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Bacterial biofilm-like aggregates have been observed in plants, but their role in pathogenicity is underinvestigated. In the present study, we observed that extracellular DNA and polysaccharides colocalized with green fluorescent protein (GFP)-expressing Pseudomonas syringae pv. tomato (Pst) aggregates in Arabidopsis leaves, suggesting that Pst aggregates are biofilms. GFP-expressing Pst, Pst ΔalgU ΔmucAB (Pst algU mutant), and Pst ΔalgD ΔalgU ΔmucAB (Pst algU algD mutant) were examined to explore the roles of (1) alginate, a potential biofilm component; (2) Pst AlgU, thought to regulate alginate biosynthesis and some type III secretion system effector genes; and (3) intercellular salicylic acid (SA) accumulation during pathogen-associated molecular pattern-triggered immunity (PTI). Pst formed extensive aggregates in susceptible plants, whereas aggregate numbers and size were reduced in Pst algU and Pst algD algU mutants, and both multiplied poorly in planta, suggesting that aggregate formation contributes to Pst success in planta. However, in SA-deficient sid2-2 plants, Pst algD algU mutant multiplication and aggregate formation were partially restored, suggesting plant-produced SA contributes to suppression of Pst aggregate formation. Pst algD algU mutants formed fewer and smaller aggregates than Pst algU mutants, suggesting both AlgU and AlgD contribute to Pst aggregate formation. Col-0 plants accumulated low levels of SA in response to Pst and both mutants (Pst algU and Pst algD algU), suggesting the regulatory functions of AlgU are not involved in suppressing SA-mediated plant defence. Plant PTI was associated with highly reduced Pst aggregate formation and accumulation of intercellular SA in flg22-induced PTI-responding wild-type Col-0, but not in PTI-incompetent fls2, suggesting intercellular SA accumulation by Arabidopsis contributes to suppression of Pst biofilm-like aggregate formation during PTI.

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Endogenous signaling compounds are intermediaries in signaling pathways that plants use to respond to the perception of harmful and beneficial organisms. The plant elicitor peptides (Peps) of plants are important endogenous signaling molecules that induce elements of defense responses such as hormone production, increased expression of defensive genes, the activation of phosphorelays, and the induction of cell secondary messenger synthesis. The processes by which Peps confer resistance to pathogenic microorganisms have been extensively studied in Arabidopsis but are less known in crop plants. Tomato and many other solanaceous plants have an endogenous signaling polypeptide, systemin, that is involved in the defense against herbivorous insects and necrotrophic pathogens. This paper explores the similarity of the effects and chemical properties of Pep and systemin in tomato. Additionally, the relationship of the Pep receptor and systemin receptors is explored, and the identification of a second tomato Pep receptor in the literature is called into question. We suggest future directions for research on Pep signaling in solanaceous crops during interactions with microbes.

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Plants have the ability to recognize the essential chitin molecule present in the fungal cell wall, which stimulates the immune response. Phytopathogenic fungi have developed various strategies to inhibit the chitin-triggered immune response. Here, we identified a chitin deacetylase of Puccinia striiformis f. sp. tritici (Pst), known as PsCDA2, that was induced during the initial invasion of wheat and acted as an inhibitor of plant cell death. Knockdown of PsCDA2 in wheat enhanced its resistance against Pst, highlighting the significance of PsCDA2 in the host-pathogen interaction. Moreover, PsCDA2 can protect Pst urediniospores from being damaged by host chitinase in vitro. PsCDA2 also suppressed the basal chitin-induced plant immune response, including the accumulation of callose and the expression of defence genes. Overall, our results demonstrate that Pst secretes PsCDA2 as a chitin deacetylase involved in establishing infection and modifying the acetyl group to prevent the breakdown of chitin in the cell wall by host endogenous chitinases. Our research unveils a mechanism by which the fungus suppresses plant immunity, further contributing to the understanding of wheat stripe rust control. This information could have significant implications for the development of suitable strategies for protecting crops against the devastating effects of this disease.

