Effects Of Exogenous Calcium On Photosynthetic Carbon Metabolism In Wheat Seedlings Under NaCl Stress

Soil salinization is the main environmental factor that affects plant growth and restricts agricultural production worldwide.It is of great significance to study and analyze the mechanism of plant salt stress response for developing new approaches to improve crop salt tolerance.Ca²⁺is an important signaling substance in plants,which plays an important role in enhancing photosynthetic capacity and plant growth.Its entire cellular signaling network is critical for plant response to abiotic stress.However,the research on the effect of exogenous Ca²⁺on the photosynthesis of sodium-salt-stressed plants mainly focuses on the study of photosystem Ⅱ（PSⅡ）,and there are few related studies on the accumulation of photosynthetic products and the distribution relationship between source and sink tissues under sodium salt stress.In addition,the mechanism of the Ca²⁺-CaM messenger system in the regulation of photosynthesis needs to be further elucidated.In our previous study,different concentrations of NaCl,including 150 mmol·L^-1NaCl,significantly inhibited the growth of wheat seedlings,and showed obvious toxic effects on"Xihan No.3"seedlings.In this experiment,150 mmol·L^-1NaCl or/and different concentrations of CaCl₂were treated alone or in combination to analyze the effects of exogenous calcium on the growth and photosynthetic characteristics of wheat seedlings under sodium salt stress.Furthermore,calcium effectors such as calcium chelator（EGTA）and calcium channel blocker（LaCl₃）were used to further explore the mechanism of exogenous CaCl₂on photosynthetic carbon metabolism of wheat seedlings under NaCl stress from two aspects of enhancing and preventing calcium signaling system in plants.The main results are as follows:1.Compared with the control,150 mmol·L^-1NaCl stress significantly inhibited the growth of stem and root of wheat seedlings,significantly increased the proline content,while the total ascorbic acid（AsA）content notably decreased in leaves.Different concentrations of CaCl₂(0.1,0.5 and 1 mmol·L^-1)showed different effects on the growth of wheat seedlings,which significantly reduced the proline content but increased the total AsA content.In addition,the addition of 0.5 and 1 mmol·L^-1CaCl₂significantly promoted the gro???祦?????槒??賮苟?????燕懔?ted wheat seedlings,which significantly rose the dry weight and total AsA content of stem in comparison with those treated with sodium salt alone.2.Photosynthetic pigment content of wheat seedlings decreased significantly,while chlorophyll a/chlorophyll b（Chl a/b）elevated obviously due to150 mmol·L^-1NaCl stress.The maximum photochemical efficiency（Fv/Fm）,actual photochemical efficiency（Y（Ⅱ））,electron transfer rate（ETR）and photochemical quenching coefficient（qP）of chlorophyll fluorescence parameters reduced remarkably,while the yield of dissipative electron（Y（NPQ））and non-photochemical quenching coefficient（NPQ）increased significantly.At the same time,the net photosynthetic rate（Pn）,transpiration rate（Tr）,intercellular CO₂concentration（Ci）and stomatal conductance（Gs）of wheat seedlings were significantly decreased under sodium salt treatment.The amount of lutein,ETR,Pn and Tr were significantly rose when wheat seedlings treated with 0.1 mmol·L^-1CaCl₂alone,while other indexes had no significant changes in comparison with the control.The photosynthetic pigment content,Fv/Fm,Y（Ⅱ）,qP,ETR,Pn and Ci of wheat leaves increased significantly under 0.5 and 1 mmol·L^-1CaCl₂treatment alone or in combination with NaCl,while Y（NPQ）and NPQ reduced significantly,and 150 mmol·L^-1NaCl+0.5mmol·L^-1CaCl₂treatment caused the largest change.However,different concentration???鸟??酅??槒?篟?担????腾奸??lone or in combination with NaCl did not affect the relative content of zeaxanthin.3.Compared with control,sodium salt treatment significantly decreased Ca²⁺and calmodulin（CaM）contents in leaves and roots of wheat seedlings,as well as calcium-dependent protein kinases（CDPKs）and Ca²⁺-ATPase activities.Ca²⁺content significantly elevated in wheat leaves,while this parameter was not been affected in the roots when wheat seedlings treated with 0.5 and 1 mmol·L^-1CaCl₂alone or in combination with NaCl.In addition,the two concentrations of calcium alone led to a significant increase in the CaM content of seedlings,the activity of CDPKs in leaves was significantly enhanced in the early treatment period,while the Ca²⁺-ATPase activity was only significantly enhanced on the 6 days of treatment.Further study found that the addition of these two concentrations of CaCl₂effectively slowed down the decrease of CaM content,CDPKs and Ca²⁺-ATPase activities caused by NaCl treatment.And on