Cell wall (CW) is the foremost interface at which interactions between plants and fungi take place. Pectin, one of the main components of CW, is secreted in a high methylesterified form. Pectin methylesterases (PMEs), catalyse the removal of the methyl esters producing homogalacturonans, releasing methanol (MeOH) and protons. Specific plant PMEs are induced in different plants by necrotrophs but the knowledge of the molecular mechanisms underlying PME-related immunity remain currently limited. PMEs could assist the production of Oligogalacturonides (OGs), elicitors of defence responses. Intriguingly, MeOH, a volatile organic compound released by PME activity, may function as a systemic intra-plant or inter-plant alarm signal. PMEs activity can also reinforce pectin structure, through Ca2+ mediate crosslinks, influencing the penetration of the fungus. PMEs induction could also lead to an apoplast acidification, a potential plant defence response to contrast the host alkalinisation, exploited by Fungi to improve their infectious potential. Available microarray data regarding the expression of Arabidopsis PMEs upon infection by different microbes indicates that AtPME17 is induced by all pathogens analysed and can be considered a potential biomarker for microbial infection. In this project, the contribution of AtPME17 on the PME activity, the release of OGs, MeOH, and changes in apoplastic pH and Ca2+ concentration will be investigated in Arabidopsis during Botrytis cinerea infection. A multidisciplinary approach including reverse genetics, molecular biology, glycan immune-histochemistry, LC-MS/GC-Mass Spectrometry and confocal and electron microscopy will be exploited. The AtPME17 structure and activity and its regulation will be explored by using different biochemical approaches. The project aims to understand if and how AtPME17, and its orthologs in crop species, could represent new promising genetic determinants useful to improve plant resistance to fungal disease.
The project aims to use new analytical tools to identify unexplored targets. The study of both gene expression and cell wall modifications is usually conducted by plant physiologist and pathologist at precise time points during pathogen infection. Monitoring changes in both wall polysaccharides and apoplastic physiological features such as pH and Ca2+ as they occur and at the point of fungal penetration thanks to the new probes available by confocal microscopy gives the project an innovative nature. The possibility of combining the biochemistry of proteins with the plant physiology associated with plant protection is a strength of the project. The possibility of expressing and crystallizing the complete form of PME17 will certainly lead to a strong gain in terms of knowledge on the role of the pro-region in the regulation of PME activity. The use of the laser micro dissector will allow us to define more precisely the location and strategy of the plant defense intervention based on the PME activity. The multidisciplinary work plan in will lead to a better knowledge of the complex plant-fungal pathogen interactions in the model plant Arabidopsis thaliana. Our efforts will provide new experimental tools and platforms to analyse, at molecular level, the plant-pathogen interactions.
Recent researches revealed that post-harvest losses of fruit and vegetable are estimated to be 40-50% (1). Due to post-harvest losses more than 30% of harvested fruits will not reach the consumers plate. Fungal pathogens play a key role in those losses, as they cause most of the vegetable and fruit rots and the customers complaints. Development of resistant cultivars represents a major environmentally friendly solution for both breeders and plant pathologists. The molecular dynamics and underlying genes identified in the project in the Arabidopsis thaliana model plant will represent excellent tools to produce, in a next future, crop varieties with a durable resistance to B. cinerea, either by traditional breeding or by genetic engineering. The results foreseen in this proposal will have an impact well beyond the species and pathogen here studied: most likely, analogous molecular mechanisms control PME activity during infection with other necrotrophic pathogens and in other crop species. The availability of new resistant genotypes can reduce the need of fungicides and minimize their impact on the environment, ensuring a high standard of plant yield and food quality resulting in a long-term benefit for citizens, economy and society. Taken together all the research activity of the project addresses in a most considerable way to food security, sustainable agriculture and bio-economy goals of the tackling social challenge targets.
1. Alkan N, Fortes AM. Insights into molecular and metabolic events associated with fruit response to post-harvest fungal pathogens. Front Plant Sci. 2015