Physiological Mechanism Of 24-Epibrassinolide-Regulated Salt Stress Tolerance In Medicago Sativa

Prevailing in arid and semi-arid soil, salinization has a strong impact on plant growth, development and yield, and has become a severe problem of modernized agriculture. Brassinosteroids(BRs), generally existed in plant and extensively used in agricultural production, is a new type of sterols phytohormone which could regulate growth and development of plants and enhance their resistance to adverse stress. As an vital growth regulator in the response process of plant to biotic and abiotic stress, BRs was increasingly concerned in the respect of improving plant Na Cl resistance, while hitherto researches about its regulating effect on alfalfa resistance to Na Cl remains scarce. This study aimed to explicit the physiological mechanism of exogenous BRs on regulating alfalfa Na Cl resistance by treating the seed and seedling of alfalfa with 24-epibrassinolide(EBR). Taken Medicago sativa L. cv. Gannong No. 3 as material, effects of exogenous EBR on seed germination, seedling growth, antioxidant system, photosynthesis, ion metabolism and carbon and nitrogen metabolism under Na Cl stress were determined. Emerging from the results were that:(1) Seed germination, growth of seedling and root of alfalfa were restrained obviously under 150 mmol·L?1 Na Cl stress, root vigor, aboveground and belowground biomass of seedling reduced as well. Under Na Cl stress, exogenous EBR facilitated seed germination, growth of seedling and root of alfalfa remarkably. The promotion effect of low concentration of exogenous EBR was superior to others, and this effect declined even turned to inhibiting effect with the growth of concentration. Seed germination rate, germination potential, germination index, vigor index, radical and bud length increased apparently after adding 0.1 μmol·L?1 EBR, and the plant height, axial root length, root system length, root total surface area, root volume, root tip number and root vigor were increased as well as biomass accumulation of alfalfa seedling. Compared with Na Cl treatment, germination rate and germination potential were facilitated by 50.85% and 98.70%, and root vigor, aboveground and belowground biomass were enhanced by 35.23%, 17.50% and 25.87%, respectively.(2) The antioxidant enzymes activities in the root system of alfalfa seedling decreased remarkably under Na Cl stress, which also resulted in an increase in GSH and As A content in leaf and root, and a fall in GSH/GSSG and As A/DHA ratio. Na Cl stress could induce the rise of ROS level in alfalfa seedling, the increase of relative permeability of plasma and the aggravation of membrane lipid peroxidation. Under Na Cl stress, exogenous EBR boosted the activity of SOD、APX、GPX、CAT and GR as well as As A and GSH content in alfalfa seedling. As A/DHA, GSH/GSSG ratio and the activity of DHAR and MDHAR were enhanced, while AAO activity was declined. As A-GSH circulation and total antioxidant capacity of alfalfa seedling were accelerated, while content of MDA and H2O2, OFR generation rate, OH concentration and relative permeability of plasma were decreased significantly under this condition, which validly mitigated the oxidative damage caused by Na Cl stress.(3) Under Na Cl stress, photosynthetic pigment content and Pn of alfalfa seedling decreased significantly. The activity of PSⅡ reaction center declined as well. Na Cl stress also led to less optical energy which absorbed by chloroplast be used in photochemical reaction and growth in antenna heat dissipation and excess energy of reaction center, which lowered the photosynthetic capacity of alfalfa seedling. Under Na Cl stress, exogenous EBR increased the content of Chla, Chlb, Chla+b and Chlx·c as well as Chla/Chlb ratio significantly. Chl/Car ratio was decreased but Pn, Tr, Gs and WUE increased. Response curve of Pn-SPR and Pn-CO2 indicated that there were notable growth in Lsp, AQY, Pnmax, Cisat, α and Pmax, and significant drop in Γ and Rp. With dramatical reduce in leaf F0 and NPQ, and remarkable improvement in Fm, Fv/F0, Fv/Fm, ФPSⅡ, Fv′/Fm′, q P and ETR, the utilization of exogenous EBR under Na Cl stress increased the energy assigned in photochemical reaction, lowered the antenna heat dissipation and excess energy of reaction center and enhanced photochemistry activity of PSⅡ. The partition equilibrium of excitation energy between PSⅠ and PSⅡ was regulated effectively. Carbon assimilation efficiency and photosynthetic capacity of alfalfa seedling were facilitated after adding exogenous EBR under Na Cl stress.