the 4days of complex treatment of NaCl and different concentrations of CaCl₂,these parameters showed a trend of first increasing and then decreasing with the increase of the concentration of exogenous CaCl₂.That is,when 0.5 mmol·L^-1CaCl₂was added,the CaM content and Ca²⁺-ATPase in the leaves of the seedlings treated with sodium salt for4 days increased the most compared with those treated with sodium salt alone.4.Quantitative PCR analysis showed that the expressions of TaCaM,TaCDPK and TaCa²⁺-ATPase in leaves of wheat seedlings treated with sodium salt for 2,4 and 6 days were significantly down-regulated compared with the control.The expression of TaCaM was up-regulated in the leaves of seedlings treated with different concentrations of CaCl₂alone at different stages.The difference was that TaCDPK in leaves of mid-stage seedlings treated with three concentrations of CaCl₂alone and TaCa²⁺-ATPase expression in early and mid-stage treatments with 0.5 and 1 mmol·L^-1CaCl₂were significantly up-regulated.In addition,the addition of three concentrations of CaCl₂effectively relieved the inhibitory effect of sodium salt treatment on the expression of TaCaM and TaCa²⁺-ATPase in seedling leaves,especially the expression of these two genes in the early and middle stages of sodium salt treatment.Only 1 mmol·L^-1exogenous CaCl₂significantly increased the expression of TaCDPK in leaves of sodium salt-treated seedlings.5.150 mmol·L^-1NaCl or different calcium effector(0.5 mmol·L^-1LaCl₃,0.5mmol·L^-1EGTA)significantly increased the amount of soluble sugar and sucrose,whereas the starch content was significantly reduced in wheat seedlings.The activities ofα-amylase,β-amylase,total amylase,GAPDH and PGK were significantly increased,while ribulose-1,5-bisphosphate carboxylation/oxygenase（Rubisco）and Rubisco activase（RCA）activities were significantly reduced.The addition of 0.5 mmol·L^-1CaCl₂significantly alleviated the increase of soluble sugar and sucrose content and the decrease of starch content induced by NaCl.However,the activities of Rubisco and RCA were notably increased and the activities of amylase,GAPDH and PGK were obviously decreased.In comparison with the combined treatment of NaCl+0.5 mmol·L^-1CaCl₂,the addition of LaCl₃and EGTA eliminated the changes of various indexes of sodium salt-treated wheat seedlings caused by the presence of exogenous CaCl₂,and even enhanced the damage of NaCl stress on wheat seedlings.6.Salt stress significantly elevated the activities of sucrose phosphosynthase（SPS）and sucrose synthase（Su Sy）,while the activities of acid invertase（SAInv）and alkaline/neutral invertase（A/N-Inv）were obviously weakened,compared with the control.Meanwhile,the expressions of TaSPS and TaSu Sy were up-regulated while the expressions of TaSAInv and TaA/N-Inv were down-regulated.The addition of 0.5mmol·L^-1CaCl₂increased the activity of SA-Inv in sodium salt-treated wheat seedlings,but decreased the activity of SPS and Su Sy.And the expression of TaSAInv、TaA/N-Inv was promoted,while without affecting A/N-Inv activity.However,the further addition of EGTA and LaCl₃significantly weakened the effect of CaCl₂and aggravated the toxic effect of sodium salt on seedlings.In conclusion,the addition of an appropriate concentration of calcium effectively alleviated the inhibitory effect of sodium salt stress on the growth and photosynthetic efficiency of wheat seedlings,which is mainly manifested in the increase of salt-induced photosynthetic pigments,the acceleration of light energy conversion efficiency and electron transfer efficiency,which might be related to the increase of photosynthetic pigments,light energy conversion efficiency and electron transfer efficiency in wheat seedlings treated with sodium salt due to the addition of exogenous CaCl₂,or it might be related to the addition of exogenous calcium to increase the intracellular calcium content and activate the calcium signal regulation mechanism.Appropriate concentration of exogenous calcium could regulate the balance of Ca²⁺uptake in leaves of sodium salt-treated wheat seedlings by enhancing the expression of calcium signal-related proteins in leaves.In addition,the addition of calcium at an appropriate concentration inhibited the activities of SPS and Su Sy and gene expression of sodiun salt-treated wheat seedlings,and enhanced the anabolism of starch,resulting in a significant decrease in sucrose content and a significant increase in starch content.However,the addition of different calcium effectors（LaCl₃and EGTA）blocked the transmission of calcium signal,affected the process of photosynthetic carbon assimilation,inhibited the metabolism of carbohydrates,and further increased the damage of sodium salt stress on wheat seedlings.