Cell wall-associated proteins from Phaeomoniella chlamydospora trigger calcium-dependent oxidative burst in grapevine cells

: Esca is a major grapevine trunk disease causing devastating economic losses in vineyards. This complex disease results from trunk colonization by fungi such as Phaeomoniella chlamydospora ( P.ch ). Because the characterization of early molecular events occurring during infection is difficult to perform directly in grapevine trunks, we previously developed a procedure based on stomatal movement to investigate early signaling events during pathogen perception. Assays performed on epidermal peels revealed that cell wall–associated proteins, but not polysaccharides, from P.ch induced stomatal closure in Vitis vinifera . Using the fluorescent probe H 2 DCFDA, we further showed that the perception of these cell wall proteins triggered a rapid accumulation of reactive oxygen species (ROS) in grapevine cells. The addition of diphenyleneiodonium chloride (DPI), an inhibitor of NADPH oxidase–dependent ROS production, prevented the stomatal closure induced by the cell wall proteins. Furthermore, treatment with the calcium (Ca)-chelating agent EGTA blocked both ROS accumulation and stomatal closure triggered by the cell wall proteins, suggesting that the oxidative burst induced by these proteins is dependent on Ca signaling. Altogether, these results provide new insights into the mechanisms underlying grapevine perception and early defense responses against an esca-associated pathogen.