Impacto de la matriz extracelular de Bacillus subtilis en la respuesta fisiológica de cucurbitáceas

Las plantas viven en asociación con microorganismos, y han desarrollado estrategias para discriminar entre beneficiosos y patogénicos. Bacillus subtilis, agente de biocontrol en cucurbitáceas frente a patógenos como el oídio, es capaz de colonizar distintas superficies de las plantas, como las raíces o las hojas, mediante el establecimiento de comunidades bacterianas denominadas biopelículas o biofilms. Para establecerse en la parte aérea de las plantas, B. subtilis debe enfrentarse a condiciones ambientales adversas, como son los cambios de temperatura, el viento, la radiación solar y la reducida disponibilidad de nutrientes impuesta por la cutícula vegetal, lo que limita su colonización y crecimiento en dicha superficie. Por otro lado, Bacillus es capaz de influenciar el estado fisiológico de las plantas, promover su crecimiento o inducir una resistencia sistémica frente a patógenos. Para formar biofilms, Bacillus secreta una serie de compuestos, entre los que destaca la surfactina, que promueve el ensamblaje de una matriz extracelular en la que se embeben las células, cuyos componentes mayoritarios son proteínas (por ejemplo, las proteínas TasA y TapA, que forman fibras amiloides) y exopolisacáridos. Nuestro principal objetivo es determinar cómo el establecimiento de las comunidades de Bacillus afecta al estado fisiológico de las plantas, principalmente melón y pepino, y qué papel juegan los componentes de la matriz extracelular en la interacción con las plantas, así como los mecanismos por los cuales las plantas modulan la formación de dichas comunidades en la superficie de las hojas. El análisis de la respuesta de Arabidopsis y melón a distintos componentes de la matriz extracelular de B. subtilis sugiere que tanto la surfactina como los elementos mayoritarios de la matriz (las proteínas TasA y TapA, exopolisacáridos) activan mecanismos de señalización en ambas especies, que pueden promover el crecimiento de la planta o anticipar la defensa frente a patógenos.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Estudio de la interacción de Bacillus cereus responsable de intoxicaciones alimentarias en humanos con la superficie de hojas y frutos

Bacillus cereus es un patógeno humano bastante común que se transmite por alimentos y es responsable de importantes brotes de intoxicación alimentaria. El desarrollo de la enfermedad emética está relacionado con la producción de la toxina termo-resistente cereulide, que normalmente se produce en los alimentos contaminados por B. cereus. La enfermedad diarreica es provocada por varias enterotoxinas: la hemolisina BL (Hbl), la enterotoxina no hemolítica (Nhe) y la citotoxina K (CytK) . En este trabajo, se ha estudiado el comportamiento de diferentes aislados, procedentes de intoxicaciones alimentarias, sobre la superficie de frutos y hojas de plantas como posibles vehículos de estos microorganismos a través de los cuales se producirían las toxiinfecciones alimentarias. De entre todos los aislados seleccionados se eligió un grupo de 8 aislados, seis de los cuales eran capaces de persistir en hoja y fruto y los otros dos no, que se secuenciaron y se buscó los grupos de genes que eran compartidos por los diferentes aislados capaces de persistir sobre las superficies y que sin embargo no están presentes en los aislados que no son capaces de persistir. Para el posterior estudio de la relevancia de estos grupos de genes en la persistencia en hoja y fruto, se seleccionó una cepa emética del estudio por varios motivos: i) produce la toxina emética cereulide, y una enterotoxina no hemolítica, siendo por tanto de gran interés en seguridad alimentaria, ii) en las distintas plantas ensayadas se mantuvo a concentraciones de 105 UFC por gramo de hoja o fruto inoculado de los que al menos un 40% apareció en la forma de esporas y iii) es una cepa manipulable genéticamente. Con todos estos datos, nos encontramos posicionados para comprender que herramientas, conocidas o aún por conocer utiliza esta cepa para interaccionar con la planta y frutos, y de qué forma coordina su persistencia con una eventual producción de toxinas que tienen como diana al hombre

ERECTA and BAK1 Receptor Like Kinases Interact to Regulate Immune Responses in Arabidopsis

