Modulation of plant autophagy during pathogen attack

In plants, the highly conserved catabolic process of autophagy has long been known as a means of maintaining cellular homeostasis and coping with abiotic stress conditions. Accumulating evidence has linked autophagy to immunity against invading pathogens, regulating plant cell death, and antimicrobial defences. In turn, it appears that phytopathogens have evolved ways not only to evade autophagic clearance but also to modulate and co-opt autophagy for their own benefit. In this review, we summarize and discuss the emerging discoveries concerning how pathogens modulate both host and self-autophagy machineries to colonize their host plants, delving into the arms race that determines the fate of interorganismal interaction.Fil: Leary, Alexandre Y. Imperial College London; Reino UnidoFil: Sanguankiattichai, Nattapong. University of Oxford; Reino UnidoFil: Duggan, Cian. Imperial College London; Reino UnidoFil: Tumtas, Yasin. Imperial College London; Reino UnidoFil: Pandey, Pooja. Imperial College London; Reino UnidoFil: Segretin, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Salguero Linares, Jose. Imperial College London; Reino UnidoFil: Savage, Zachary D. Imperial College London; Reino UnidoFil: Yow, Rui Jin. Imperial College London; Reino UnidoFil: Bozkurt, Tolga O.. Imperial College London; Reino Unid

Arabidopsis metacaspase MC1 localizes in stress granules, clears protein aggregates and delays senescence

Stress granules (SGs) are highly conserved cytoplasmic condensates that assemble in response to stress and contribute to maintaining protein homeostasis. These membraneless organelles are dynamic, disassembling once the stress is no longer present. Persistence of SGs due to mutations or chronic stress has been often related to age-dependent protein-misfolding diseases in animals. Here, we find that the metacaspase MC1 is dynamically recruited into SGs upon proteotoxic stress in Arabidopsis (Arabidopsis thaliana). Two predicted disordered regions, the prodomain and the 360 loop, mediate MC1 recruitment to and release from SGs. Importantly, we show that MC1 has the capacity to clear toxic protein aggregates in vivo and in vitro, acting as a disaggregase. Finally, we demonstrate that overexpressing MC1 delays senescence and this phenotype is dependent on the presence of the 360 loop and an intact catalytic domain. Together, our data indicate that MC1 regulates senescence through its recruitment into SGs and this function could potentially be linked to its remarkable protein aggregate-clearing activity

A phloem‐localized Arabidopsis metacaspase (AtMC3) improves drought tolerance

Increasing drought phenomena pose a serious threat to agricultural productivity. Although plants have multiple ways to respond to the complexity of drought stress, the underlying mechanisms of stress sensing and signaling remain unclear. The role of the vasculature, in particular the phloem, in facilitating inter-organ communication is critical and poorly understood.Combining genetic, proteomic and physiological approaches, we investigated the role of AtMC3, a phloem-specific member of the metacaspase family, in osmotic stress responses in Arabidopsis thaliana. Analyses of the proteome in plants with altered AtMC3 levels revealed differential abundance of proteins related to osmotic stress pointing into a role of the protein in water-stress-related responses.Overexpression of AtMC3 conferred drought tolerance by enhancing the differentiation of specific vascular tissues and maintaining higher levels of vascular-mediated transportation, while plants lacking the protein showed an impaired response to drought and inability to respond effectively to the hormone abscisic acid.Overall, our data highlight the importance of AtMC3 and vascular plasticity in fine-tuning early drought responses at the whole plant level without affecting growth or yield.ISSN:0028-646XISSN:1469-813

The Changing Landscape for Stroke\ua0Prevention in AF: Findings From the GLORIA-AF Registry Phase 2

