Regulation Of NO On Physiological Characteristics And Gene Expression In Rice Under Alkali Stress

Rice is the main food crop in Ningxia Yellow River diversion irrigation area.Salinization is common in the soil of the Yellow River diversion irrigation area.Rice is subjected to saline-alkali stress in the process of growth and development,which affects the yield and quality.It is of great practical significance to alleviate the damage caused by saline-alkali stress and improve the alkali tolerance of rice.Nitric oxide(nitricoxide,NO)is an important signal molecule in plants,which plays an important role in regulating plant response to abiotic stress.In this study,rice Zhonghua 11(ZH11,alkali sensitive)and Ningjing 52(NJ52,alkali tolerant)were used as experimental materials to study the effects of NO donor sodium nitroprusside(SNP)on seed germination and seedling growth of rice under alkali stress,so as to provide a theoretical basis for the application of NO as a growth regulator in rice production in saline-alkali soil.The main results are as follows:1.Low concentration of SNP(10～50 μmol·L-1)increased the germination rate of ZH11 and NJ52 seeds under alkali stress,and promoted the growth of buds and roots.However,high concentration of SNP(100 μmol·L-1)inhibited the seed germination,bud and root growth of the two kinds of rice under alkali stress.Among them,SNP 30 μmol·L-1 could significantly increase the GA content of rice seeds under alkali stress,reduce the ABA content,promote seed germination,and effectively alleviate the damage to buds caused by alkali stress.The mitigation effect on seed germination and bud growth of ZH11 was stronger than that of NJ52.2.Compared with alkali stress alone,alkali stress+30 μmol·L-1SNP treatment significantly enhanced amylase activity and up-regulated amylase synthesis gene expression in germinated seeds of ZH11 and NJ52.Among them,β-amylase activity and amylase synthesis gene expression(OsAmy3D,OsBMY2 and OsBMY6)in NJ52 were significantly higher than those in ZH11.3.Compared with alkali stress alone,alkali stress+30 umol·L-1SNP treatment decreased Na+/K+ratio,fructose,sucrose,reactive oxygen species(H2O2 and O2-·),MDA content and LOX activity in ZH11 and NJ52 buds under alkali stress,significantly increased glucose,proline malic acid,citric acid and antioxidant AsA content,AsA/GSH and GSH/GSSG ratio,antioxidant enzyme(SOD,POD,APX)activity,and changed sugar synthase gene expression.The expression of proline synthesis gene OsP5CS and antioxidant enzyme gene was up-regulated,and the expression of lipoxygenase gene OsLOX5 was down-regulated.Among them,the contents of proline,malic acid,citric acid and antioxidant substances,activities of antioxidant enzymes(CAT and DHAR),expression of carbohydrate synthase gene(OsNIN1 and OsSPS11),proline synthase gene OsP5CS and antioxidant enzyme gene(OSCATC,OsDHAR1 and OsGPX4)in NJ52 buds under alkali stress+30 μmol·L-1SNP were significantly higher than those in ZH11.4.Compared with alkali stress alone,alkali stress+30 μmol·L-1SNP treatment significantly increased the activities of nitrate reductase(NR)and S-nitrosoglutathione reductase(GSNOR)in ZH11 and NJ52 buds under alkali stress,significantly increased NO content,and up-regulated the expression of NO synthesis related genes OsNR1.2,OsNR2,OsNOA1 and S-nitrosoglutathione reductase gene OsGSNOR1.Among them,NO content,GSNOR activity,OsNR1.2 and OsNOA1 expression in NJ52 were significantly higher than those in ZH11.5.OPLS-DA combined with variable projection importance(VIP)analysis showed that relative bud length,relative root length,germination rate,β-amylase activity,soluble protein content,NO content,GSH/GSSG ratio,OsNR1.2 and