ERECTA (ER) receptor-like kinase (RLK) regulates Arabidopsis thaliana organ growth, and inflorescence and stomatal development by interacting with the ERECTA-family genes (ERf) paralogs, ER-like 1 (ERL1) and ERL2, and the receptor-like protein (RLP) TOO MANY MOUTHS (TMM). ER also controls immune responses and resistance to pathogens such as the bacterium Pseudomonas syringae pv. tomato DC3000 (Pto) and the necrotrophic fungus Plectosphaerella cucumerina BMM (PcBMM). We found that er null-mutant plants overexpressing an ER dominant-negative version lacking the cytoplasmic kinase domain (ERΔK) showed an enhanced susceptibility to PcBMM, suggesting that ERΔK associates and forms inactive complexes with additional RLKs/RLPs required for PcBMM resistance. Genetic analyses demonstrated that ER acts in a combinatorial specific manner with ERL1, ERL2, and TMM to control PcBMM resistance. Moreover, BAK1 (BRASSINOSTEROID INSENSITIVE 1-associated kinase 1) RLK, which together with ERf/TMM regulates stomatal patterning and resistance to Pto, was also found to have an unequal contribution with ER in regulating immune responses and resistance to PcBMM. Co-immunoprecipitation experiments in Nicotiana benthamiana further demonstrated BAK1-ER protein interaction. The secreted epidermal pattern factor peptides (EPF1 and EPF2), which are perceived by ERf members to specify stomatal patterning, do not seem to regulate ER-mediated immunity to PcBMM, since their inducible overexpression in A. thaliana did not impact on PcBMM resistance. Our results indicate that the multiproteic receptorsome formed by ERf, TMM and BAK1 modulates A. thaliana resistance to PcBMM, and suggest that the cues underlying ERf/TMM/BAK1-mediated immune responses are distinct from those regulating stomatal pattering

Non‐branched β‐1,3‐glucan oligosaccharides trigger immune responses in Arabidopsis

[EN] Fungal cell walls, which are essential for environmental adaptation and host colonization by the fungus, have been evolutionarily selected by plants and animals as a source of microbe-associated molecular patterns (MAMPs) that, upon recognition by host pattern recognition receptors (PRRs), trigger immune responses conferring disease resistance. Chito-oligosaccharides [b-1,4-N-acetylglucosamine oligomers, (GlcNAc)n] are the only glycosidic structures from fungal walls that have been well-demonstrated to function as MAMPs in plants. Perception of (GlcNAc)4–8 by Arabidopsis involves CERK1, LYK4 and LYK5, three of the eight members of the LysM PRR family. We found that a glucan-enriched wall fraction from the pathogenic fungus Plectosphaerella cucumerina which was devoid of GlcNAc activated immune responses in Arabidopsis wild-type plants but not in the cerk1 mutant. Using this differential response, we identified the non-branched 1,3-b-D-(Glc) hexasaccharide as a major fungal MAMP. Recognition of 1,3-b-D-(Glc)6 was impaired in cerk1 but not in mutants defective in either each of the LysM PRR family members or in the PRR-co-receptor BAK1. Transcriptomic analyses of Arabidopsis plants treated with 1,3-b-D-(Glc)6 further demonstrated that this fungal MAMP triggers the expression of immunity-associated genes. In silico docking analyses with molecular mechanics and solvation energy calculations corroborated that CERK1 can bind 1,3-b-D-(Glc)6 at effective concentrations similar to those of (GlcNAc)4. These data support that plants, like animals, have selected as MAMPs the linear 1,3-b-D-glucans present in the walls of fungi and oomycetes. Our data also suggest that CERK1 functions as an immune co-receptor for linear 1,3-b-D-glucans in a similar way to its proposed function in the recognition of fungal chito-oligosaccharides and bacterial peptidoglycan MAMPs.S

Alteration of cell wall xylan acetylation triggers defense responses that counterbalance the immune deficiencies of plants impaired in the β‐subunit of the heterotrimeric G‐protein

