The Mechanism Of Synergistic Regulation Of 5-aminolevulinic Acid And Calcium Ion On Salt Tolerance In Peanut

Peanut(Arachis hypogaea L.),which is an oil crop,occupies an important position in the oilseed market in China and in the world.Because of its rich in lipids and proteins,it has great nutritional and economic value.However,the increasing global soil salinization and the abuse of pesticides and fertilizers have made peanuts available for use in growing areas and seriously affect the quality of peanuts.In order to improve the salt tolerance of peanut plants and reduce the adverse effects of salt stress on peanut crops,it is necessary to find and dig salt-tolerant molecular mechanism of peanut.Under the premise of not competing with grain,expand the use of peanuts on saline-alkali land.5-Aminolevulinic acid(ALA)is the precursor of chlorophyll synthesis.A large number of studies have shown that ALA can be used as a growth regulator to improve the stress resistance of plants.This experiment was conducted to study the effects of exogenous ALA on peanut salt stress and the effects of Ca2+ on salt tolerance of peanut induced by ALA.The gene of AhHEMA1,a key enzyme gene for ALA synthesis in peanut,was transferred into tobacco by genetic engineering.The salt resistance of transgenic tobacco was preliminarily studied.Firstly,six concentration gradients of 0,1,10,25,50 and 100 mg·L-1 were set up to study the effects of different concentrations of ALA pretreatment on the salt tolerance of peanut seedlings huayu22 under salt stress.The results showed that low concentrations of exogenous ALA pretreatment could increase the accumulation of chlorophyll by up-regulating the expression levels of HEMA1,CHLH and CHLD genes in chlorophyll biosynthesis.ALA could significantly reduce the accumulation of H2O2 and O2-,increase the activities of antioxidant enzymes such as SOD,POD,CAT and APX,reduce the accumulation of malondialdehyde(MDA)and proline,and protect the peanut seedlings.ALA also up-regulated the CaM gene and CaM content under salt stress.Therefore,ALA may enhance salt tolerance of peanut by regulating this signaling pathway.Secondly,we studied the effect of Ca2+ on chlorophyll under salt stress.The results showed that chlorophyll a,chlorophyll b and chlorophyll a/b were increased under salt stress.On the one hand,under the salt stress conditions,exogenous application of a certain concentration of calcium ions significantly enhanced the activity of antioxidant enzymes such as SOD,POD,CAT,and APX,and delayed the degradation of chlorophyll in leaves.On the other hand,the expressions of CHLD and CHLH genes were up regulated after Ca2+ treatment,the expression of CHLI gene was down-regulated and the chlorophyll biosynthesis was increased.This indicated that Ca2+ could regulate the expression of magnesium chelatase components and promote chlorophyll synthesis.Exogenous calcium had no effect on HEMA1 gene expression,and calcium and its inhibitors EGTA,CPZ and LaCl3 had no effect on ALA accumulation in leaves under salt stress.This indicated that Ca2+ might be involved in the regulation of chlorophyll mainly in the downstream of ALA.Exogenous ALA and calcium ion enhanced the salt tolerance of peanut,while calcium inhibitors LaCl3,CPZ and EGTA inhibited the alleviation effect of ALA on salt stress,indicating that calcium ion was involved in the salt tolerance of peanut induced by ALA.Further analysis showed that ALA and calcium ion enhanced Ca2+and CaM content and enhanced plasma membrane Ca2+-ATPase and CDPK activity in peanut leaves.Exogenous calcium ion treatment and calcium-related inhibitors prevented this promotion and did not increase ALA production.This suggests that calcium ions may regulate peanut salt tolerance downstream of ALA.The AhHEMAl gene was studied for its salt tolerance by cloning AhHEMAl gene from peanut and transferring it into wild type tobacco.The results showed that chlorophyll content in AhHEMA1 transgenic tobacco was significantly higher than that in wild type.After 24h of salt stress,the activities of SOD,POD,CAT and APX in AhHEMA1 transgenic tobacco were significantly higher than those in wild type,indicating that AhHEMA1 enhanced the salt tolerance of tobacco.