The Stomatal Regulation Mechanisms Of Drought Priming Induced Drought Tolerance In Wheat

Drought is a major environmental factor limiting wheat growth and productivity.More and more studies have shown that drought-priming during the wheat early growth period can effectively alleviate drought tolerance during the late growth period,and it is one of the strategies for wheat production to actively respond to drought stress.Under drought stress,compared with the non-primed plants,the primed plants showed higher stomatal conductance,so as to better adapt to drought stress,but the stomatal regulation mechanism of drought priming is still unclear.Abscisic acid(ABA),hydrogen peroxide(H2O2)and nitric oxide(NO)are important signal substances involved in the stomatal regulation of in plants,but their regulation in regulating the stomatal response to drought priming in wheat is still unclear.In this experiment,Yangmai 16(drought priming sensitive type)and Sumai 188(drought priming retardation type)were used as materials to study the the role and regulatory mechanism of abscisic acid(ABA),hydrogen peroxide(H2O2)and nitric oxide(NO)in drought-priming induced drought tolerance during wheat grain filling stage,then clarify the stomatal response mechanism of these signal substances in drought priming to improve wheat drought tolerance.Two experiments were as follows:(1)pretreatment of exogenous ABA,H2O2 and NO inhibitors and scavengers in wheat four-leaf stage and six-leaf stage,followed by drought priming after 2 days(soil water potential-0.3 Mpa～-0.4 Mpa)3d,severe drought stress treatment(-0.8 Mpa～-1 Mpa)was carried out during the filling stage(7 days after flowering),and rehydration was completed immediately after the treatment for 5 days.(2)The drought priming(10%PEG6000 water potential:-0.58MPa)was carried out in the three-leaf stage and the drought stress(20%PEG6000 water potential:-0.96MPa)was carried out after 10 days of recovery.The stomatal response mechanism of drought priming to improve the drought tolerance of wheat was studied.The main findings are as follows:1.The Role and Physiological Mechanism of Abscisic Acid and Hydrogen Peroxide in Drought-priming Induced Drought Tolerance during Wheat Grain Filling StageDrought stress during grain filling stage caused the significant decrease of 1000-grain weight and yield,water potential dry matter accumulation,photosynthetic rate and stomatal conductance.Compared with the non-primed plants(CD),the primed treatment(PD)increased the levels of ABA and NO in wheat leaves by up-regulating the expression of the 9-cis epoxy carotenoid dioxygenase gene NCED1,aldehyde oxidase gene AO1,AO2 and NAD(P)H oxidase encoding gene RBOH,then significantly up-regulated the expression of the △l-pyrroline-5-carboxylic acid synthase coding gene P5CS and the betaine aldehyde dehydrogenase encoding gene BADH,and down-regulated the expression of proline-degraded pyruvate dehydrogenase-encoding gene PDH,increased the content of osmotic adjustment substances in wheat leaves to enhance the drought tolerance of the plants.Further analysis of the drought priming process,found that drought priming reduced leaf water potential,shoot biomass and photosynthetic capacity of wheat,and up-regulated the expression level of proline synthesis related gene P5CS,betaine-related synthetic BADH and down-regulated proline decomposition related gene PDH,then increased the content of soluble sugar,proline and betaine of the wheat leaves.Drought priming significantly induced the production of ABA and H2O2 in leaves,which is related to up-regulation of the key genes of ABA synthesis,NCED1,AO1,AO2 and NAD(P)H-oxidase synthesis gene RBOH.Inhibition of ABA and H2O2 production in part exacerbated the damage caused by drought priming,reduced the increase of osmotic adjustment substances caused by drought priming and the up-regulation of synthetic related genes P5CS and BADH.Inhibition of ABA significantly reduced the content of ABA and H2O2 in leaves,but inhibited H2O2 had no significant effect on ABA content in leaves.And drought stress after ABA inhibition had no significant priming effect on wheat.In conclusion,ABA and H2O2 play important roles in drought-priming induced drought tolerance.H2O2 is mainly produced by NADPH oxidase on the plasma membrane,and ABA plays a role in the upstream of H2O2 signal,regulating downstream osmotic regulation,stomatal movement and photo synthetic capacity,then the drought tolerance of wheat under drought stress was enhanced.2.The Role and Physiological Mechanism of Nitric Oxide in Drought-priming Induced Drought Tolerance during Wheat Grain Filling StageUnder drought priming,exogenous NO scavenger c-PTIO,NR inhibitor Tungstate and NOS inhibitor L-NAME significantly reduced the NO content in wheat leaves,but did not significantly reduce the ABA and H2O2 content of wheat leaves.Compared with primed treatment(P),inhibition of NO during drought priming decreased the leaf water potential,shoot biomass and photosynthetic capacity,while reduced the increase of soluble sugar,proline and betaine content of wheat leaves caused by drought priming and exacerbated the damage caused by drought priming.Compared with non-priming treatment,the ABA and NO of the primed plants were significantly increased,the content of H2O2 was decreased,the photosynthetic capacity and osmotic adjustment capacity increased significantly,and reduced yield decline caused by direct drought stress.At the same time,the relief effect of drought-priming after NO scavenger,NR and NOS inhibitor treatment was significantly reduced,which affected osmotic adjustment and regulated its drought tolerance.In conclusion,NO plays an important role in drought priming induced drought tolerance during grain filling stage.NO was in the downstream of H2O2 in drought priming induced drought tolerance,and both NR and NOS were involved in NO synthesis in wheat leaves.2.The Stomatal regulation Mechanisms of Drought Priming induced Drought Tolerance in WheatThe dynamic change of stomata after drought priming showed that there was a significant difference in stomatal opening after 0.5 h of drought priming compare with control.Therefore,the response mechanism of stomata to drought priming was analyzed at 0.5 h after drought priming.The main results showed that drought priming significantly reduced the stomatal opening of wheat leaves.Laser confocal scanning microscopy showed that a large amount of H2O2 and NO accumulated in the guard cells after drought priming.Laser Capture Microdissection and gene expression analysis of guard cells showed that the expression level of the guard cell’s stomatal regulation of the fast anion channel encoding gene QUAC1 and the slow anion channel encoding gene SLAC1 was significantly up-regulated.Inhibition of ABA,H2O2 and NO during drought priming significantly slowed the decrease of stomatal opening caused by drought priming,and Inhibition of ABA and H2O2 during drought priming decreased H2O2 content in guard cells and the expression of SLAC1,but the inhibition of NO had no significant effect on H2O2 accumulation.Inhibition of ABA,H2O2 and NO during drought priming significantly decreased NO content in guard cells.Primed treatment(PD)significantly reduced the H2O2 content of guard cells,increased NO content,and significantly up-regulated the expression levels of the anion channel-encoding genes QUAC1 and SLAC1 involved in stomatal regulation,resulting in a significant decrease in stomatal opening.The above results indicated that the stomata partial closure was significantly induced after 0.5h of drought priming,but the stomata closure did not reduce the photosynthetic rate,which could reduce transpiration and maintain plant water potential.This may be an adaptive protection mechanism that does affect the stomatal closure of wheat leaves.