[EN] Arabidopsis heterotrimeric G-protein complex modulates pathogen-associated molecular pattern-triggeredimmunity (PTI) and disease resistance responses to different types of pathogens. It also plays a role in plantcell wall integrity as mutants impaired in the Gb-(agb1-2)orGc-subunits have an altered wall compositioncompared with wild-type plants. Here we performed a mutant screen to identify suppressors ofagb1-2(sgb) that restore susceptibility to pathogens to wild-type levels. Out of the foursgbmutants (sgb10–sgb13)identified,sgb11is a new mutant allele ofESKIMO1(ESK1), which encodes a plant-specific polysaccharideO-acetyltransferase involved in xylan acetylation. Null alleles (sgb11/esk1-7)ofESK1restore to wild-typelevels the enhanced susceptibility ofagb1-2to the necrotrophic fungusPlectosphaerella cucumerina BMM(PcBMM), but not to the bacteriumPseudomonas syringaepv.tomatoDC3000 or to the oomyceteHyaloper-onospora arabidopsidis. The enhanced resistance toPcBMMof theagb1-2 esk1-7double mutant was notthe result of the re-activation of deficient PTI responses inagb1-2. Alteration of cell wall xylan acetylationcaused byESK1impairment was accompanied by an enhanced accumulation of abscisic acid, the constitu-tive expression of genes encoding antibiotic peptides and enzymes involved in the biosynthesis of trypto-phan-derived metabolites, and the accumulation of disease resistance-related secondary metabolites anddifferent osmolites. Theseesk1-mediated responses counterbalance the defective PTI andPcBMMsuscepti-bility ofagb1-2plants, and explain the enhanced drought resistance ofesk1plants. These results suggestthat a deficient PTI-mediated resistance is partially compensated by the activation of specific cell-wall-trig-gered immune responses.SISpanish Ministry of Economy and Competitiveness (MINECO) grants BIO2012-32910 to A.M.The Australian Research Council Centre of Excellence in Plant Cell Walls and matching funding from KTH (grants to V.B) and NIGMS (R01GM065989) and NSF (MCB-1713880) to A.M.J. The Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences of the US Department of Energy through the grant DE-FG02-05er15671 to A.M.J. funded technical support in this study. H.M. was supported by an IEF grant (SignWALLINg-624721) from the European Union, E.M. by a Juan de la Cierva Postdoctoral Fellow from MINECO, S.S. by the BRAVE Erasmus Mundi Program (European Union), and A.M.-B. was the recipient of a PIF fellow from Universidad Politécnica de Madrid

YODA Kinase Controls a Novel Immune Pathway of Tomato Conferring Enhanced Disease Resistance to the Bacterium Pseudomonas syringae

© 2020 Téllez, Muñoz-Barrios, Sopeña-Torres, Martín-Forero, Ortega, Pérez, Sanz, Borja, de Marcos, Nicolas, Jahrmann, Mena, Jordá and Molina.Mitogen-activated protein kinases (MAPK) play pivotal roles in transducing developmental cues and environmental signals into cellular responses through pathways initiated by MAPK kinase kinases (MAP3K). AtYODA is a MAP3K of Arabidopsis thaliana that controls stomatal development and non-canonical immune responses. Arabidopsis plants overexpressing a constitutively active YODA protein (AtCA-YDA) show broad-spectrum disease resistance and constitutive expression of defensive genes. We tested YDA function in crops immunity by heterologously overexpressing AtCA-YDA in Solanum lycopersicum. We found that these tomato AtCA-YDA plants do not show developmental phenotypes and fitness alterations, except a reduction in stomatal index, as reported in Arabidopsis AtCA-YDA plants. Notably, AtCA-YDA tomato plants show enhanced resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 and constitutive upregulation of defense-associated genes, corroborating the functionality of YDA in tomato immunity. This function was further supported by generating CRISPR/Cas9-edited tomato mutants impaired in the closest orthologs of AtYDA [Solyc08g081210 (SlYDA1) and Solyc03g025360 (SlYDA2)]. Slyda1 and Slyda2 mutants are highly susceptible to P. syringae pv. tomato DC3000 in comparison to wild-type plants but only Slyda2 shows altered stomatal index. These results indicate that tomato orthologs have specialized functions and support that YDA also regulates immune responses in tomato and may be a trait for breeding disease resistance.This work was supported by grants BIO2015-64077-R of the Spanish Ministry of Economy and Competitiveness (MINECO), RTI2018-096975-B-I00 of Spanish Ministry of Science, Innovation and Universities, and grant P190050072 of Plant Response Biotech SL to AMo. Research in the Montaña Mena’s laboratory is supported by the MINECO (PPII10-0194-4164) and the Junta de Comunidades de Castilla-La Mancha (SBPLY/17/180501/000394), complemented with EU FEDER funds. JT was financially supported by a PhD fellow of the FPI program from MINECO (BES-2016-076708). AM-F and AO were recipients of research and PhD fellowships from JCCM

YODA MAPK kinase kinase regulates a novel immunity pathway conferring broad-spectrum resistance to pathogens

Plant mitogen-activated protein kinase (MAPK) casca des transduce environmental molecular signals and developmental cues into cellular responses. Among these signals are the pathogen-associated molecular patterns (PAMPs) that upon recognition by plant pattern recognition receptors (PRR), including Receptor-Like Kinases (RLKs), activate MAPK cascades that regulate PAMP-triggered immunity responses (PTI)

