◽

Philip A. Bronstein ◽

Alan Collmer

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Death ◽

Pseudomonas Syringae ◽

Type Iii Secretion ◽

Type Iii Secretion System ◽

Secretion System ◽

Dependent Manner ◽

Tomato Dc3000 ◽

Type Iii ◽

Pathogen Effector

The injection of nearly 30 effector proteins by the type III secretion system underlies the ability of Pseudomonas syringae pv. tomato DC3000 to cause disease in tomato and other host plants. The search for effector functions is complicated by redundancy within the repertoire and by plant resistance (R)-gene sentinels, which may convert effector virulence activities into a monolithic defense response. On the premise that some effectors target universal eukaryotic processes and that yeast (Saccharomyces cerevisiae) lacks R genes, the DC3000 effector repertoire was expressed in yeast. Of 27 effectors tested, HopAD1, HopAO1, HopD1, HopN1, and HopU1 were found to inhibit growth when expressed from a galactose-inducible GAL1 promoter, and HopAA1-1 and HopAM1 were found to cause cell death. Catalytic site mutations affecting the tyrosine phosphatase activity of HopAO1 and the cysteine protease activity of HopN1 prevented these effectors from inhibiting yeast growth. Expression of HopAA1-1, HopAM1, HopAD1, and HopAO1 impaired respiration in yeast, as indicated by tests with ethanol glycerol selective media. HopAA1-1 colocalized with porin to yeast mitochondria and was shown to cause cell death in yeast and plants in a domain-dependent manner. These results support the use of yeast for the study of plant-pathogen effector repertoires.

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