Loss of the arabidopsis protein kinases ANPs affects root cell wall composition, and triggers the cell wall damage syndrome

The Arabidopsis NPK1-related Protein kinases ANP1, ANP2 and ANP3 belong to the MAP kinase kinase kinase (MAPKKK) superfamily and were previously described to be crucial for cytokinesis, elicitor-induced immunity and development. Here we investigate the basis of their role in development by using conditional β-estradiol-inducible triple mutants to overcome lethality. In seedlings, lack of ANPs causes root cell bulging, with the transition zone being the most sensitive region. We uncover a role of ANPs in the regulation of cell wall composition and suggest that developmental defects of the triple mutants, observed at the cellular level, might be a consequence of the alterations of the pectic and cellulosic cell wall components. Lack of ANPs also induced a typical cell wall damage syndrome (CWDS) similar to that observed in plants treated with the cellulose biosynthesis inhibitor isoxaben (ISX). Moreover, anp double mutants and plants overexpressing single ANPs (ANP1 or ANP3) respectively showed increased and reduced accumulation of jasmonic acid and PDF1.2 transcripts upon ISX treatment, suggesting that ANPs are part of the pathway targeted by this inhibitor and play a role in cell wall integrity surveillance

Luminol-based Assay for Detection of Immunity Elicitor-induced Hydrogen Peroxide Production in Arabidopsis thaliana Leaves

In Arabidopsis thaliana, one of the very early immune-related responses induced after elicitor perception is the oxidative burst, i.e. reactive oxygen species (ROS) generation including superoxide anion and hydrogen peroxide (H2O2). ROS production plays different roles in a wide range of biotic and abiotic stress responses, including the closure of stomata and the regulation of cell expansion. In particular, elicitor-induced H2O2 is produced mainly by the membrane localized NAD(P)H oxidases RESPIRATORY BURST OXIDASE HOMOLOGUE D and F. In this protocol, we describe a simple and reproducible luminol/peroxidase-based assay to detect and evaluate immunity-related accumulation of H2O2 produced in Arabidopsis leaf discs treated with immunity elicitors, such as oligogalacturonides (OGs), flagellin (flg22) or the elongation factor-thermo-unstable (EF-Tu - elf18). This method is based on the detection of the luminescence released by excited-luminol molecules generated after the horseradish peroxidase (HRP)-catalyzed oxidation of luminol molecules in the presence of H2O2. Levels as well as duration of the luminescence are proportional to the amount of H2O2 produced by elicited leaf discs

Camalexin Quantification in Arabidopsis thaliana Leaves Infected with Botrytis cinerea

Phytoalexins are heterogeneous low molecular mass secondary metabolites with antimicrobial activity produced in response to pathogen invasion attempts at the infection site and represent an important part of the plant defense repertoire. Camalexin (3-Thiazol-2′-yl-indole) is a known phytoalexin first detected and isolated in Camelina sativa, from which it takes its name, infected with Alternaria brassicae (Browne et al., 1991). Production of camalexin is also induced in Arabidopsis thaliana leaves by a range of biotrophic and necrotrophic plant pathogens (bacteria, oomycetes, fungi and viruses) (Ahuja et al., 2012) as well as by abiotic stresses, such as UV and chemicals (e.g. acifluorfen, paraquat, chlorsulfuron and α-amino butyric acid) (Zhao et al., 1998; Tierens et al., 2002). Camalexin originates from tryptophan and CYP79B2 and CYP71B15 (PAD3) are P450 enzymes that catalyze important steps in its biosynthetic pathway (Glawischnig, 2007). In this protocol the detection and quantification of camalexin produced in Arabidopsis leaves infected with the necrotrophic fungus Botrytis cinerea is described

Corrigendum: Loss of the Arabidopsis Protein Kinases ANPs Affects Root Cell Wall Composition, and Triggers the Cell Wall Damage Syndrome

The Arabidopsis NPK1-related Protein kinases ANP1, ANP2 and ANP3 belong to the MAP kinase kinase kinase (MAPKKK) superfamily and were previously described to be crucial for cytokinesis, elicitor-induced immunity and development. Here we investigate the basis of their role in development by using conditional β-estradiol-inducible triple mutants to overcome lethality. In seedlings, lack of ANPs causes root cell bulging, with the transition zone being the most sensitive region. We uncover a role of ANPs in the regulation of cell wall composition and suggest that developmental defects of the triple mutants, observed at the cellular level, might be a consequence of the alterations of the pectic and cellulosic cell wall components. Lack of ANPs also induced a typical cell wall damage syndrome (CWDS) similar to that observed in plants treated with the cellulose biosynthesis inhibitor isoxaben (ISX). Moreover, anp double mutants and plants overexpressing single ANPs (ANP1 or ANP3) respectively showed increased and reduced accumulation of jasmonic acid and PDF1.2 transcripts upon ISX treatment, suggesting that ANPs are part of the pathway targeted by this inhibitor and play a role in cell wall integrity surveillance.Highlights: The loss of ANP function affects cell wall composition and leads to typical cell wall damage-induced phenotypes, such as ectopic lignification and jasmonic acid accumulation