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The plant immune system is constituted of two functionally interdependent branches that provide the plant with an effective defense against microbial pathogens. They can be considered separate since one detects extracellular pathogen-associated molecular patterns by means of receptors on the plant surface, while the other detects pathogen-secreted virulence effectors via intracellular receptors. Plant defense depending on both branches can be effectively suppressed by host-adapted microbial pathogens. In this review we focus on bacterially driven suppression of the latter, known as effector-triggered immunity (ETI) and dependent on diverse NOD-like receptors (NLRs). We examine how some effectors secreted by pathogenic bacteria carrying type III secretion systems can be subject to specific NLR-mediated detection, which can be evaded by the action of additional co-secreted effectors (suppressors), implying that virulence depends on the coordinated action of the whole repertoire of effectors of any given bacterium and their complex epistatic interactions within the plant. We consider how ETI activation can be avoided by using suppressors to directly alter compromised co-secreted effectors, modify plant defense-associated proteins, or occasionally both. We also comment on the potential assembly within the plant cell of multi-protein complexes comprising both bacterial effectors and defense protein targets.

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The constant battle of survival between pathogens and host plants has played a crucial role in shaping the course of their co-evolution. However, the major determinants of the outcome of this ongoing arms race are the effectors secreted by pathogens into host cells. These effectors perturb the defense responses of plants to promote successful infection. In recent years, extensive research in the area of effector biology has reported an increase in the repertoire of pathogenic effectors that mimic or target the conserved ubiquitin-proteasome pathway. The role of the ubiquitin-mediated degradation pathway is well known to be indispensable for various aspects of a plant's life, and thus targeting or mimicking it seems to be a smart strategy adopted by pathogens. Therefore, this review summarizes recent findings on how some pathogenic effectors mimic or act as one of the components of the ubiquitin-proteasome machinery while others directly target the plant's ubiquitin-proteasome system.

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Pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) are required for host defense against pathogens. Although PTI and ETI are intimately connected, the underlying molecular mechanisms remain elusive. In this study, we demonstrate that flg22 priming attenuates Pseudomonas syringae pv. tomato DC3000 (Pst) AvrRpt2-induced hypersensitive cell death, resistance, and biomass reduction in Arabidopsis. Mitogen-activated protein kinases (MAPKs) are key signaling regulators of PTI and ETI. The absence of MPK3 and MPK6 significantly reduces pre-PTI-mediated ETI suppression (PES). We found that MPK3/MPK6 interact with and phosphorylate the downstream transcription factor WRKY18, which regulates the expression of AP2C1 and PP2C5, two genes encoding protein phosphatases. Furthermore, we observed that the PTI-suppressed ETI-triggered cell death, MAPK activation, and growth retardation are significantly attenuated in wrky18/40/60 and ap2c1 pp2c5 mutants. Taken together, our results suggest that the MPK3/MPK6-WRKYs-PP2Cs module underlies PES and is essential for the maintenance of plant fitness during ETI.

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As a destructive plant pathogen, Phytophthora infestans secretes diverse host-entering RxLR effectors to facilitate infection. One critical RxLR effector, PiAvr3b, not only induces effector-triggered immunity (ETI), which is associated with the potato resistance protein StR3b, but also suppresses pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). To date, the molecular basis underlying such dual activities remains unknown. Based on phylogenetic analysis of global P. infestans isolates, we found two PiAvr3b isoforms that differ by three amino acids. Despite this sequence variation, the two isoforms retain the same properties in activating the StR3b-mediated hypersensitive response (HR) and inhibiting necrosis induced by three PAMPs (PiNpp, PiINF1, and PsXeg1) and an RxLR effector (Pi10232). Using a combined mutagenesis approach, we found that the dual activities of PiAvr3b were tightly linked and determined by 88 amino acids at the C-terminus. We further determined that either the W60 or the E134 residue of PiAvr3b was essential for triggering StR3b-associated HR and inhibiting PiNpp- and Pi10232-associated necrosis, while the S99 residue partially contributed to PTI suppression. Additionally, nuclear localization of PiAvr3b was required to stimulate HR and suppress PTI, but not to inhibit Pi10232-associated cell death. Our study revealed that PiAvr3b suppresses the plant immune response at different subcellular locations and provides an example in which a single amino acid of an RxLR effector links ETI induction and cell death suppression.

