Effects Of Mechanical Wounding On Cold Tolerance In Wheat And Its Mechanisms

The mechanism of physiological responses of wheat plant to cold tolerance induced by mechanical wounding were studied under field experiments and hydroponic culture.Three experiments were conducted:(1)Under field conditions,a field air temperature control system(FATC)was used to simulate cold stress in jointing stage.The effects of mechanical wounding in tillering stage on cold tolerance in jointing stage of wheat plant were investigated;(2)Under hydroponic culture,synthetic inhibitors or scavenger were applied to detect the role of NO and H2O2 in mediating local mechanical wounding induced freezing tolerance in systemic leaves of wheat plant;(3)Under hydroponic culture,the inhibitors or scavenger of H2O2 were used to explore the alleviation effects of mechanical wounding on the newly occurred leaves of wheat plant.The main results of this study were listed below:1.Mechanical wounding induces cold tolerance in jointing stage of wheat plantLocal mechanical wounding before jointing stage enhanced cold tolerance of wheat plant in jointing stage.The Pn and activities of antioxidant enzymes in the first fully expanded leaves were increased significantly while H2O2 content and membrane peroxidation were decreased in wounding pretreated plants under cold conditions.In addition,wounding-induced cold stress mitigation was closely associated with the contents of total soluble sugars and free amino acids.Meanwhile,wounding increased the snapping resistanc of wheat second basal intemode at 5,15 and 25 days after anthesis.The activities of crutial enzymes in plant secondary metabolism of the second basal internode were increased in wounding treatment.In contrast,the gravity center height and internode length were decreased while external diameter and thickness of the second basal internode were increased under mechanical wounding,leading to the higher lodging resistance index.Thus,mechanical wounding could enhance cold tolerance in jointing stage by strengthening the antioxidant system and photosystem in wheat plant.Furthermore,mechanical wounding could also increase the snapping resistance and regulate the secondary metabolism of wheat stem after anthesis.2.Mechanical wounding induces freezing tolerance in untreated systemic leaves of wheat plantLocal mechanical wounding enhanced freezing tolerance in untreated systemic leaves of wheat plant,and meanwhile the signal molecules hydrogen peroxide(H2O2)and nitric oxide(NO)were accumulated systemically.Pharmacological study showed that H2O2 acts upstream of NO in wounding signal transduction pathways.Cytochemical and vascular tissues localizations approved that RBOH-dependent H2O2 acts as long-distance signal in wounding response.Transcriptome analysis revealed that 279 genes were up-regulated in plants treated with wounding and freezing,but not in plants treated with freezing alone.Importantly,freezing-and wounding-induced genes were significantly enriched in the categories of "photosynthesis" and "signaling".Thus,mechanical wounding can induce freezing tolerance in wheat through the systemic accumulation of NO and H2O2,and further modifications in photosystem and antioxidant system.3.Mechanical wounding induces freezing tolerance in newly occurred leaves of wheat plantLocal mechanical wounding enhanced freezing tolerance in the newly occurred systemic leaves of wheat plant.The Pn,Fv/Fm and activities of antioxidant enzymes in the newly occurred leaves were significanty increased after wounding while alleviated freezing-induced chlorophyll decomposition,electrolyte leakage,water lose and membrane peroxidation.In addition,wounding-induced freezing stress mitigation was closely associated with the ratio between reduced and oxidized glutathione(GSH/GSSG),and the ratio between ascorbate and dehydroascorbate(AsA/DHA),as well as the contents of total soluble sugars and free amino acids.Importantly,pharmacological study showed that wounding-induced freezing tolerance was substantially arrested by pretreatment of wheat leaves with the scavenger of hydrogen peroxide(H2O2)or the inhibitor of NADPH oxidase(RBOH).Thus,local mechanical wounding-induced freezing tolerance in newly occurred leaves is largely attributed to RBOH-dependent H2O2 production,and through strengthening the antioxidant system and photosystem,modifications of AsA-GSH cycle,adjustion of osmolytes such as soluble sugar and free amino acids in wheat plant.In conclusion,we demonstrated that mechanical wounding could alleviate cold stress in jointing stage as well as enhance freezing tolerance of the upper unwounded leaves and the newly occurred leaves of wheat plant.Importantly,physiological and transcriptome analysis revealed that endogenous NO and H2O2 signal transduction,photosystem,antioxidant system as well as the AsA-GSH cycle play vital roles in wounding-induced cold tolerance of wheat plant.