Background GLORIA-AF (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients with Atrial Fibrillation) is a prospective, global registry program describing antithrombotic treatment patterns in patients with newly diagnosed nonvalvular atrial fibrillation at risk of stroke. Phase 2 began when dabigatran, the first non\u2013vitamin K antagonist oral anticoagulant (NOAC), became available. Objectives This study sought to describe phase 2 baseline data and compare these with the pre-NOAC era collected during phase 1. Methods During phase 2, 15,641 consenting patients were enrolled (November 2011 to December 2014); 15,092 were eligible. This pre-specified cross-sectional analysis describes eligible patients\u2019 baseline characteristics. Atrial fibrillation disease characteristics, medical outcomes, and concomitant diseases and medications were collected. Data were analyzed using descriptive statistics. Results Of the total patients, 45.5% were female; median age was 71 (interquartile range: 64, 78) years. Patients were from Europe (47.1%), North America (22.5%), Asia (20.3%), Latin America (6.0%), and the Middle East/Africa (4.0%). Most had high stroke risk (CHA2DS2-VASc [Congestive heart failure, Hypertension, Age  6575 years, Diabetes mellitus, previous Stroke, Vascular disease, Age 65 to 74 years, Sex category] score  652; 86.1%); 13.9% had moderate risk (CHA2DS2-VASc = 1). Overall, 79.9% received oral anticoagulants, of whom 47.6% received NOAC and 32.3% vitamin K antagonists (VKA); 12.1% received antiplatelet agents; 7.8% received no antithrombotic treatment. For comparison, the proportion of phase 1 patients (of N = 1,063 all eligible) prescribed VKA was 32.8%, acetylsalicylic acid 41.7%, and no therapy 20.2%. In Europe in phase 2, treatment with NOAC was more common than VKA (52.3% and 37.8%, respectively); 6.0% of patients received antiplatelet treatment; and 3.8% received no antithrombotic treatment. In North America, 52.1%, 26.2%, and 14.0% of patients received NOAC, VKA, and antiplatelet drugs, respectively; 7.5% received no antithrombotic treatment. NOAC use was less common in Asia (27.7%), where 27.5% of patients received VKA, 25.0% antiplatelet drugs, and 19.8% no antithrombotic treatment. Conclusions The baseline data from GLORIA-AF phase 2 demonstrate that in newly diagnosed nonvalvular atrial fibrillation patients, NOAC have been highly adopted into practice, becoming more frequently prescribed than VKA in Europe and North America. Worldwide, however, a large proportion of patients remain undertreated, particularly in Asia and North America. (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients With Atrial Fibrillation [GLORIA-AF]; NCT01468701

TRY plant trait database – enhanced coverage and open access

Plant traits - the morphological, anatomical, physiological, biochemical and phenological characteristics of plants - determine how plants respond to environmental factors, affect other trophic levels, and influence ecosystem properties and their benefits and detriments to people. Plant trait data thus represent the basis for a vast area of research spanning from evolutionary biology, community and functional ecology, to biodiversity conservation, ecosystem and landscape management, restoration, biogeography and earth system modelling. Since its foundation in 2007, the TRY database of plant traits has grown continuously. It now provides unprecedented data coverage under an open access data policy and is the main plant trait database used by the research community worldwide. Increasingly, the TRY database also supports new frontiers of trait‐based plant research, including the identification of data gaps and the subsequent mobilization or measurement of new data. To support this development, in this article we evaluate the extent of the trait data compiled in TRY and analyse emerging patterns of data coverage and representativeness. Best species coverage is achieved for categorical traits - almost complete coverage for ‘plant growth form’. However, most traits relevant for ecology and vegetation modelling are characterized by continuous intraspecific variation and trait–environmental relationships. These traits have to be measured on individual plants in their respective environment. Despite unprecedented data coverage, we observe a humbling lack of completeness and representativeness of these continuous traits in many aspects. We, therefore, conclude that reducing data gaps and biases in the TRY database remains a key challenge and requires a coordinated approach to data mobilization and trait measurements. This can only be achieved in collaboration with other initiatives

Understanding plant immunity through the identification of immunogenic cell death indicators and the characterization of plant metacaspases in Arabidopsis thaliana

