Effect Of Exogenous Calcium On Nitrogen Use And Quality Of Rice Under Acid Rain Stress

The emission of acid gas cannot be effectively controlled in the short term,resulting in the harm caused by acid rain to agriculture and forestry still continuing.In order to alleviate the threat of acid rain pollution to grain production and food safety,it is very important to find effective methods based on clarifying the impact of acid rain on crop yield and quality.In this paper,food crops rice（resistant rice“Wuyou 308”and sensitive rice“Nanjing 9108”）were taken as research materials,and the purpose was to clarify the regulation mechanism of exogenous Ca²⁺on rice yield and quality under acid rain stress.The effect of exogenous Ca²⁺（5 m M）on yield and nutritional quality（amino acid and protein content）of the two rice cultivars under simulated acid rain（SAR,pH 4.5/3.0）stress was studied with an amino acid analyzer.The effect of exogenous Ca²⁺on nitrogen uptake capacity and transport efficiency of roots in the two rice cultivars under SAR was analyzed by real-time quantitative PCR and Western Blot.Clarifying the regulation mechanism of exogenous Ca²⁺and comparing different resistant cultivars can not only enrich the mechanism of plant acid resistance,but also provide new thinking directions for finding effective ways to alleviate the decline of crop yield and nutritional quality caused by acid rain,and reduce the pressure of agricultural production.The major results were as follows:（1）Effect of exogenous Ca²⁺on the yield and nutritional quality of different resistant rice under SAR was clarified.The results showed that low-intensity SAR（pH 4.5）had no effect on the yield（100-grain weight,grains per ear,and seed setting rate）of resistant rice“Wuyou 308”,but decreased the yield of sensitive rice“Nanjing 9108”.At the same time,low-intensity SAR（pH 4.5）caused a decrease in the total amino acid and essential amino acid contents of the two rice cultivars.High-intensity SAR（pH 3.0）reduced the yield and nutritional quality（total amino acid content,essential amino acid,and protein content）of the two rice cultivars,and the decline was greater than that under low-intensity SAR（pH 4.5）treatment,and sensitive rice“Nanjing 9108”decreased more.Combined with nitrogen content changes at different growth stages of the two rice cultivars under SAR（pH 4.5/3.0）treatment,it was speculated that the larger drop in yield in sensitive rice“Nanjing 9108”was related to lower nitrogen content at the seedling and booting stages.Single exogenous Ca²⁺treatment had no effect on the yield and nutritional quality of the two rice cultivars.Under the combined treatment of Ca²⁺and SAR（pH 4.5/3.0）,the yield and nutritional quality of the two rice cultivars were higher than those under SAR（pH 4.5/3.0）treatment.Among them,under the combined treatment of Ca²⁺and SAR1（pH 4.5）,the above indicators of the two rice cultivars reached the control level,and even the 100-grain weight of resistant rice“Wuyou 308”was higher than that of the control.Under the combined treatment of Ca²⁺and SAR2（pH 3.0）,the yield of resistant rice“Wuyou308”reached the control level,but the nutritional quality was lower than that of the control,and the yield and nutritional quality were both still lower than thant of the control.This was consistent with the fact that exogenous Ca²⁺effectively alleviated the reduction of total nitrogen content at the seedling and booting stages of the two rice cultivars under SAR1（pH 4.5）treatment,while the mitigation effect of exogenous Ca²⁺under SAR2（pH 3.0）was restricted by SAR intensity,resistant rice“Wuyou 308”had a better mitigation effect.（2）Effect mechanism of exogenous Ca²⁺on nitrogen uptake and transport in different resistant rice under SAR was clarified.The results showed that SAR1（pH 4.5）increased the uptake of NH₄⁺in the two rice cultivars and the uptake of NO₃^-in resistant rice“Wuyou 308”,but decreased the uptake of NO₃^-in sensitive rice“Nanjing 9108”.This may be because SAR1（pH 4.5）induced an increase in the phosphorylation level of plasma membrane H⁺-ATPase in the two rice cultivars,which promoted activity increased and increased ATP decomposition,while up-regulated the expression of ammonium transporter OsAMT1;3 to enhance NH₄⁺uptake ability.SAR1（pH 4.5）up-regulated the expression of nitrate transporter OsNRT2.1 in the roots of resistant rice“Wuyou 308”,but down-regulated the expression of this protein in sensitive rice“Nanjing 9108”.This was one of the reasons why the nitrogen content of resistant rice“Wuyou 308”was higher than that of sensitive rice“Nanjing 9108”under SAR1（pH 4.5）.SAR2（pH3.0）resulted in NO₃^-and NH₄⁺uptake decreasing both in the two rice cultivars.This may be because SAR2（pH 3.0）not only reduced genes（subfamily I,II,and III）expression and activity of H⁺-ATPase in the roots of the two rice cultivars,but also reduced the decomposition of ATP,resulting in a decrease in energy supply.And SAR2（pH 3.0）treatment down-regulated the expression of OsNRT2.1,OsNRT2.2 and OsAMT1;3,reducing the absorption