He Mechanism Of Ascorbic Acid Regulating The Adaptation Of Tomato Seedlings To Salt Stress

[Objective]Soil salinization is one of the main obstacles restricting the growth and development of processing tomato in Xinjiang.Salt stress causes ion poisoning,osmotic stress and oxidative stress,destroys the photosynthetic mechanism,leads to the decline of photosynthesis and resistance of processing tomato in Xinjiang,and the problems of crop yield and quality become increasingly prominent.Therefore,it is of great scientific and practical significance to study the adaptive mechanism of tomato salt.Ascorbic acid（As A）,as an electron donor and antioxidant in redox reactions,plays an important role in plant growth and development and stress resistance.Previous studies have shown that an appropriate concentration of As A can improve the salt tolerance of tomato seedlings and promote their growth,but the specific mechanism is still unclear.In addition,studies have shown that nitric oxide（NO）,as an important signaling molecule,is often accompanied by an increase in endogenous As A levels while improving plant resistance.In the non-enzymatic pathway of NO production,As A can react with NO₂^-to produce NO.It shows that there is a certain relationship between NO and As A.Therefore,this study,on the one hand,analyzed the mechanism of As A improving tomato salt adaptability,and on the other hand,explored the relationship between As A and NO in the process of improving salt adaptability.[Methods]To investigate the mechanism of salt adaptation of tomato under salt stress by As A,this experiment used processing tomato（Solanum lycopersicum L.）cv.‘Ligeer 87-5’as the experimental material,which was grown in nutrient solution and treated with 100 m M Na Cl to simulate salt stress,and foliar sprays of 0.5 m M As A（NA treatment）,0.25 m M LYC（lycorine,an inhibitor of As A synthesis）（NL treatment）and LYC+As A（NLA treatment）to construct different endogenous As A levels under 100 m M Na Cl simulated salt stress treatment,and to investigate the effects of different treatments on ion homeostasis,osmoregulation,redox（redox）status and photosynthetic fluorescence parameters.Transcriptomic,targeted metabolomic and protein ubiquitinomic analyses were also carried out on tomato leaves under salt stress and NA treatment.To resolve the relationship between As A and NO in the regulation of salt acclimation in tomato,the As A synthesis inhibitor LYC（0.25 m M）and NO scavenger PTIO（1-lambda1-Oxidanyl-4,4,5,5-tetramethyl-3-oxido-2-phenylimidazol-3-ium,0.05 m M）were used.,the roles of As A and NO in regulating ion homeostasis,osmoregulation,redox status,photosynthetic fluorescence parameters and their interrelationships were investigated by foliar spraying of As A,SNP（sodium nitroprusside,NO donor）,LYC,PTIO,PTIO+As A and LYC+SNP setting different treatments under salt stress.[Results]（1）Exogenous As A significantly increased the activity and gene expression of L-galactosyl-1,4-lactone dehydrogenase（Gal LDH）and L-galactose dehydrogenase（Gal DH）in tomato seedling leaves under Na Cl stress and NL treatment,and increased the content of endogenous As A;Exogenous As A regulates multiple ion transporters through SOS pathway to rebuild the intracellular ion homeostasis,improve the selective transport ability of K⁺,Ca²⁺and Mg²⁺in the shoot,promote the efflux and regionalization of Na⁺and Cl^-,and alleviate the ion toxicity caused by salt stress.By enhancing the activity and gene expression of proline dehydrogenase（Pro DH）and inhibiting the activity and transcriptional level of△1-pyrroline-5-carboxylate synthase（P5CS）and ornithine-δ-aminotransferase（OAT）,the excessive Pro content in leaves can be reduced to alleviate osmotic stress.By improving the activities of antioxidant enzymes and key enzymes of As A-GSH cycle and gene expression,effectively eliminating reactive oxygen species（ROS）,reducing membrane lipid peroxidation and alleviating oxidative stress.In addition,transcriptome analysis showed that As A was involved in regulating hormone signaling through the regulation of transcription factors and gene expression,decreasing abscisic acid（ABA）and 1-aminocyclopropane-1-carboxylic acid（ACC）content and increasing jasmonic acid（JA）,salicylic acid（SA）,cytokinins（CKs）,gibberellins（GA）and indoleacetic acid（IAA）content.