Els patògens vegetals representen una gran amenaça per la seguretat alimentària, causant al voltant del 20-40% de pèrdues en la producció de cultius. En el context actual de canvi climàtic, els patògens, que evolucionen ràpidament, poden superar fàcilment la resistència proporcionada pels pesticides tradicionals. Per tant, entendre exhaustivament el sistema immunitari de les plantes és de gran importància per generar cultius resistents. Com a estratègia per combatre la invasió de patògens, les cèl·lules vegetals infectades desencadenen un tipus de mort cel·lular regulada coneguda com a resposta hipersensible (HR). La regulació de la HR és essencial per confinar la resposta immunitària exclusivament al lloc d’entrada del patogen. Actualment, el nostre coneixement de com es produeix la zonació de la mort cel·lular i com responen les cèl·lules veïnes a l’infecció és escàs. En el primer capítol de la meva tesi, he explorat com la HR es regula de manera espaciotemporal a nivell transcripcional a la planta model Arabidopsis thaliana. Aquests resultats ens van permetre identificar marcadors transcripcionals específics de la HR. A més, proporcionem a la comunitat una línia transgènica reportera fluorescent que mostra una forta senyal espaciotemporal en cèl·lules destinades a patir HR. L’ús d’aquesta línia reportera per tècniques específiques que involucrin estudis “òmics” de cèl·lules individuals permetrà una major comprensió del caracter zonal de la immunitat vegetal. En la darrera dècada, un nombre creixent d’estudis suggereixen que les proteases desenvolupen papers fonamentals durant la HR. En el segon capítol de la meva tesi, he estudiat el paper de la metacaspasa 1 (AtMC1) d’Arabidopsis, un tipus de cisteïna proteasa, en la immunitat vegetal. Originalment, es va descriure a la AtMC1 com un regulador positiu de la HR en plantes joves, per contra, les plantes adultes que no tenen AtMC1 mostren una activació constitutiva de la immunitat en condicions basals, actuant així com un regulador negatiu de la immunitat. Les mutacions en el lloc catalític de la proteasa desencadenen una autoimmunitat greu. A través d’una combinació d’experiments genètics, bioquímics i de biologia cel·lular, mostrem que la versió catalíticament inactiva d’AtMC1 actua com una plataforma d’acoblament per a components relacionats amb la immunitat, inclosos receptors i.possiblement, evita la seva correcta degradació. Basant-nos en aquestes dades i en la literatura anterior, inferim que l’AtMC1 podria controlar directa o indirectament l’homeostasi dels receptors d’immunitat. Per tant, interferir amb la funció de la proteasa wild-type té un impacte negatiu en el creixement de la planta. Finalment, he participat en una segona línia d’investigació on hem intentat entendre la funció d’AtMC1 en condicions d’estrés proteotòxic. AtMC1 és dinàmicament reclutada a condensats citoplasmàtics, coneguts com granuls d’estrés, regulant la senescència (Capítol 3). Per caracteritzar bioquímicament aquesta funció, vam eliminar alguns dominis predits com a altament propensos a l’agregació i vam aconseguir expressar i aïllar la proteasa de forma recombinant. Això ens va permetre demostrar que AtMC1 té una gran capacitat, evolutivament conservada, de netejar agregats de proteïnes, inclosos els formats per formes de proteïnes patològiques que causen malalties mortals en humans. La implementació de proteïnes recombinants amb alta activitat de neteja d’agregats pot obrir noves vies per a la intervenció terapèutica en malalties causades per proteïnes mal plegades. En els darrers cinc anys, la comunitat científica de plantes ha presenciat un gran progrés en la nostra comprensió del sistema immunitari vegetal gràcies a estudis mecanístics dels receptors d’immunitat i vies de senyalització que regulen i condueixen la HR. Aprofitar aquest coneixement per aconseguir resistència a malalties en cultius bàsics i econòmicament importants serà una prioritat en els pròxims anys. Espero que els treballs presentats en aquesta tesi puguin contribuir a aconseguir aquests ambiciosos objectius.Los patógenos vegetales representan una gran amenaza para la seguridad alimentaria, causando alrededor del 20-40% de pérdidas en la producción de cultivos. En un contexto de cambio climático, los patógenos que evolucionan rápidamente pueden superar fácilmente la resistencia proporcionada por los pesticidas tradicionales. En consecuencia, comprender exhaustivamente el sistema inmune de las plantas es de suma importancia para generar cultivos resistentes. Como estrategia para contrarrestar la invasión de patógenos, las células vegetales infectadas desencadenan un tipo de muerte celular regulada conocida como la respuesta hipersensible (HR). La regulación de la HR es esencial para confinar la respuesta inmune exclusivamente al lugar de ingreso del patógeno. Sin embargo, nuestra comprensión de cómo se logra la zonificación de la muerte celular y cómo las células vecinas responden a la infección es fragmentaria. En el primer capítulo de mi tesis, exploro cómo la HR se regula espacio-temporalmente a nivel transcripcional en la planta modelo Arabidopsis thaliana. Estos resultados nos permitieron identificar indicadores transcripcionales genuinos de la HR. Además, proporcionamos a la comunidad una línea transgénica reportera fluorescente que muestra un fuerte señal espacio-temporal en células destinadas a sufrir HR. El uso de esta línea reportera para técnicas específicas que involucren estudios “ómicos” de células individuales permitirá una mayor disección del aspecto espacial de la inmunidad vegetal. En la última década, evidencias crecientes sugieren que las proteasas desempeñan papeles cruciales durante la HR. En el segundo capítulo de mi tesis, intento desentrañar el papel de la metacaspasa 1 (AtMC1) de Arabidopsis, un tipo de cisteína proteasa, en la inmunidad vegetal. Aunque originalmente se describió como un regulador positivo de la HR en plantas jóvenes, plantas adultas que carecen de AtMC1 exhiben una activación constitutiva de la inmunidad en condiciones basales, actuando por consiguiente como un regulador negativo de la