of NO₃^-and NH₄⁺,and sensitive rice“Nanjing 9108”decreased more.This may be one of the reasons why SAR2（pH 3.0）inhibited the total nitrogen level of the two rice cultivars to a greater extent,and sensitive rice“Nanjing 9108”decreased more.Single exogenous Ca²⁺treatment had no effect on the absorption and transport of NO₃^-and NH₄⁺in the two rice cultivars,which may be the reason why the total nitrogen level of the two rice cultivars remained unchanged.The absorption of NO₃^-and NH₄⁺in the two rice cultivars maintained the control level under the treatments of Ca²⁺and SAR1（pH 4.5）.The absorption of NO₃^-and NH₄⁺under combination of Ca²⁺and SAR2（pH3.0）treatment was higher than that under SAR2（pH 3.0）treatment,but still lower than the control,and resistant rice“Wuyou 308”was closer to the control level.This may be because that under the combined of Ca²⁺and SAR（pH 4.5/3.0）treatment,genes（I,II,III subfamily）expression and activity of H⁺-ATPase,ATP content,expressions of OsNRT2.1 and OsAMT1;3 in the roots of the two rice cultivars were closer to the control compared with SAR（pH 4.5/3.0）,which was beneficial to maintain the stable absorption of NO₃^-and NH₄⁺.However,the above indexes were still lower than the control under the combination of Ca²⁺and SAR2（pH 3.0）（except OsNRT2.1 expression in resistant rice“Wuyou 308”）,and the absorption of NO₃^-and NH₄⁺of the two rice cultivars were still lower than that of the control.This may be the reason why exogenous Ca²⁺can alleviate the decrease in total nitrogen level of the two rice cultivars under SAR（pH 4.5/3.0）treatment,but the regulation effect was limited by SAR intensity,and the regulation effect on resistant rice“Wuyou 308”was better.（3）The mechanism of the effect of exogenous Ca²⁺on the nitrogen assimilation efficiency of different resistant rice seedlings under SAR was understood.The results showed that although SAR（pH 4.5/3.0）increased in glutamate dehydrogenase（GDH）activity,the glutamine synthase/glutamate synthase（GS/GOGAT）cycle efficiency decreased,and led to a decrease in NH₄⁺assimilation ability,resulting in the reduction of free amino acid content in leaves.SAR2（pH 3.0）inhibited the GS/GOGAT cycle efficiency more and inhibited the assimilation of NH₄⁺to a greater extent,and also reduced nitrate reductase（NR）activity and reduced the assimilation of NO₃^-,resulting in a decrease in soluble protein content.Sensitive rice“Nanjing 9108”showed a greater degree of inhibition.This may be one of the reasons why SAR（pH 4.5/3.0）decreased the nutritional quality of the two rice cultivars,and the sensitive rice“Nanjing 9108”decreased more.The single Ca²⁺treatment had no effect on the nitrogen assimilation process and products of the two rice cultivars,which may be one of the reasons why the nutritional quality of the two rice cultivars did not change under this treatment.Exogenous Ca²⁺increased the expression of OsGDH1 and GDH activity of leaves in two rice cultivars under SAR（pH 4.5/3.0）treatment,and alleviated the decreased in GS/GOGAT cycle efficiency and NR activity caused by increased in the expression of OsGS1;1,OsGS1;2 and decreased in the expression of OsNADH-GOGAT2 to alleviate the decline in NO₃^-and NH₄⁺assimilation,which was beneficial to alleviate the decrease in free amino acid and soluble protein content.Under combination of Ca²⁺and SAR2（pH 3.0）treatment,the regulation effect of exogenous Ca²⁺on NH₄⁺assimilation ability of the two rice cultivars and NO₃^-assimilation ability of sensitive rice“Nanjing 9108”was limited by SAR intensity.and the alleviation effect of soluble protein content in sensitive rice“Nanjing 9108”was limited.This may be one of the reasons why exogenous Ca²⁺had a mitigating effect on the nutritional quality of different resistant rice under SAR（pH 4.5/3.0）,and the mitigating effect was limited by SAR intensity and variety.In conclusion,the reduction of root nitrogen absorption capacity and leaf nitrogen assimilation capacity of different resistant rice under acid rain stress was one of the reasons for the decline in yield and nutritional quality.Exogenous Ca²⁺maintained root nitrogen uptake by alleviating the inhibition of H⁺-ATPase activity,genes expression of nitrate transporter protein and ammonium transporter under acid rain stress,reducing the decrease in final yield,and even increasing the yield of resistant rice.At the same time,exogenous Ca²⁺alleviated the inhibition of SAR on NR activity,and maintained the balance of GS/GOGAT,and improved GDH activity.These were conducive to the conversion of inorganic nitrogen to organic nitrogen,and maintained amino acids and protein contents in grains,and alleviated the deterioration of nutritional quality.The regulation of exogenous Ca²⁺on rice yield and nutritional quality under acid rain stress was not only limited by acid rain intensity,but also had differences among cultivars.These can provide new thinking directions and a theoretical basis for finding effective ways to reduce the adverse effects of acid rain on agricultural and forestry production.