Brassinosteroid Stimulates Hydrogen Peroxide Biosynthesis and Reduces the Effect of Cold Stress

Cold stress is an adverse environmental condition that limits the growth and yield of grapes. Brassinosteroids (BRs) are a class of polyhydroxylated steroidal phytohormones that induce plant resistance to biotic and abiotic stresses. Hydrogen peroxide (H2O2) is a crucial signaling molecule that participates in various physiological processes and abiotic stresses. However, the role of endogenous H2O2 in BR-induced cold stress tolerance of grape seedlings was unclear. Here, we showed that BR application significantly induced cold tolerance of grape seedlings via maintaining higher antioxidases activities, osmoprotectant levels, maximum photosystem II quantum yield, chlorophyll content, and less cell membrane damage, and regulating phytohormone levels. Even more importantly, exogenous BR treatment triggered the upregulation of VvRBOHa, VvRBOHb and VvRBOHe genes expression, the activity of NADPH oxidase, as well as the levels of endogenous H2O2 in the seedlings exposed to cold stress. Pretreatment with dimethylthiourea (DMTU, an H2O2 scavenger) significantly blocked BR-induced H2O2 production under cold stress by reducing NADPH oxidase encoding genes expression and NADPH oxidase activity. Moreover, inhibiting endogenous H2O2 eliminated BR-induced increase in Fv/Fm value and superoxide dismutase, catalase, ascorbate peroxidase, peroxidase activities, and the changes of phytohormone levels. Taken together, we provide evidence supporting the essential roles of RBOH-dependent H2O2 in BR-induced cold tolerance in grape seedlings.