Differential Expression of Fungal Genes Determines the Lifestyle of Plectosphaerella Strains During Arabidopsis thaliana Colonization

16 Päg.The fungal genus Plectosphaerella comprises species and strains with different lifestyles on plants, such as P. cucumerina, which has served as model for the characterization of Arabidopsis thaliana basal and nonhost resistance to necrotrophic fungi. We have sequenced, annotated, and compared the genomes and transcriptomes of three Plectosphaerella strains with different lifestyles on A. thaliana, namely, PcBMM, a natural pathogen of wild-type plants (Col-0), Pc2127, a nonpathogenic strain on Col-0 but pathogenic on the immunocompromised cyp79B2 cyp79B3 mutant, and P0831, which was isolated from a natural population of A. thaliana and is shown here to be nonpathogenic and to grow epiphytically on Col-0 and cyp79B2 cyp79B3 plants. The genomes of these Plectosphaerella strains are very similar and do not differ in the number of genes with pathogenesis-related functions, with the exception of secreted carbohydrate-active enzymes (CAZymes), which are up to five times more abundant in the pathogenic strain PcBMM. Analysis of the fungal transcriptomes in inoculated Col-0 and cyp79B2 cyp79B3 plants at initial colonization stages confirm the key role of secreted CAZymes in the necrotrophic interaction, since PcBMM expresses more genes encoding secreted CAZymes than Pc2127 and P0831. We also show that P0831 epiphytic growth on A. thaliana involves the transcription of specific repertoires of fungal genes, which might be necessary for epiphytic growth adaptation. Overall, these results suggest that in-planta expression of specific sets of fungal genes at early stages of colonization determine the diverse lifestyles and pathogenicity of Plectosphaerella strains.This work was supported by the Spanish Ministry of Economy and Competitiveness (MINECO) grant BIO2015-64077-R and the Spanish Research Agency (AEI) grant RTI2018-096975-B-I00 to A. Molina and by the “Severo Ochoa Programme for Centers of Excellence in R&D” grant SEV-2016-0672 (2017-2021) to the CBGP (UPM-INIA). In the frame of SEV-2016-0672 program, H. Mélida was supported with a postdoctoral contract. A. Muñoz-Barrios was financially supported by the Universidad Politécnica de Madrid (UPM) Ph.D. students PIF program, I. del Hierro was a FPU fellow (Spanish Ministry of Education, Culture and Sports grant FPU16/07118), V. Fernández-Calleja was supported by the Consejería de Educacíon e Investigacíon of Comunidad de Madrid YEI program for postdoctoral researchers (PEJD-2016/BIO-3327), and the work was further supported through a Comunidad de Madrid YEI program for laboratory technicians grant (PEJ16/BIO/TL-1570).Peer reviewe

Functional characterization of YODA, a mitogen-activated protein kinase kinase kinase (MAP3K) that regulates a novel innate immunity pathway in Arabidopsis thaliana

