The Molecular Physiological Mechanisms Of Melatonin Regulating Tomato Adaptation To Saline-alkali And Drought Stress

Saline-alkali and drought stress are serious environmental factors that affecting crop distribution and growth.Tomato?Solanum lycopersicum L.?is widely planted all over the world,it is susceptible to saline-alkali and drought stress.So,studying the mechanism of salinity-alkalinity stress tolerance of tomato for the sustainable production of tomato in saline-alkali land and arid area has important theory and application value.Melatonin is a small molecular reductive substance in plants,which is involved in plant growth,development and resistance regulation.We constructed tomato plants with over-expression of caffeic acid-O-methyltransferase?SlCOMT1?,which was proved as an important enzyme controlling melatonin synthesis in tomato,and its overexpression could significantly improve salt and drought resistance of tomato.Based on this,we studied the physiological and molecular mechanisms of salt and drought resistance regulated by SlCOMT1.In addition,we also studied the regulation of exogenous melatonin in S-nitrosation of tomato protein under saline-alkali stress.The main results are as follows:1.Overexpression of SlCOMT1 increased tomato tolerance to salt stressOverexpression of SlCOMT1 signigicantly increased melatonin accumulation and salt tolerance in tomato plants,and increased cholrophyll content,enhanced photochemical electron transfer efficiency and dark reaction rate under salt stress,and promoted the growth of tomato plants under salt stress.In addition,overexpression of SlCOMT1 increased the mineral nutrition level by promoting the uptake of N,P and K under salt stress.Overexpression of SlCOMT1 could activate the expression of SOS pathway key genes including SOS1,NHX1,NHX2,HKT1.1 and HKT1.2.Firstly,SOS1 was activated to promote Na⁺efflux from root,by upregulating the HKT1.1 and HKT1.2 expression to inhibit Na⁺transport from root to stem and leaf tissues,and then NHX1 and NHX2 were used to enhance intracellular Na⁺regional detoxification,thereby reducing the accumulation of Na⁺in various tissues of tomato plants,increasing K⁺uptake,maintaining higher K⁺/Na⁺in tissues and cells under salt stress.Therefor,the toxic effect of Na⁺on cell was alleviated.Salt could cause ROS?O₂.^-and H₂O₂?burst in tomato plants,while overexpression of SlCOMT1 enhanced antioxodant enzyme activity?SOD,POD,CAT,GR and DHAR?and antioxidants?AsA and GSH?contents,reducing the ROS accumulation and membrane lipid peroxidation in tomato plants induced by salt stress.Furthermore,overexpression of SlCOMT1 also activated the expression of plant stress response genes?AREB,AIM1,MAPK1,WRKY33,DREB and CDPK1?,which might be an important mechanism that Overexpression of SlCOMT1improved salt tolerance in tomato plants.2.Overexpression of SlCOMT1 increased tomato tolerance to drought stressOverexpression of SlCOMT1 inhibited the degradation of chlorophyll,promoted photosynthesis,increased root,plant height,stem diameter and plant weight in tomato under drought stress.Further studies showed that overexpression of SlCOMT1 activited the expression of ABA synthesisi-related genes?ZEP,NCED3 and ABA3?and increased the expression of ABA receptor genes?PYL1,PYL2,PYL4,PYL8,PYL9 and PYL10?,which could induce ABA signal transduction,controll stomatal closure,and reduce transpiration rate and water dissipation,thus maintaining high water content of tomato plants under drought stress.Overexpression of SlCOMT1 eliminated ROS accumulation and reduced membrane lipid peroxidation injury by increasing antioxidant enzyme activity?SOD,POD,CAT,GR and DHAR?and antioxidant?AsA and GSH?content in tomato plants under drought stress.Moreover,overexpression of SlCOMT1 also activitated the expression of stress response factors?DREB1,AREB1,ERF5,WRKY33,MYB and LEA?,which played an important role in improving drought tolerance of tomato plants.3.Regulation of exogenous melatonin on protein S-nitrosation modification in tomato plantunder NaHCO₃ stressNaHCO₃ stress significantly induced nitrosative damage and led to an increase of protein nitrosation modification in tomato plants.Melatonin could react with intracellular reactive nitrogen?RNS?to remove nitrosative damage induced by stress and reduced the level of nitrosation modification of protein.Proteomic analysis of S-nitrosylated proteins based on Iodo TMT indicated that the S-nitrosylated proteins induced by NaHCO₃ stress were mainly distributed in subcellular organelles with high energy metabolism,such as chloroplasts,cytoplasm,mitochondria,and so on.Melatonin could effectively promote the denitrification of protein in chloroplasts and cytoplasm under NaHCO₃ stress.Through the combined analysis of GO,Pathway and key physiological indicators,the following results were found:melatonin reduced the nitrosation modification of photosynthetic pathway proteins and increased chlorophyll content,maximum photosynthetic efficiency?Fv/Fm?,antenna conversion efficiency?Fv'/Fm'?,photochemical efficiency??PS??,photochemical quenching coefficient?qP?and electron transport rate?ETR?,reduced the non-photochemical quenching coefficient?NPQ?,therefore,the absorption and utilization of light by tomato leaves under NaHCO₃ stress were improved,and photosynthesis was finally enhanced;Under NaHCO₃ stress,melatonin significantly reduced the S-nitrosation of amide biosynthetic proteins,and inceased the activities of glutamine synthase?GS?,glutamate dehydrogenase?GDH?and glutamate synthase?GOGAT?during nitrogen metabolism,thus improving the efficiency of nitrogen assimilation;melatonin reduced the nitrosation modification of fatty acid pathway proteins under NaHCO₃ stress,increased the contents of unsaturated fatty acids?oleic acid,linoleic acid,gamma-linolenic acid,alpha-linolenic acid,cis-11,14-eicosatrienoic acid,cis-11,14,17-eicosatrienoic acid and Erucic acid?,decreased the contents of saturated fatty acids?lauric acid,myristic acid and Erucic acid?,the ratio of unsaturated fatty acid to saturated fatty acid was increased,so the stability and fluidity of membrane under NaHCO₃stress were enhanced.