(4) Na Cl stress increased the content of Na+, Cl? and Cu2+ in different organs(leaf, stem and root) of alfalfa seedling dramatically, while reduced the content of K+, Ca2+, Mg2+, Fe2+, Mn2+, Zn2+ and the ratio of K+/Na+, Ca2+/Na+, Mg2+/Na+, Fe2+/Na+, Mn2+/Na+, Cu2+/Na+ and Zn2+/Na+ significantly, which caused the imbalance of metabolic processes in alfalfa seedling such as the absorption, transportation and allocation of ionic, and destroy of ion homeostasis in cell. Under Na Cl stress, exogenous EBR lowered the content of Na+, Cl? and Cu2+ remarkably, while increased the content of K+, Ca2+, Mg2+, Fe2+, Mn2+, Zn2+ and the ratio of K+/Na+, Ca2+/Na+, Mg2+/Na+, Fe2+/Na+, Mn2+/Na+, Cu2+/Na+ and Zn2+/Na+ notably. Meanwhile, the antagonism between Na+ and other positive ion such as K+, Ca2+, Mg2+, Fe2+, Mn2+, Cu2+ and Zn2+ were decreased under this condition. As a result, there were promotions in inorganic ion absorption and validly regulation in transportation and allocation of inorganic ion in alfalfa seedling, which contributed to ion homeostasis maintenance in cell.(5) Proteolytic enzymatic activity in alfalfa seedling was enhanced significantly under Na Cl stress, which facilitated the protein hydrolysis. The activity of NR、GS and GOGAT were decreased and GDH activity increased. Na Cl stress resulted in the NH4+-N accumulation and suppression in absorption and transportation of NO3?-N. Nitrogen metabolic capacity was refined and total nitrogen content was reduced. Under Na Cl stress, exogenous EBR decreased proteolytic enzymatic activity, suppressed the protein hydrolysis and reduced the content of free amino acid and free proline dramatically. Meanwhile, the activity of NR, GS and GOGAT were facilitated and GDH activity was decreased notably. This indicated that exogenous EBR could improve the absorption and utilization of NO3?-N and accelerate ammoniac assimilation by synergistic effects such as GS-GOGAT, GDH and NR activity alteration. The content of NH4+-N in alfalfa seedling was decreased and total nitrogen content of plant was boosted. The utilization of exogenous EBR under Na Cl stress validly regulated nitrogen metabolism and mitigated the ammoniac poison to alfalfa seedling caused by Na Cl.(6) Na Cl stress suppressed the transportation of sucrose from leaf to root in alfalfa seedling significantly. The content of starch, fructose and glucose in leaf were boosted dramatically, while that in root was dropped. Na Cl stress, which inhabited the activity of enzymes associated with carbohydrates metabolism, leaded to the disruption of synthesis and decomposition balance of sucrose as well as the disorder of carbohydrates metabolism. Under Na Cl stress, exogenous EBR facilitated the transportation of sucrose from leaf to root in alfalfa seedling. The content of starch in leaf was decreased but that in root was increased. The content of sucrose, fructose and glucose and the activity of SS and SPS in alfalfa seedling were enhanced. There was also a regulation in the activity of AI, NI, S and SP. The utilization of exogenous EBR under Na Cl stress lowered the negative feedback of carbohydrates accumulation on photosynthesis by the synergistic effects among the approaches of Ivr, SS and SPS. As a result, the synthesis and decomposition balance of sucrose was maintained, and synthesis and accumulation of photosynthesis assimilation production were facilitated, which contributed to the regular performance of carbohydrates metabolism.In conclusion, under Na Cl stress, exogenous EBR could facilitate seed germination and seedling growth of alfalfa, and enhance the activity of antioxidant system in seedling. EBR also could contribute to acceleration of the As A-GSH circulation, decrease of the ROS level and increase of the carbon assimilation efficiency and photosynthesis capacity. The absorption, transportation and distribution of ion were also validly regulated under this condition. Nitrogen metabolism capacity of seedling was increased by synergistic effects among GS-GOGAT, GDH and NR activity alteration, and regular carbohydrates metabolism was maintained by the synergistic effects among the approaches of Ivr, SS and SPS as well after adding exogenous EBR under Na Cl stress, which resulted in an improvement in seedling salt resistance.