Las cascadas de señalización mediadas por proteína quinasas activadas por mitógeno (MAP quinasas) son capaces de integrar y transducir señales ambientales en respuestas celulares. Entre estas señales se encuentran los PAMPs/MAMPs (Pathogen/Microbe-Associated Molecular Patterns), que son moléculas de patógenos o microorganismos, o los DAMPs (Damaged-Associated Molecular Patterns), que son moléculas derivadas de las plantas producidas en respuesta a daño celular. Tras el reconocimiento de los PAMPs/DAMPs por receptores de membrana denominados PRRs (Pattern Recognition Receptors), como los receptores con dominio quinasa (RLKs) o los receptores sin dominio quinasa (RLPs), se activan respuestas moleculares, incluidas cascadas de MAP quinasas, que regulan la puesta en marcha de la inmunidad activada por PAMPs (PTI). Esta Tesis describe la caracterización funcional de la MAP quinasa quinasa quinasa (MAP3K) YODA (YDA), que actúa como un regulador clave de la PTI en Arabidopsis. Se ha descrito previamente que YDA controla varios procesos de desarrollo, como la regulación del patrón estomático, la elongación del zigoto y la arquitectura floral. Hemos caracterizado un alelo mutante hipomórfico de YDA (elk2 o yda11) que presenta una elevada susceptibilidad a patógenos biótrofos y necrótrofos. Notablemente, plantas que expresan una forma constitutivamente activa de YDA (CA-YDA), con una deleción en el dominio N-terminal, presentan una resistencia de amplio espectro frente a diferentes tipos de patógenos, incluyendo hongos, oomicetos y bacterias, lo que indica que YDA juega un papel importante en la regulación de la resistencia de las plantas a patógenos. Nuestros datos indican que esta función es independiente de las respuestas inmunes mediadas por los receptores previamente caracterizados FLS2 y CERK1, que reconocen los PAMPs flg22 y quitina, respectivamente, y que están implicados en la resistencia de Arabidopsis frente a bacterias y hongos. Hemos demostrado que YDA controla la resistencia frente al hongo necrótrofo Plectosphaerella cucumerina y el patrón estomático mediante su interacción genética con la RLK ERECTA (ER), un PRR implicado en la regulación de estos procesos. Por el contrario, la interacción genética entre ER y YDA en la regulación de otros procesos de desarrollo es aditiva en lugar de epistática. Análisis genéticos indicaron que MPK3, una MAP quinasa que funciona aguas abajo de YDA en el desarrollo estomático, es un componente de la ruta de señalización mediada por YDA para la resistencia frente a P. cucumerina, lo que sugiere que el desarrollo de las plantas y la PTI comparten el módulo de transducción de MAP quinasas asociado a YDA. Nuestros experimentos han revelado que la resistencia mediada por YDA es independiente de las rutas de señalización reguladas por las hormonas de defensa ácido salicílico, ácido jasmónico, ácido abscísico o etileno, y también es independiente de la ruta de metabolitos secundarios derivados del triptófano, que están implicados en inmunidad vegetal. Además, hemos demostrado que respuestas asociadas a PTI, como el aumento en la concentración de calcio citoplásmico, la producción de especies reactivas de oxígeno, la fosforilación de MAP quinasas y la expresión de genes de defensa, no están afectadas en el mutante yda11. La expresión constitutiva de la proteína CA-YDA en plantas de Arabidopsis no provoca un aumento de las respuestas PTI, lo que sugiere la existencia de mecanismos de resistencia adicionales regulados por YDA que son diferentes de los regulados por FLS2 y CERK1. En línea con estos resultados, nuestros datos transcriptómicos revelan una sobre-representación en plantas CA-YDA de genes de defensa que codifican, por ejemplo, péptidos antimicrobianos o reguladores de muerte celular, o proteínas implicadas en la biogénesis de la pared celular, lo que sugiere una conexión potencial entre la composición e integridad de la pared celular y la resistencia de amplio espectro mediada por YDA. Además, análisis de fosfoproteómica indican la fosforilación diferencial de proteínas relacionadas con la pared celular en plantas CA-YDA en comparación con plantas silvestres. El posible papel de la ruta ER-YDA en la regulación de la integridad de la pared celular está apoyado por análisis bioquímicos y glicómicos de las paredes celulares de plantas er, yda11 y CA-YDA, que revelaron cambios significativos en la composición de la pared celular de estos genotipos en comparación con la de plantas silvestres. En resumen, nuestros datos indican que ER y YDA forman parte de una nueva ruta de inmunidad que regula la integridad de la pared celular y respuestas defensivas, confiriendo una resistencia de amplio espectro frente a patógenos. ABSTRACT Plant mitogen-activated protein kinase (MAPK) cascades transduce environmental signals and developmental cues into cellular responses. Among these signals are the pathogen- or microbe-associated molecular patterns (PAMPs or MAMPs) and the damage-associated molecular patterns (DAMPs). These PAMPs/DAMPs, upon recognition by plant pattern recognition receptors (PRRs), such as Receptor-Like Kinases (RLKs) and Receptor-Like Proteins (RLPs), activate molecular responses, including MAPK cascades, which regulate the onset of PAMP-triggered immunity (PTI). This Thesis describes the functional characterization of the MAPK kinase kinase (MAP3K) YODA (YDA) as a key regulator of Arabidopsis PTI. YDA has been previously described to control several developmental processes, such as stomatal patterning, zygote elongation and inflorescence architecture. We characterized a hypomorphic, non-embryo lethal mutant allele of YDA (elk2 or yda11) that was found to be highly susceptible to biotrophic and necrotrophic pathogens. Remarkably, plants expressing a constitutive active form of YDA (CA-YDA), with a deletion in the N-terminal domain, showed broad-spectrum resistance to different types of pathogens, including fungi, oomycetes and bacteria, indicating that YDA plays a relevant function in plant resistance to pathogens. Our data indicated that this function is independent of the immune responses regulated by the well characterized FLS2 and CERK1 RLKs, which are the PRRs recognizing flg22 and chitin PAMPs, respectively, and are required for Arabidopsis resistance to bacteria and fungi. We demonstrate that YDA controls resistance to the necrotrophic fungus Plectosphaerella cucumerina and stomatal patterning by genetically interacting with ERECTA (ER) RLK, a PRR involved in regulating these processes. In contrast, the genetic interaction between ER and YDA in the regulation of other ER-associated developmental processes was additive, rather than epistatic. Genetic analyses indicated that MPK3, a MAP kinase that functions downstream of YDA in stomatal development, also regulates plant resistance to P. cucumerina in a YDA-dependent manner, suggesting that the YDA-associated MAPK transduction module is shared in plant development and PTI. Our experiments revealed that YDA-mediated resistance was independent of signalling pathways regulated by defensive hormones like salicylic acid, jasmonic acid, abscisic acid or ethylene, and of the tryptophan-derived metabolites pathway, which are involved in plant immunity. In addition, we showed that PAMP-mediated PTI responses, such as the increase of cytoplasmic Ca2+ concentration, reactive oxygen species (ROS) burst, MAPK phosphorylation, and expression of defense-related genes are not impaired in the yda11 mutant. Furthermore, the expression of CA-YDA protein does not result in enhanced PTI responses, further suggesting the existence of additional mechanisms of resistance regulated by YDA that differ from those regulated by the PTI receptors FLS2 and CERK1. In line with these observations, our transcriptomic data revealed the over-representation in CA-YDA plants of defensive genes, such as those encoding antimicrobial peptides and cell death regulators, and genes encoding cell wall-related proteins, suggesting a potential link between plant cell wall composition and integrity and broad spectrum resistance mediated by YDA. In addition, phosphoproteomic data revealed an over-representation of genes encoding wall-related proteins in CA-YDA plants in comparison with wild-type plants. The putative role of the ER-YDA pathway in regulating cell wall integrity was further supported by biochemical and glycomics analyses of er, yda11 and CA-YDA cell walls, which revealed significant changes in the cell wall composition of these genotypes compared with that of wild-type plants. In summary, our data indicate that ER and YDA are components of a novel immune pathway that regulates cell wall integrity and defensive responses, which confer broad-spectrum resistance to pathogens