inmunidad. Mutaciones en el sito catalítico de la proteasa desencadenan una autoinmunidad grave. A través de una combinación de experimentos genéticos, bioquímicos y de biología celular, mostramos que la versión cataliticamente inactiva de AtMC1 actúa como un sitio de acoplamiento pegajoso para componentes relacionados con la inmunidad, incluidos receptores inmunes, posiblemente evitando su oportuna degradación. En base a estos datos y a la literatura previa, inferimos que AtMC1 podría controlar directa o indirectamente la homeostasis de los receptores inmunes. Por lo tanto, interferir con la función de la proteasa wild-type tiene impactos negativos en el crecimiento de la planta. Finalmente, participé en una segunda línea de investigación en la que intentamos entender la función de AtMC1 ante el estrés proteotóxico. AtMC1 es dinámicamente reclutada a condensados citoplásmicos altamente conservados, conocidos como granulos de estrés, regulando la senescencia (Capítulo 3). Para caracterizar bioquímicamente esta función, eliminamos ciertos dominios previstos de ser altamente propensos a la agregación y logramos expresar e aislar la proteasa de forma recombinante. Esto nos permitió demostrar que AtMC1 tiene una gran capacidad, evolutivamente conservada, de limpiar agregados de proteínas, incluidos los formados por formas de proteínas patológicas que causan enfermedades mortales en los humanos. La implementación de proteínas recombinantes con alta actividad de limpieza de agregados puede abrir nuevas vías para la intervención terapéutica en enfermedades causadas por proteínas mal plegadas. En los últimos cinco años, la comunidad científica de plantas ha presenciado un gran avance en nuestra comprensión del sistema inmune vegetal gracias a estudios mecanísticos en receptores inmunes y vías de señalización que regulan y conducen a la HR. Aprovechar este conocimiento para lograr resistencia a enfermedades en cultivos básicos y económicamente importantes será una prioridad en los próximos años. Espero que los trabajos presentados en esta tesis puedan contribuir a futuros esfuerzos para lograr esos ambiciosos objetivos.Losses to plant pathogens pose a major threat to food security, bringing about serious economic and societal burdens across the globe. with around 20-40% of crop production lost to plant diseases. In the face of climate change, rapidly evolving pathogens can easily overcome resistance provided by traditional pesticides. Consequently, a thorough understanding of the plant immune system is of paramount importance to breed disease-resistant crops. As a strategy to counteract pathogen invasion, infected plant cells elicit type of regulated cell death known as the hypersensitive response (HR). Tight regulation of HR is critical for confinement of the immune response exclusively to the pathogen ingress site. However, our understanding of how cell death zonation is achieved and how by-stander cells (neighbouring cells) respond to infection remains fragmentary. In the first chapter of my PhD thesis, I explored how HR is spatiotemporally regulated at the transcriptional level in the plant model Arabidopsis thaliana. These results allowed us to identify bona fide transcriptional indicators of HR. Moreover, we provide for the community a fluorescent reporter transgenic line that displays a strong spatiotemporally resolved signal specifically in cells destined to undergo HR. Use of this reporter line for specific and-or high-throughput techniques involving single-cell “omics” will enable further dissection of the spatial aspect of plant immunity. Over the last decade, accumulating evidence suggest that plant proteases play crucial roles during HR. In the second chapter of my PhD, I attempt to unravel the role of Arabidopsis metacaspase 1 (AtMC1), a type of cysteine protease, in plant immunity. Although originally described as a positive regulator of HR in young plants, adult plants lacking AtMC1 exhibit constitutive activation of immune responses under basal conditions, thus acting as a negative regulator of plant immunity. We report that mutating the catalytic cysteine of the protease causes severe autoimmunity. Through a combination of genetic, biochemical and cell biology experiments we show that catalytically inactive AtMC1 acts as a sticky docking site for immune-related components including immune receptors, possibly preventing their timely turnover. Based on this data and previous literature, we infer that AtMC1 might directly or indirectly control the homeostasis of immune receptors. Therefore, interfering with the wild-type function of the protease has negative impacts on plant growth. Finally, I actively participated in a second line of research in which we try to understand the function of AtMC1 upon proteotoxic stress. AtMC1 is dynamically recruited to highly conserved cytoplasmic condensates, known as stress granules, regulating senescence (Chapter 3). To biochemically characterize this function, we removed certain domains predicted to be high aggregation-prone and successfully expressed and isolated the protease. This major step forward allowed us to prove that AtMC1 exhibits a strong and evolutionary conserved capacity to clear protein aggregates, including those formed by pathological protein forms that cause a diversity of life-threatening pathologies in humans. The implementation of recombinant proteins with high aggregate-clearance activity may open new avenues for therapeutic intervention in diseases caused by misfolded proteins. In the last five years, the plant science community has particularly witnessed a quantum leap in our understanding of the plant immune system thanks to mechanistic studies on plant immune receptors and signalling pathways regulating and leading to HR. Leveraging this knowledge to engineer disease resistance in staples and economically important crops will be a priority in the years to come. I hope that the works herein and conclusions drawn from this thesis can contribute to future endeavours to achieve such ambitious goals.Universitat Autònoma de Barcelona. Programa de Doctorat en Biologia i Biotecnologia Vegeta