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Bacterial wilt, caused by Ralstonia solanacearum, one of the most destructive phytopathogens, leads to significant annual crop yield losses. Type III effectors (T3Es) mainly contribute to the virulence of R. solanacearum, usually by targeting immune-related proteins. Here, we clarified the effect of a novel E3 ubiquitin ligase (NEL) T3E, RipAW, from R. solanacearum on pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) and further explored its action mechanism. In the susceptible host Arabidopsis thaliana, we monitored the expression of PTI marker genes, flg22-induced ROS burst, and callose deposition in RipAW- and RipAWC177A-transgenic plants. Our results demonstrated that RipAW suppressed host PTI in an NEL-dependent manner. By Split-Luciferase Complementation, Bimolecular Fluorescent Complimentary, and Co-Immunoprecipitation assays, we further showed that RipAW associated with three crucial components of the immune receptor complex, namely FLS2, XLG2, and BIK1. Furthermore, RipAW elevated the ubiquitination levels of FLS2, XLG2, and BIK1, accelerating their degradation via the 26S proteasome pathway. Additionally, co-expression of FLS2, XLG2, or BIK1 with RipAW partially but significantly restored the RipAW-suppressed ROS burst, confirming the involvement of the immune receptor complex in RipAW-regulated PTI. Overall, our results indicate that RipAW impairs host PTI by disrupting the immune receptor complex. Our findings provide new insights into the virulence mechanism of R. solanacearum.

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Plant immunity is tightly controlled by a complex and dynamic regulatory network, which ensures optimal activation upon detection of potential pathogens. Accordingly, each component of this network is a potential target for manipulation by pathogens. Here, we report that RipAC, a type III-secreted effector from the bacterial pathogen Ralstonia solanacearum, targets the plant E3 ubiquitin ligase PUB4 to inhibit pattern-triggered immunity (PTI). PUB4 plays a positive role in PTI by regulating the homeostasis of the central immune kinase BIK1. Before PAMP perception, PUB4 promotes the degradation of non-activated BIK1, while after PAMP perception, PUB4 contributes to the accumulation of activated BIK1. RipAC leads to BIK1 degradation, which correlates with its PTI-inhibitory activity. RipAC causes a reduction in pathogen-associated molecular pattern (PAMP)-induced PUB4 accumulation and phosphorylation. Our results shed light on the role played by PUB4 in immune regulation, and illustrate an indirect targeting of the immune signalling hub BIK1 by a bacterial effector.

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Subtilisin, a serine protease, can trigger defense responses in a wide variety of plants, both locally and systemically, to protect against pathogens. However, key residues of subtilisin to improve resistance to plant diseases remain unknown. In this study, Nicotiana benthamiana (N. benthamiana) leaves expressing subtilisin from Bacillus velezensis LJ02 were shown to improve protection against Botrytis cinerea (B. cinerea). Furthermore, the underlying mechanism that LJ02 subtilisin improved the protective effect was explored, and the direct inhibitory effect of subtilisin on B. cinerea was excluded in vitro. Subsequently, reactive oxygen species (ROS) burst and upregulation of resistance-related genes in systemic leaves of N. benthamiana further verified that subtilisin could induce systemic protection against B. cinerea. G307A/T308A and S213A/L214A/G215A subtilisin significantly reduced the ability to resist B. cinerea infection in N. benthamiana. Furthermore, the ROS content and expression levels of resistance-related genes of both mutants were significantly decreased compared with that of wild-type subtilisin. This work identified key residues essential for the activation function of subtilisin plant immunity and was crucial in inducing plant defense responses against B. cinerea.

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The soil-borne fungus Verticillium dahliae is causing a devastating vascular disease in more than 200 species of dicotyledonous plants. The pathogen attacks susceptible plants through the roots, colonizes the plant vascular system, and causes the death of aerial tissues. In this study, we used Arabidopsis and eggplants to examine the plant protective and immunization effects of autoclaved V. dahliae spores against V. dahliae. We observed that the application of V. dahliae autoclaved spores in eggplants and Arabidopsis resulted in enhanced protection against V. dahliae, since the disease severity and pathogen colonization were lower in the plants treated with V. dahliae autoclaved spores when compared to controls. In addition, upregulation of the defense related genes PR1 and PDF1.2 in the Arabidopsis plants treated with the V. dahliae autoclaved spores was revealed. Furthermore, pathogenicity experiments in the Arabidopsis mutant cerk1, defective in chitin perception, revealed a loss of protection against V. dahliae in the cerk1 treated with the V. dahliae autoclaved spores. The participation of the chitin receptor CERK1 is evident in Arabidopsis immunization against V. dahliae using autoclaved spores of the pathogen.