Mitogen-Activated Protein Kinase Phosphatase 1 (MKP1) Negatively Regulates the Production of Reactive Oxygen Species During Arabidopsis Immune Responses

[EN] Genetic ablation of the b subunit of the heterotrimeric G protein complex in agb1-2 confers defective activation of microbe-associated molecular pattern (MAMP)-triggered immunity, resulting in agb1-2 enhanced susceptibility to pathogens like the fungus Plectosphaerella cucumerina BMM. A mutant screen for suppressors of agb1-2 susceptibility (sgb) to P. cucumerina BMM identified sgb10, a new null allele (mkp1-2) of the mitogen-activated protein kinase phosphatase 1 (MKP1). The enhanced susceptibility of agb1-2 to the bacterium Pseudomonas syringae pv. tomato DC3000 and the oomycete Hyaloperonospora arabidopsidis is also abrogated by mkp1-2. MKP1 negatively balances production of reactive oxygen species (ROS) triggered by MAMPs, since ROS levels are enhanced in mkp1. The expression of RBOHD, encoding a NADPH oxidase–producing ROS, is upregulated in mkp1 upon MAMP treatment or pathogen infection. Moreover, MKP1 negatively regulates RBOHD activity, because ROS levels upon MAMP treatment are increased in mkp1 plants constitutively overexpressing RBOHD (35S::RBOHD mkp1). A significant reprograming of mkp1 metabolic profile occurs with more than 170 metabolites, including antimicrobial compounds, showing differential accumulation in comparison with wild-type plants. These results suggest that MKP1 functions downstream of the heterotrimeric G protein during MAMP-triggered immunity, directly regulating the activity of RBOHD and ROS production as well as other immune responses.SIThis work was supported by the Secretaría de Estado de Inves-tigación, Desarrollo e Innovación grants BIO2015-64077-R and BIO2012-3291, H2020 Marie Skłodowska-Curie Actions grant (SignWALLINg-624721) from the European Union, the BRAVE Erasmus Mundi Program (European Union), and a PIF fellowship from Universidad Politécnica de Madrid. Work in the Jones lab was supported by grants from the NIGMS (R01GM065989) and National Science Foundation (MCB-0718202)