Understanding plant immunity through the identification of immunogenic cell death indicators and the characterization of plant metacaspases in Arabidopsis thaliana

Els patògens vegetals representen una gran amenaça per la seguretat alimentària, causant al voltant del 20-40% de pèrdues en la producció de cultius. En el context actual de canvi climàtic, els patògens, que evolucionen ràpidament, poden superar fàcilment la resistència proporcionada pels pesticides tradicionals. Per tant, entendre exhaustivament el sistema immunitari de les plantes és de gran importància per generar cultius resistents. Com a estratègia per combatre la invasió de patògens, les cèl·lules vegetals infectades desencadenen un tipus de mort cel·lular regulada coneguda com a resposta hipersensible (HR). La regulació de la HR és essencial per confinar la resposta immunitària exclusivament al lloc d'entrada del patogen. Actualment, el nostre coneixement de com es produeix la zonació de la mort cel·lular i com responen les cèl·lules veïnes a l'infecció és escàs. En el primer capítol de la meva tesi, he explorat com la HR es regula de manera espaciotemporal a nivell transcripcional a la planta model Arabidopsis thaliana. Aquests resultats ens van permetre identificar marcadors transcripcionals específics de la HR. A més, proporcionem a la comunitat una línia transgènica reportera fluorescent que mostra una forta senyal espaciotemporal en cèl·lules destinades a patir HR. L'ús d'aquesta línia reportera per tècniques específiques que involucrin estudis "òmics" de cèl·lules individuals permetrà una major comprensió del caracter zonal de la immunitat vegetal. En la darrera dècada, un nombre creixent d'estudis suggereixen que les proteases desenvolupen papers fonamentals durant la HR. En el segon capítol de la meva tesi, he estudiat el paper de la metacaspasa 1 (AtMC1) d'Arabidopsis, un tipus de cisteïna proteasa, en la immunitat vegetal. Originalment, es va descriure a la AtMC1 com un regulador positiu de la HR en plantes joves, per contra, les plantes adultes que no tenen AtMC1 mostren una activació constitutiva de la immunitat en condicions basals, actuant així com un regulador negatiu de la immunitat. Les mutacions en el lloc catalític de la proteasa desencadenen una autoimmunitat greu. A través d'una combinació d'experiments genètics, bioquímics i de biologia cel·lular, mostrem que la versió catalíticament inactiva d'AtMC1 actua com una plataforma d'acoblament per a components relacionats amb la immunitat, inclosos receptors i.possiblement, evita la seva correcta degradació. Basant-nos en aquestes dades i en la literatura anterior, inferim que l'AtMC1 podria controlar directa o indirectament l'homeostasi dels receptors d'immunitat. Per tant, interferir amb la funció de la proteasa wild-type té un impacte negatiu en el creixement de la planta. Finalment, he participat en una segona línia d'investigació on hem intentat entendre la funció d'AtMC1 en condicions d'estrés proteotòxic. AtMC1 és dinàmicament reclutada a condensats citoplasmàtics, coneguts com granuls d'estrés, regulant la senescència (Capítol 3). Per caracteritzar bioquímicament aquesta funció, vam eliminar alguns dominis predits com a altament propensos a l'agregació i vam aconseguir expressar i aïllar la proteasa de forma recombinant. Això ens va permetre demostrar que AtMC1 té una gran capacitat, evolutivament conservada, de netejar agregats de proteïnes, inclosos els formats per formes de proteïnes patològiques que causen malalties mortals en humans. La implementació de proteïnes recombinants amb alta activitat de neteja d'agregats pot obrir noves vies per a la intervenció terapèutica en malalties causades per proteïnes mal plegades. En els darrers cinc anys, la comunitat científica de plantes ha presenciat un gran progrés en la nostra comprensió del sistema immunitari vegetal gràcies a estudis mecanístics dels receptors d'immunitat i vies de senyalització que regulen i condueixen la HR. Aprofitar aquest coneixement per aconseguir resistència a malalties en cultius bàsics i econòmicament importants serà una prioritat en els