Guard cells and stomatal movement reveal early molecular interaction between grapevine cells and esca-associated pathogens

Abstract

Esca is one of the major grapevine trunk diseases that cause vineyards decline and important economic losses in vineyards. Studies have shown that wood differently recognizes Phaeomoniella chlamydospora (P.ch) and Phaeoacremonium minimum (P.min), two esca associated pathogens, at tissular, metabolomic and transcriptional levels. However, the characterization of molecular events occurring throughout the first hours of interaction between grapevine cells and esca-associated pathogens is difficult to perform directly in the trunk. To overcome this difficulty, we proposed a cellular model based on stomata. Stomata are specialized structures consisting of two guard cells surrounding a pore, that perceive stimuli and respond by inducing their opening/closing, which can be quantified. The aim of this study is to decipher the early interaction between esca-associated pathogens and the grapevine cells by using guard-cells model.  Epidermal peels of V.vinifera and A.thaliana are observed and the ostiole aperture is measured before and after immersion in a bathing solution containing either complete cell wall of P.ch or P.min, their parietal proteins or their parietal polysaccharides. Our first results suggest that P.ch’s parietal proteins are recognized by grapevine and A.thaliana cells, whereas it is the polysaccharide part of P.min’s cell wall is recognized. In addition, the use of 1-methylcyclopropene, a competitive inhibitor of ethylene, blocked the stomatal closure in response to P.ch, suggesting an involvement of the ethylene cell signaling in grapevine response to this pathogen.  We develop further this topic by the use of mutants involved in the ethylene signaling. Reactive oxygen species (ROS) production and calcium accumulation are signatures of molecular plant response to pathogens. By using fluorescent dyes, we are determining the kinetics of ROS and calcium accumulation on guard cell protoplasts inoculated with pathogens.  Overall results showed that guard cell and stomatal responses are efficient and promising tools to establish a cell pathosystem model for characterize and explore molecular events that occur during grapevine and esca-associated pathogens. In addition to these results, other cellular contexts will be explored the most complete vision of these interactions.