（2）Exogenous As A could significantly increase the photosynthetic pigment content（Chl and Car）in tomato seedling leaves under Na Cl stress and NL treatment to improve the light energy absorption and transformation,and increase the leaf gas exchange efficiency under salt stress by increasing stomatal opening and reducing stomatal density,thus increasing the net photosynthetic rate（P_n）of tomato seedling leaves.As A treatment alleviates the decrease of maximum photochemical efficiency（F_v/F_m）of PSII,the maximal P700 changes（P_m）,non-photochemical quenching coefficient（NPQ）,effective quantum yield of PSII and PSI[Y（II）and Y（I）]under salt stress.By restoring the excitation energy balance between the two photosystems.A high level of electron flux for the photosynthetic carbon reduction[Je（PCR）]was maintained,and cyclic electron flow（CEF）around PSI was activated to alleviate the inhibition of PSI and PSII in tomato seedlings under salt stress.In addition,As A increased the electron transfer activity（φE_o andψ_o）on the acceptor side of PSII,increased the size of the PQ pool（S_m）,and decreased the maximum rate at which Q_A was reduced（M_o）.At the same time,As A counteracted the negative effects of salt stress（with or without LYC spray）on the activity density of PSII per reaction centers（RCs）,the utilization and dissipation of absorbed light energy.（3）A total of 489 ubiquitinated proteins,843 ubiquitinated peptides and 972 lysine ubiquitination sites were identified in Control,Na Cl-treated and NA-treated tomato leaves using ubiquitinated Label-free quantitative proteomics techniques,of which 848 lysine ubiquitination sites were quantified for 446 ubiquitinated proteins.The comparative group analysis of NA＿vs＿Na Cl showed that As A up-regulated 97 differentially ubiquitinated peptides and down-regulated 98 differentially ubiquitinated peptides under Na Cl stress.These differentially ubiquitinated peptides correspond to proteins involved in multiple metabolic pathways such as ascorbate and aldehyde metabolism,photosynthetic biocarbon sequestration,glutathione metabolism,and photosynthesis.The analysis revealed that exogenous application of As A significantly down-regulated the ubiquitination levels of proteins related to ascorbic acid and glutathione metabolism,in addition to being involved in regulating the ubiquitination levels of proteins related to photosynthesis and biological carbon sequestration.（4）Under salt stress,exogenous As A can significantly increase the level of endogenous NO,and NO can also significantly increase the level of endogenous As A,indicating the mutual response between As A and NO.Spraying As A and SNP alone can maintain intracellular ion homeostasis through SOS pathway;Regulate Pro anabolism to alleviate osmotic stress;Improve the activity of antioxidant enzymes,remove ROS,and reduce oxidative stress;Improve thermal dissipation,maintain excitation energy distribution balance,activate CEF photoprotection mechanism,and effectively alleviate photoinhibition.But exogenous LYC and PTIO increased the damage of salt stress on plants.And PTIO had less effect on the protective effect induced by As A,while LYC could reduce the protective function of SNP.[Conclusion]Exogenous As A improves salt acclimation of tomato seedlings by mediating endogenous As A synthesis and regulating the SOS pathway,Pro anabolism,ROS scavenging system,phytohormone signaling,and photosynthesis;As A regulates the level of protein ubiquitination modification in tomato seedlings under salt stress and protects proteins from degradation under salt stress,thus improving salt acclimation of tomato seedlings;NO as a downstream signal involved in the regulation of ion toxicity,osmoregulation of oxidative stress and photosynthetic damage of tomato seedlings by As A under salt stress.