pròxims anys. Espero que els treballs presentats en aquesta tesi puguin contribuir a aconseguir aquests ambiciosos objectius.Los patógenos vegetales representan una gran amenaza para la seguridad alimentaria, causando alrededor del 20-40% de pérdidas en la producción de cultivos. En un contexto de cambio climático, los patógenos que evolucionan rápidamente pueden superar fácilmente la resistencia proporcionada por los pesticidas tradicionales. En consecuencia, comprender exhaustivamente el sistema inmune de las plantas es de suma importancia para generar cultivos resistentes. Como estrategia para contrarrestar la invasión de patógenos, las células vegetales infectadas desencadenan un tipo de muerte celular regulada conocida como la respuesta hipersensible (HR). La regulación de la HR es esencial para confinar la respuesta inmune exclusivamente al lugar de ingreso del patógeno. Sin embargo, nuestra comprensión de cómo se logra la zonificación de la muerte celular y cómo las células vecinas responden a la infección es fragmentaria. En el primer capítulo de mi tesis, exploro cómo la HR se regula espacio-temporalmente a nivel transcripcional en la planta modelo Arabidopsis thaliana. Estos resultados nos permitieron identificar indicadores transcripcionales genuinos de la HR. Además, proporcionamos a la comunidad una línea transgénica reportera fluorescente que muestra un fuerte señal espacio-temporal en células destinadas a sufrir HR. El uso de esta línea reportera para técnicas específicas que involucren estudios "ómicos" de células individuales permitirá una mayor disección del aspecto espacial de la inmunidad vegetal. En la última década, evidencias crecientes sugieren que las proteasas desempeñan papeles cruciales durante la HR. En el segundo capítulo de mi tesis, intento desentrañar el papel de la metacaspasa 1 (AtMC1) de Arabidopsis, un tipo de cisteína proteasa, en la inmunidad vegetal. Aunque originalmente se describió como un regulador positivo de la HR en plantas jóvenes, plantas adultas que carecen de AtMC1 exhiben una activación constitutiva de la inmunidad en condiciones basales, actuando por consiguiente como un regulador negativo de la inmunidad. Mutaciones en el sito catalítico de la proteasa desencadenan una autoinmunidad grave. A través de una combinación de experimentos genéticos, bioquímicos y de biología celular, mostramos que la versión cataliticamente inactiva de AtMC1 actúa como un sitio de acoplamiento pegajoso para componentes relacionados con la inmunidad, incluidos receptores inmunes, posiblemente evitando su oportuna degradación. En base a estos datos y a la literatura previa, inferimos que AtMC1 podría controlar directa o indirectamente la homeostasis de los receptores inmunes. Por lo tanto, interferir con la función de la proteasa wild-type tiene impactos negativos en el crecimiento de la planta. Finalmente, participé en una segunda línea de investigación en la que intentamos entender la función de AtMC1 ante el estrés proteotóxico. AtMC1 es dinámicamente reclutada a condensados citoplásmicos altamente conservados, conocidos como granulos de estrés, regulando la senescencia (Capítulo 3). Para caracterizar bioquímicamente esta función, eliminamos ciertos dominios previstos de ser altamente propensos a la agregación y logramos expresar e aislar la proteasa de forma recombinante. Esto nos permitió demostrar que AtMC1 tiene una gran capacidad, evolutivamente conservada, de limpiar agregados de proteínas, incluidos los formados por formas de proteínas patológicas que causan enfermedades mortales en los humanos. La implementación de proteínas recombinantes con alta actividad de limpieza de agregados puede abrir nuevas vías para la intervención terapéutica en enfermedades causadas por proteínas mal plegadas. En los últimos cinco años, la comunidad científica de plantas ha presenciado un gran avance en nuestra comprensión del sistema inmune vegetal gracias a estudios mecanísticos en receptores inmunes y vías de señalización que regulan y conducen a la HR. Aprovechar este conocimiento para lograr resistencia a enfermedades en cultivos