Functions And Mechanisms Of Phosphatidic Acid On Cellular Membrane Stability Under Water Stress

Wheat ranked second as the main crop globally.Drought is one of an important restrict in wheat yield.Deeply understanding mechanisms of plants in response to drought stress provides guidances for breeding selection and improvement.Phospholipase D(PLD)is an important signal transduction enzyme under a variety of stresses.N-butanol(BA)has been used in many studies to stimulate PLD activity,and some researchers applied it to inhibit PLD-mediated transfer from phospholipids to phosphatidic acid(PA).Since PLD is a multi-gene family,and each PLD isoform has substrate preferences and properties for responses to various stresses.PA produces specific compositions of molecular species of lipids under different stresses.In this study,we adopted ESI-MS/MS technique to identify molecular species composition under drought stress,and trace back to probable origins of these PA species.We used BA to increase PA product,and we measured changes of molecular species of PA,as well as the effects of PA on drought resistances of wheat.After that,iTRAQ and LC/MS technique were applied to analyze proteins in various metabolite pathways and explore mechanisms of regulation of PA on drought resistance.The main conclusions are as follows:(1)BA promotes PLD activity in wheat seedlings under drought stressPA(34:3),PA(34:2),PA(36:6),PA(36:5)and PA(36:4)exhibited signal feature,rising firstly and then falling down rapidly.BA treatment promoted PA production at 10 min and 1h under drought stress.PA(34:3),PA(34:2),PA(36:5),PA(36:4)were elevated significantly.Since BA has an activating effect on PLD and inhibits the transacylation of PLD,the results of this experiment indicated that the promotion of PLD is more predominant in the leaves of Jinmai 47 seedlings.(2)PA negatively affects plant drought resistance under drought stressBy comparing the growth rate,membrane lipid peroxidation degree,membrane stability and photosynthetic parameters of PEG+BA treatment and PEG treatment,it was found that the growth of winter wheat seedling leaves was inhibited,net photosynthetic rate was decreased,photosynthetic non-stomatal limitation and cell membrane ion leakage rate increased significantly in PEG+BA treatment,and malonaldehyde(MDA)was increased,indicating that BA reduced cell membrane stability.(3)Function mechanisms of PA under drought stressComparing the proteomes after 4 days of PEG treatment and PEG+BA treatment,the results showed that the content of photosynthesis-related proteins decreased,all transportation-related proteins decreased,proteins involved in lipid metabolism reduced,so as the most proteins function in nucleic acid metabolism in nucleus.However,almost all antioxidant proteins that scavenge reactive oxygen species were up-regulated,57% of drought-resistant or tolerant-related proteins were up-regulated,and 68.4% of carbohydrate metabolism-related proteins were elevated,with the reduction of both starch and sucrose,and the accumulation of glucose.As mentioned above,the product of PLD,PA,mediates a variety of physiological processes under stress,and phospholipase A(PLA)is significantly reduced here.At present,only BA has been found to inhibit the transformation of PLD,but there is no report on whether BA laid an inhibition effect on PLA production.Further researches are needed.(4)Dynamic responses of lipids in wheat leaves to drought stressLipidomic analysis showed that phosphatidylcholine(PC),phosphatidylethanolamine(PE)and phosphatidylglycerol(PG)were the main targets of degradation,and the main origins of PA.PC was preferentially hydrolyzed among them.Monogalactosyldiacylglycerol(MGDG)(34:3),Digalactosyldiacylgly(DGDG)(34:3)began to decline earlier than the time when two major molecular specie,MGDG(36:6)and DGDG(36:6),began to fall.The result suggested that various molecular species might play different roles in distinct subcellular structure.