OsAmy3E expression were important differences in distinguishing the effects of SNP treatment on seed germination of ZH11 and NJ52 under alkali stress.Among them,alkali stress+30 μmol·L-1SNP treatment effectively alleviated the damage of alkali stress to NJ52 by significantly affecting the soluble protein content and OsAmy3E expression of NJ52 during germination,and alleviated the damage of ZH11 caused by alkali stress by significantly affecting the relative bud length,relative root length,germination rate,β-amylase activity,NO content,GSH/GSSG ratio and OsNR1.2 expression of ZH11.6.Compared with alkali stress alone,alkali stress+50 umol·L-1SNP treatment increased green leaf coefficient,survival rate,seedling height,root length,shoot dry and fresh weight,root dry and fresh weight and root activity of ZH11 and NJ52 seedlings under alkali stress,and decreased the relative electrical conductivity of leaves.Among them,the green leaf coefficient and root activity of NJ52 were significantly higher than those of ZH11 under alkali stress+50 μmol·L-1SNP treatment.7.Compared with alkali stress alone,alkali stress+50 umol·L-1SNP treatment significantly decreased the ratio of Na+/K+,fructose,sucrose,active oxygen(H2O2 and O2-·),MDA,abscisic acid content and LOX activity in ZH11 and NJ52 seedlings under alkali stress,and significantly increased the contents of chlorophyll a,chlorophyll b,total chlorophyll,glucose,proline,malic acid,citric acid,gibberellin and antioxidants(AsA and GSH),and antioxidant enzymes(POD,CAT and GPX)activities.The expression of carbohydrate synthase gene and chlorophyll synthesis related gene was changed,the expression of proline synthesis gene OsP5CS,antioxidant enzyme gene(OsCu/Zn-SOD,OsPOXl,OsAPX2 and OsDHAR1)and gibberellin synthesis related gene(OsGA3ox2 and OsGA20ox1)was significantly up-regulated,and the expression of lipoxygenase gene OsLOX5 was significantly downregulated.Among them,the contents of chlorophyll a,chlorophyll b,total chlorophyll,sucrose,glucose,proline,malic acid,citric acid,gibberellin and antioxidants,antioxidant enzyme activities(SOD,POD,DHAR,GPX and GR),chlorophyll synthesis gene expression(OsYGL1 and OsCAO1),sugar synthase gene expression(OsNIN1,OsSUS5,OsSUS7 and OsSSPS1)in NJ52 seedlings under alkali stress+50μmol·L-1SNP,The expression levels of antioxidant enzyme synthesis genes(OsCu/Zn-SOD,OsPOX1,OsAPX2,OsDHAR1,OsGPX4 and OsGR3),gibberellin synthesis genes OsGA3ox2,OsGA20ox1 and abscisic acid decomposition gene OsABA8ox1 were significantly higher than those of ZH11.8.Compared with alkali stress alone,alkali stress+50 umol·L-1SNP treatment increased the contents of NO,NO3-and SNO in ZH11 and NJ52 seedlings under alkali stress,enhanced NR activity,and significantly up-regulated the expression levels of OsNR1.2 and OsNR2.Among them,NO3-content,NR activity,OsNR1.2,OsNR2,OsNOA1 and OsGSNOR1 expression in NJ52 were significantly higher than those in ZH11.9.OPLS-DA combined with variable projection importance(VIP)analysis showed that relative root length,relative root dry weight,fructose content,DHA content,GSSG content,OsSUS5 expression,OsNR2 expression and OsPOXI expression were important difference indexes to distinguish the effects of SNP treatment on ZH11 and NJ52 seedlings under alkali stress.Among them,alkali stress+50 umol·L-1SNP treatment alleviated the damage to NJ52 caused by alkali stress by significantly affecting relative root length,relative root dry weight,fructose content,DHA content,OsSUS5 expression,OsNR2 expression and OsPOX1 expression in NJ52 varieties,while exogenous SNP treatment alleviated the damage caused by alkali stress to ZH11 by significantly affecting GSSG content in ZH11 varieties.