básicos y económicamente importantes será una prioridad en los próximos años. Espero que los trabajos presentados en esta tesis puedan contribuir a futuros esfuerzos para lograr esos ambiciosos objetivos.Losses to plant pathogens pose a major threat to food security, bringing about serious economic and societal burdens across the globe. with around 20-40% of crop production lost to plant diseases. In the face of climate change, rapidly evolving pathogens can easily overcome resistance provided by traditional pesticides. Consequently, a thorough understanding of the plant immune system is of paramount importance to breed disease-resistant crops. As a strategy to counteract pathogen invasion, infected plant cells elicit type of regulated cell death known as the hypersensitive response (HR). Tight regulation of HR is critical for confinement of the immune response exclusively to the pathogen ingress site. However, our understanding of how cell death zonation is achieved and how by-stander cells (neighbouring cells) respond to infection remains fragmentary. In the first chapter of my PhD thesis, I explored how HR is spatiotemporally regulated at the transcriptional level in the plant model Arabidopsis thaliana. These results allowed us to identify bona fide transcriptional indicators of HR. Moreover, we provide for the community a fluorescent reporter transgenic line that displays a strong spatiotemporally resolved signal specifically in cells destined to undergo HR. Use of this reporter line for specific and-or high-throughput techniques involving single-cell "omics" will enable further dissection of the spatial aspect of plant immunity. Over the last decade, accumulating evidence suggest that plant proteases play crucial roles during HR. In the second chapter of my PhD, I attempt to unravel the role of Arabidopsis metacaspase 1 (AtMC1), a type of cysteine protease, in plant immunity. Although originally described as a positive regulator of HR in young plants, adult plants lacking AtMC1 exhibit constitutive activation of immune responses under basal conditions, thus acting as a negative regulator of plant immunity. We report that mutating the catalytic cysteine of the protease causes severe autoimmunity. Through a combination of genetic, biochemical and cell biology experiments we show that catalytically inactive AtMC1 acts as a sticky docking site for immune-related components including immune receptors, possibly preventing their timely turnover. Based on this data and previous literature, we infer that AtMC1 might directly or indirectly control the homeostasis of immune receptors. Therefore, interfering with the wild-type function of the protease has negative impacts on plant growth. Finally, I actively participated in a second line of research in which we try to understand the function of AtMC1 upon proteotoxic stress. AtMC1 is dynamically recruited to highly conserved cytoplasmic condensates, known as stress granules, regulating senescence (Chapter 3). To biochemically characterize this function, we removed certain domains predicted to be high aggregation-prone and successfully expressed and isolated the protease. This major step forward allowed us to prove that AtMC1 exhibits a strong and evolutionary conserved capacity to clear protein aggregates, including those formed by pathological protein forms that cause a diversity of life-threatening pathologies in humans. The implementation of recombinant proteins with high aggregate-clearance activity may open new avenues for therapeutic intervention in diseases caused by misfolded proteins. In the last five years, the plant science community has particularly witnessed a quantum leap in our understanding of the plant immune system thanks to mechanistic studies on plant immune receptors and signalling pathways regulating and leading to HR. Leveraging this knowledge to engineer disease resistance in staples and economically important crops will be a priority in the years to come. I hope that the works herein and conclusions drawn from this thesis can contribute to future endeavours to achieve such ambitious goals