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Two conserved Glycine max (soybean) mitogen activated protein kinase 3 (MAPK3) paralogs function in defense to the parasitic soybean cyst nematode Heterodera glycines. Gene Ontology analyses of RNA seq data obtained from MAPK3-1-overexpressing (OE) and MAPK3-2-OE roots compared to their control, as well as MAPK3-1-RNA interference (RNAi) and MAPK3-2-RNAi compared to their control, hierarchically orders the induced and suppressed genes, strengthening the hypothesis that their heterologous expression in Gossypium hirsutum (upland cotton) would impair parasitism by the root knot nematode (RKN) Meloidogyne incognita. MAPK3-1 expression (E) in G. hirsutum suppresses the production of M. incognita root galls, egg masses, and second stage juveniles (J2s) by 80.32%, 82.37%, and 88.21%, respectfully. Unexpectedly, egg number increases by 28.99% but J2s are inviable. MAPK3-2-E effects are identical, statistically. MAPK3-1-E and MAPK3-2-E decreases root mass 1.49-fold and 1.55-fold, respectively, as compared to the pRAP15-ccdB-E control. The reproductive factor (RF) of M. incognita for G. hirsutum roots expressing MAPK3-1-E or MAPK3-2-E decreases 60.39% and 50.46%, respectively, compared to controls. The results are consistent with upstream pathogen activated molecular pattern (PAMP) triggered immunity (PTI) and effector triggered immunity (ETI) functioning in defense to H. glycines. The experiments showcase the feasibility of employing MAPK3, through heterologous expression, to combat M. incognita parasitism, possibly overcoming impediments otherwise making G. hirsutum's defense platform deficient. MAPK homologs are identified in other important crop species for future functional analyses.

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Glycine max root cells developing into syncytia through the parasitic activities of the pathogenic nematode Heterodera glycines underwent isolation by laser microdissection (LM). Microarray analyses have identified the expression of a G. max DOESN'T MAKE INFECTIONS3 (DMI3) homolog in syncytia undergoing parasitism but during a defense response. DMI3 encodes part of the common symbiosis pathway (CSP) involving DMI1, DMI2, and other CSP genes. The identified DMI gene expression, and symbiosis role, suggests the possible existence of commonalities between symbiosis and defense. G. max has 3 DMI1, 12 DMI2, and 2 DMI3 paralogs. LM-assisted gene expression experiments of isolated syncytia under further examination here show G. max DMI1-3, DMI2-7, and DMI3-2 expression occurring during the defense response in the H. glycines-resistant genotypes G.max [Peking/PI548402] and G.max [PI88788] indicating a broad and consistent level of expression of the genes. Transgenic overexpression (OE) of G. max DMI1-3, DMI2-7, and DMI3-2 impairs H. glycines parasitism. RNA interference (RNAi) of G. max DMI1-3, DMI2-7, and DMI3-2 increases H. glycines parasitism. The combined opposite outcomes reveal a defense function for these genes. Prior functional transgenic analyses of the 32-member G. max mitogen activated protein kinase (MAPK) gene family has determined that 9 of them act in the defense response to H. glycines parasitism, referred to as defense MAPKs. RNA-seq analyses of root RNA isolated from the 9 G. max defense MAPKs undergoing OE or RNAi reveal they alter the relative transcript abundances (RTAs) of specific DMI1, DMI2, and DMI3 paralogs. In contrast, transgenically-manipulated DMI1-3, DMI2-7, and DMI3-2 expression influences MAPK3-1 and MAPK3-2 RTAs under certain circumstances. The results show G. max homologs of the CSP, and defense pathway are linked, apparently involving co-regulated gene expression.

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Efficient plant immune responses depend on the ability to recognise an invading microbe. The 22-amino acids in the N-terminal domain and the 28-amino acids in the central region of the bacterial flagellin, called flg22 and flgII-28, respectively, are important elicitors of plant immunity. Plant immunity is activated after flg22 or flgII-28 recognition by the plant transmembrane receptors FLS2 or FLS3, respectively. There is strong selective pressure on many plant pathogenic and endophytic bacteria to overcome flagellin-triggered immunity. Here we provide an overview of recent developments in our understanding of the evasion and suppression of flagellin pattern recognition by plant-associated bacteria.

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Sclerotinia sclerotiorum is a devastating pathogen that infects a broad range of host plants. The mechanism underlying plant defence against fungal invasion is still not well characterized. Here, we report that ANGUSTIFOLIA (AN), a CtBP family member, plays a role in the defence against S. sclerotiorum attack. Arabidopsis an mutants exhibited stronger resistance to S. sclerotiorum at the early stage of infection than wild-type plants. Accordingly, an mutants exhibited stronger activation of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) responses, including mitogen-activated protein kinase activation, reactive oxygen species accumulation, callose deposition, and the expression of PTI-responsive genes, upon treatment with PAMPs/microbe-associated molecular patterns. Moreover, Arabidopsis lines overexpressing AN were more susceptible to S. sclerotiorum and showed defective PTI responses. Our luminometry, bimolecular fluorescence complementation, coimmunoprecipitation, and in vitro pull-down assays indicate that AN interacts with allene oxide cyclases (AOC), essential enzymes involved in jasmonic acid (JA) biosynthesis, negatively regulating JA biosynthesis in response to S. sclerotiorum infection. This work reveals AN is a negative regulator of the AOC-mediated JA signalling pathway and PTI activation.

