The function of plant cell walls in plant-microbe interactions: Characterizing receptor-mediated cell wall integrity sensing during defense

Plants constitute the largest source of biomass on Earth. Along with being a vital part of human nutrition, plants are being used to generate the novel oils and plastics, materials, and therapeutic molecules of the future. Much of it is derived from a specialized plant cell structure called the cell wall (CW). The plant CW is composed of a complex network of polysaccharides and glycoproteins that serve as a rigid exoskeleton for plants. This structure allows plants to withstand even the harshest of environments and is the plant's first line of defense against pathogens. During plant development, the three most abundant CW polysaccharides, cellulose, hemicellulose, and pectin are known to be modified through the action of CW proteins. Pectins are composed of long chains of ɑ(1→4)-linked D-galacturonic acid and are extensively modified at the CW to alter CW properties. Pectin is deposited at the CW in a highly methylated form and is demethylesterified in muro by CW enzymes. The maintenance of pectin at the CW is vital for plant growth and development as demonstrated by the large number of pectin-associated genes present within plant species and the altered growth and defense phenotypes that arise if they are perturbed. Furthermore, microbes such as the fungal vascular pathogen Fusarium oxysporum, are known to benefit from increased pectin demethylation at the plant CW. Pectin status is monitored by plasma membrane (PM)-localized receptor proteins to maintain CW integrity (CWI). Previous work identified the protein RESISTANCE TO FUSARIUM OXYSPORUM 1 (RFO1) as a putative wall-associated kinase-like receptor required for full resistance to F. oxysporum in Arabidopsis thaliana. Using a newly developed plate infection assay described here, rfo1-1 was found to suffer altered root growth responses and vascular colonization upon F. oxysporum 5176 (Fo5176) infection and decreased sensitivity to pharmacological and genetically-induced changes in pectin methylation. Natural variants of RFO1 were shown to differ in their capacity to sense pectin and defend against Fo5176 possibly due to altered protein-protein interactions. Furthermore, using confocal microscopy coupled with single-particle tracking tools RFO1 particle dynamics at the PM were found to respond to altered pectin methylation at the CW. Using in vitro biochemical methods, the ectodomain of RFO1 was shown to bind to demethylesterified pectin. Likewise, RFO1 was found to bind to CW-derived pectin fractions according to the degree of pectin methylation present before and after Fo5176 infection. Biochemical and gene expression analyses revealed that RFO1 is involved in modulating the activation of downstream CWI and defense pathways upon treatment with exogenous demethylated pectin. The work of this thesis characterizes RFO1 as a novel pectin integrity sensor that has evolved in A. thaliana to perceive CWI and function in defense against Fo5176 infection. Furthermore, it provides a road map of RFO1 amino acid adaptations that could provide future insights into RFO1 function in pectin sensing and defense

Pathogen‐induced pH changes regulate the growth‐defense balance in plants

Environmental adaptation of organisms relies on fast perception and response to external signals, which lead to developmental changes. Plant cell growth is strongly dependent on cell wall remodeling. However, little is known about cell wall‐related sensing of biotic stimuli and the downstream mechanisms that coordinate growth and defense responses. We generated genetically encoded pH sensors to determine absolute pH changes across the plasma membrane in response to biotic stress. A rapid apoplastic acidification by phosphorylation‐based proton pump activation in response to the fungus Fusarium oxysporum immediately reduced cellulose synthesis and cell growth and, furthermore, had a direct influence on the pathogenicity of the fungus. In addition, pH seems to influence cellulose structure. All these effects were dependent on the COMPANION OF CELLULOSE SYNTHASE proteins that are thus at the nexus of plant growth and defense. Hence, our discoveries show a remarkable connection between plant biomass production, immunity, and pH control, and advance our ability to investigate the plant growth‐defense balance.ISSN:0261-4189ISSN:1460-207