| Soil salinization is a global resources and ecological problem,limiting agricultural productivity worldwide.Sodium chloride is the most widespread soil salt.Almost all crop plants are non-halophytes that they can't tolerate high salt concentrations.High concentration of NaCl in soil will disrupt the ion homeostasis in plants,lead to water deficiency,and result in the inhibition of plants growth and development.In China,Cucumber(Cucumis sativus L.)is an important vegetable crop with the largest cultivation area,grown both in open fields and protected facilities.Cucumber is highly sensitive to soil salinity,showing inhibited plant growth and decreased yield.Polyamines(PAs)are low-molecular-weight aliphatic nitrogenous compounds.At physiological pH,PAs are usually protonated and positively charged,showing strong biological activities.Studies show that PAs are widely involved in many metabolic processes,and also regulates the response of plants to biotic and abiotic stresses.The application of exogenous PAs is an effective way to enhance plants' stress tolerance.Studies showed that putrescine(Put)could improve the photochemical efficiency,reduce the accumulation of Na+ and Cl-,and enhance the antioxidant defense ability,thus improving plant's salt tolerance.However,these studies mainly focused on the physiological level.The specific mechanism of Put improving salt tolerance needs to be further studied.In the present study,we adopted hydroponics culture,and used a salt-sensitive cucumber variety 'Jinyou No.4'as material,carrying out experiments to explore the underlying mechanism of exogenous Put(either sprayed to leaves with 8 mM or added to nutrient solution with 0.8 mM)alleviating salt(75 mM NaCl)induced damage.The main results were as follows:1.NaCl stress severely destroyed the leaf internal structure,inhibited photosynthetic capacity,caused the over-accumulation of starch and sucrose and resulted in the decreased biomass accumulation of cucumber seedlings.The application of Put to leaves of NaCl-stressed cucumber seedlings promoted leaf expansion and optimized leaf structure.Exogenous Put significantly increased the maximum rate of Rubisco-mediated carboxylation,light saturated rate of electron transport and triose-phosphate export rate from chloroplast,reduced CO2 compensation point,indicating an improvement of CO2 assimilation.Moreover,Put regulated the activities of sucrose-and starch-metabolizing enzymes,thus reduced the over-accumulation of starch and sucrose in cucumber leaves.As a result,Put improved the overall photosynthtic capacity of cucumber leaves and promoted the biomass accumulation.2.NaCl stress seriously decreased cucumber root growth and leaded to root cells death.Put added to nutrient solution increased the average root growth rate by 80%and improved root activity.62 differentially expressed protein spots in cucumber roots respond to NaCl and Put were successfully identified by MALDI-TOF/TOF MS.These proteins implicated in various biological processes including defense response,protein metabolism,carbohydrate metabolism and amino acid metabolism.Exogenous Put up-regulated most identified proteins involved in carbohydrate metabolism,implying an enhancement in energy generation.Proteins involved in defense response and protein metabolism were differently regulated by Put,which indicated the roles of Put in stress resistance and proteome rearrangement.Put also increased the abundance of proteins involved in amino acid metabolism.Meanwhile,physiological analysis showed that Put could further up-regulated the levels of free amino acids in salt stressed-roots.In addition,Put also regulated the transcription levels of key enzymes involved in polyamine metabolism,and improved endogenous polyamines contents.Therefore,exogenous Put comprehensively regulated carbon and nitrogen metabolism and defense reaction to promote root growth under NaCl stress conditions.3.NaCl stress significantly increased the Na+ levels in cucumber seedlings.Na+level in cucumber root gradually decreased from the outermost epidermis to the inside stele,While Na+ in stem was mainly accumulated in vascular tissue.The scan of leaf transverse sections showed that leaf vein gathered more Na+ than mesophyll cells.Na+ content in xylem sap was also elevated.NaCl stress decreased K+ levels in roots,stems and leaves of cucumber seedlings,but increased K+ content in xylem sap.Exogenous Put inhibited the NaCl-induced accumulation of Na+ in all tissues of root and leaf,but induced more Na+ gathered in the cortex and primary phloem of stem.Put reduced Na+ content in xylem sap of NaCl-stressed seedlings.For all sections scanned,the application of Put effectively alleviated the loss of K+caused by NaCl,and Put further improved the increase of K+ in xylem sap of NaCl-stressed seedlings.Analysis of the expression levels of genes encoding ion transporters showed that Put generally up-regulated SOS1 and NHX expression levels both in roots and leaves of cucumber seedlings.Put also increased the transcription level of HKT in cucumber leaves,but decreased its expression in roots.All these results indicated that exogenous Put could regulate Na+ transport and accumulation,and thus maintaining the Na+/K+ homeostasis in NaCl-stressed cucumber seedlings.4.Under NaCl stress conditions,exogenous Put significantly increased the accumulation of H2O2 in cucumber roots,mainly through improving DAO activity.The application of DMTU,a specific H2O2 scavenger,increased Na+ accumulation and decreased K+ levels in roots and leaves of cucumber.In addition,the application of AG,an inhibitor of DAO,could compromise the ability of Put in reducing Na+ accumulation and preventing K+ loss.These results suggested that the accumulation of H2O2 generated by polyamines oxidation was an important reason to the Na+ and K+ levels regulated by Put.5.NaCl stress significantly reduced the growth of cucumber plants at fruiting stage.The net photosynthetic rate(Pn),transpiration rate(Tr),stomatal conductance(GS)and intercellular CO2 concentration(Ci)were decreased.NaCl stress resulted in the cucumber fruit deformity,and decreased single fruit fresh weight.Tannin,titratable acid,free amino acid and soluble sugar content in cucumber fruit were increased,but vitamin C and soluble protein content were decreased.Exogenous Put increased the Pn and Tr,thus enhanced the photosynthetic capacity.Put also increased the maximal quantum yield of photosystem II photochemistry,the actual photochemical efficiency of photosystem II and the regulated energy dissipation,and decreased the non-regulated heat dissipation,improving the photochemical activity of photosynthetic reaction center.Moreover,exogenous Put could effectively relief fruit deformity,increase fruit weight and water content.Put decreased tannin and titratable acid contents,and increased soluble protein and vitamin C levels,indicating the improvement of fruit quality and flavor. |
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