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The genus Colletotrichum harbors many plant pathogenic species, several of which cause significant yield losses in the field and post harvest. Typically, in order to infect their host plants, spores germinate, differentiate a pressurized infection cell, and display a hemibiotrophic lifestyle after plant invasion. Several factors required for virulence or pathogenicity have been identified in different Colletotrichum species, and adaptation of cell wall biogenesis to distinct stages of pathogenesis has been identified as a major pre-requisite for the establishment of a compatible parasitic fungus-plant interaction. Here, we highlight aspects of fungal cell wall biogenesis during plant infection, with emphasis on the maize leaf anthracnose and stalk rot fungus, Colletotrichum graminicola.

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Plants evolve a prompt and robust immune system to defend themselves against pathogen infections. Pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) is the first battle layer activated upon the PAMP's perception, which leads to multiple defense responses. The plasma membrane (PM) H+-ATPases are the primary ion pumps to create and maintain the cellular membrane potential that is critical for various essential biological processes, including plant growth, development, and defense. This study discovered that the PM H+-ATPase AHA5 is negatively involved in Arabidopsis PTI against the virulent pathogen Pseudomonas syringae pvr. tomato (Pto) DC3000 infection. The aha5 mutant plants caused the reduced stomata opening upon the Pto infection, which was associated with the salicylic acid (SA) pathway. In addition, the aha5 mutant plants caused the increased levels of callose deposition, defense-related gene expression, and SA accumulation. Our results also indicate that the PM H+-ATPase activity of AHA5 probably mediates the coupling of H2O2 generation and the apoplast alkalization in PTI responses. Moreover, AHA5 was found to interact with a vital defense regulator, RPM1-interacting protein 4 (RIN4), in vitro and in vivo, which might also be critical for its function in PTI. In summary, our studies show that AHA5 functions as a novel and critical component that is negatively involved in PTI by coordinating different defense responses during the Arabidopsis-Pto DC3000 interaction.

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High atmospheric humidity levels profoundly impact host-pathogen interactions in plants by enabling the establishment of an aqueous living space that benefits pathogens. The effectors HopM1 and AvrE1 of the bacterial pathogen Pseudomonas syringae have been shown to induce an aqueous apoplast under such conditions. However, the mechanisms by which this happens remain unknown. Here, we show that HopM1 and AvrE1 work redundantly to establish an aqueous living space by inducing a major reprogramming of the Arabidopsis thaliana transcriptome landscape. These effectors induce a strong abscisic acid (ABA) signature, which promotes stomatal closure, resulting in reduced leaf transpiration and water-soaking lesions. Furthermore, these effectors preferentially increase ABA accumulation in guard cells, which control stomatal aperture. Notably, a guard-cell-specific ABA transporter, ABCG40, is necessary for HopM1 induction of water-soaking lesions. This study provides molecular insights into a chain of events of stomatal manipulation that create an ideal microenvironment to facilitate infection.

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The first layer of active plant immunity relies upon the recognition of pathogen-associated molecular patterns (PAMPs), and the induction of PTI. Flagellin is the major protein component of the bacterial flagellum. Flagellin-derived peptide fragments such as CD2-1, flg22, and flgII-28 function as PAMPs in most higher plants. To determine the distribution of CD2-1, flg22, and flgII-28 recognition systems within plant species, the inducibility of PTI by CD2-1, flg22, and flgII-28 in 8 plant species, including monocotyledonous and dicotyledonous plants, was investigated. CD2-1 caused PTI responses in Oryza sativa, Brachypodium distachyon, and Asparagus persicus; flg22 caused PTI responses in Phyllostachys nigra, A. persicus, Arabidopsis thaliana, Nicotiana tabacum, Solanum lycopersicum, and Lotus japonicus; and flgII-28 caused PTI responses only in S. lycopersicum. Furthermore, quantitative analysis of FLS2 receptor revealed that the responsiveness of flg22 in plants was dependent on the expression level of the receptor.

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