Cell death as a defense strategy against pathogens in plants and animals

Eukaryotes are endowed with sophisticated innate immune systems to recognize non-self and halt pathogen proliferation. Activation of cell death at the site of attempted pathogen ingress is a common strategy used by plants and animals to restrict pathogen proliferation and trigger immune responses in the surrounding tissues. As such, immunogenic cell death shares several features in both plants and animals that will be discussed in this article, namely: (i) it is triggered by activation of NLR immune receptors-often through oligomerization; (ii) it results in disruption of the plasma membrane (PM)/endomembrane integrity driving an imbalance in ion fluxes; and (iii) it results in the release of signaling molecules from dying cells

Cell death as a defense strategy against pathogens in plants and animals.

Eukaryotes are endowed with sophisticated innate immune systems to recognize non-self and halt pathogen proliferation. Activation of cell death at the site of attempted pathogen ingress is a common strategy used by plants and animals to restrict pathogen proliferation and trigger immune responses in the surrounding tissues. As such, immunogenic cell death shares several features in both plants and animals that will be discussed in this article, namely: (i) it is triggered by activation of NLR immune receptors-often through oligomerization; (ii) it results in disruption of the plasma membrane (PM)/endomembrane integrity driving an imbalance in ion fluxes; and (iii) it results in the release of signaling molecules from dying cells

Cell death as a defense strategy against pathogens in plants and animals

Eukaryotes are endowed with sophisticated innate immune systems to recognize non-self and halt pathogen proliferation. Activation of cell death at the site of attempted pathogen ingress is a common strategy used by plants and animals to restrict pathogen proliferation and trigger immune responses in the surrounding tissues. As such, immunogenic cell death shares several features in both plants and animals that will be discussed in this article, namely: (i) it is triggered by activation of NLR immune receptors—often through oligomerization; (ii) it results in disruption of the plasma membrane (PM)/endomembrane integrity driving an imbalance in ion fluxes; and (iii) it results in the release of signaling molecules from dying cells