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Plant immunity regulated by cell wall integrity

Final Report Summary - SIGNWALLING (Plant immunity regulated by cell wall integrity)

In their natural environments, plants are under continuous threats caused by different attackers and stresses that compromise their survival and offspring. Constitutive expression of defensive mechanisms may not always be the best strategy against these environmental threats due to trade-offs associated to such responses. Therefore, plants have developed a repertoire of monitoring systems to sense plant morphogenesis and face the continuous environmental changes, thus initiating specific compensatory responses. This cell autonomous monitoring system comprises a collection of Pattern-Recognition Receptors (PRRs), such as Receptor-Like Protein Kinases (RLKs) and Receptor-Like Proteins (RLPs), which recognise either “plant-self” endogenous molecules (Damage-Associated Molecular Patterns, DAMPs) or Pathogen-Associated Molecular Patterns (PAMPs), activating Protein Kinase (PK) cascades, which in turn regulate downstream immune or plant growth-associated responses. The plant cell wall, a dynamic and complex structure surrounding every plant cell, is among the plant monitoring systems. The cell wall is constantly being remodeled during plants’ life in response to internal and external constraints. Therefore, plants have evolved a dedicated cell wall integrity (CWI) maintenance mechanism, that integrates many different signals into overreaching signaling systems. Alteration of CWI induces a variety of innate immune and growth responses. The EU-funded IEF-SignWALLINg has explored some of the CWI-mediated mechanisms of plant resistance against pathogens and has unveiled some of its molecular basis.
The host laboratory at Centro de Biotecnología y Genómica de Plantas (Centre for Plant Biotechnology and Genomics, CBGP) holds a collection of over 70 Arabidopsis thaliana cell wall mutants. Resistance screenings against the pathogens Plectosphaerella cucumerina, Ralstonia solanacearum and Hyaloperonospora arabidopsidis have allowed to place the mutants in patho-clusters according to their enhanced resistance, which in some cases is uncoupled from their biomass or seed production. Based on the patho-cluster selection cell wall modifications from 30 mutants were monitored by Fourier Transform InfraRed (FTIR) spectroscopy, which is able to non-destructively recognize polymers and functional groups and provide abundant information about their in muro organization from very small amounts of material. Principal component analysis of the FTIR data allowed to reduce the number of mutants to 10, which were further biochemically analyzed by an array of techniques established at the host laboratory. Given the big amount of data generated, a rigorous mathematical analysis was demanded, and this was performed in collaboration with scientists from Universidad Complutense de Madrid. Modelling the data generated has allowed to unveil the key importance of a handful of cell wall epitopes in regulating plant resistance/fitness phenotypes.
DAMPs comprise plant cell wall-derived molecules or peptides that are released or synthesized, respectively, upon pathogen infection or wounding. Despite the relevant role of wall derived DAMPs in plant-pathogen interactions, a very limited number of them, such as oligogalacturonides (OGs), has been well characterized in the past. Based in our predictive model, we selected two cell wall mutants and have characterized the molecular basis of the wall-mediated resistance found in them. Several chemically extracted wall fractions were further chromatographically purified and tested for their capacity to activate plant Pattern Triggered Immunity (PTI). Interestingly, we found that mutant wall fractions enriched in different wall components were more active than the corresponding wild-type ones in activating such responses. These included intracellular calcium accumulation, phosphorylation by mitogen-activated protein kinases (MAPKs) and transcriptional regulation of immunity genes. The immune active components in these fractions were shown to be of a carbohydrate nature. Monosaccharide and glycosidic linkage analyses were performed in order to elucidate the structures of such immune active DAMPs. Transcriptomic RNAseq data confirmed that these novel wall DAMPs activate immune signaling pathways in a different way from that triggered by OGs. The novel carbohydrate DAMP structures elucidated during the tenure of SignWALLINg will be further tested for their capacity to protect crops against pathogens in collaboration with SMEs with expertise in crop protection.