Fighting salt or enemies : shared perception and signaling strategies

Plants react to a myriad of biotic and abiotic environmental signals through specific cellular mechanisms required for survival under stress. Although pathogen perception has been widely studied and characterized, salt stress perception and signaling remain largely elusive. Recent observations, obtained in the model plant Arabidopsis thaliana, show that perception of specific features of pathogens also allows plants to mount salt stress resilience pathways, highlighting the possibility that salt sensing and pathogen perception mechanisms partially overlap. We discuss these overlapping strategies and examine the emerging role of A. thaliana cell wall and plasma membrane components in activating both salt- and pathogen-induced responses, as part of exquisite mechanisms underlying perception of damage and danger. This knowledge helps understanding the complexity of plant responses to pathogens and salinity, leading to new hypotheses that could explain why plants evolved similar strategies to respond to these, at first sight, very different types of stimuli

Plant cell wall integrity maintenance in model plants and crop species-relevant cell wall components and underlying guiding principles

The walls surrounding the cells of all land-based plants provide mechanical support essential for growth and development as well as protection from adverse environmental conditions like biotic and abiotic stress. Composition and structure of plant cell walls can differ markedly between cell types, developmental stages and species. This implies that wall composition and structure are actively modified during biological processes and in response to specific functional requirements. Despite extensive research in the area, our understanding of the regulatory processes controlling active and adaptive modifications of cell wall composition and structure is still limited. One of these regulatory processes is the cell wall integrity maintenance mechanism, which monitors and maintains the functional integrity of the plant cell wall during development and interaction with environment. It is an important element in plant pathogen interaction and cell wall plasticity, which seems at least partially responsible for the limited success that targeted manipulation of cell wall metabolism has achieved so far. Here, we provide an overview of the cell wall polysaccharides forming the bulk of plant cell walls in both monocotyledonous and dicotyledonous plants and the effects their impairment can have. We summarize our current knowledge regarding the cell wall integrity maintenance mechanism and discuss that it could be responsible for several of the mutant phenotypes observed

Plant cell wall integrity maintenance in model plants and crop species-relevant cell wall components and underlying guiding principles

The walls surrounding the cells of all land-based plants provide mechanical support essential for growth and development as well as protection from adverse environmental conditions like biotic and abiotic stress. Composition and structure of plant cell walls can differ markedly between cell types, developmental stages and species. This implies that wall composition and structure are actively modified during biological processes and in response to specific functional requirements. Despite extensive research in the area, our understanding of the regulatory processes controlling active and adaptive modifications of cell wall composition and structure is still limited. One of these regulatory processes is the cell wall integrity maintenance mechanism, which monitors and maintains the functional integrity of the plant cell wall during development and interaction with environment. It is an important element in plant pathogen interaction and cell wall plasticity, which seems at least partially responsible for the limited success that targeted manipulation of cell wall metabolism has achieved so far. Here, we provide an overview of the cell wall polysaccharides forming the bulk of plant cell walls in both monocotyledonous and dicotyledonous plants and the effects their impairment can have. We summarize our current knowledge regarding the cell wall integrity maintenance mechanism and discuss that it could be responsible for several of the mutant phenotypes observed.</p

Arabidopsis root responses to salinity depend on pectin modification and cell wall sensing

Owing to its detrimental effect on plant growth, salinity is an increasing worldwide problem for agriculture. To understand the molecular mechanisms activated in response to salt in Arabidopsis thaliana, we investigated the Catharanthus roseus receptor-like kinase 1-like family, which contains sensors that were previously shown to be involved in sensing the structural integrity of the cell walls. We found that herk1 the1-4 double mutants, lacking the function of HERKULES1 (HERK1) and combined with a gain-of-function allele of THESEUS1 (THE1), strongly respond to salt application, resulting in an intense activation of stress responses, similarly to plants lacking FERONIA (FER) function. We report that salt triggers pectin methyl esterase (PME) activation and show its requirement for the activation of several salt-dependent responses. Because chemical inhibition of PMEs alleviates these salt-induced responses, we hypothesize a model in which salt directly leads to cell wall modifications through the activation of PMEs. Responses to salt partly require the functionality of FER alone or HERK1/THE1 to attenuate salt effects, highlighting the complexity of the salt-sensing mechanisms that rely on cell wall integrity