| Soil salinization has a serious inhibitory effect on plant growth and its yield,which is a serious problem faced by modern agricultural and forestry production.Brassinolide?BRs?is a new plant growth regulator,which is widely involved in various physiological processes of plants,especially in plant growth and development and its response to stress.A large number of studies have shown that BRs was increasingly concerned in the respect of improving plant NaCl resistance,while hitherto researches about its regulating effect on Atropa belladonna L resistance to NaCl remains scarce.In this study,we used Atropa belladonna L.seedlings as materials,and treated with exogenous 2,4-epibrassinolide?EBR?.This study aimed to explicit the physiological mechanism of exogenous BRs on regulating Atropa belladonna L.NaCl resistance,by studying the effect of different concentrations(0.05 mg·L-1,0.1mg·L-1,0.2 mg·L-1,0.4mg·L-1)of EBR on the photosynthesis,physiological indexes of adversity,nitrogen metabolism,secondary metabolite content,precursor substances and gene expression of key enzymes in the tropane alkaloid synthesis pathway of Atropa belladonna L.at different treatment times?5 d,10 d,15 d,20 d?under 100 mmol.L-11 NaCl stress.The main results are as follows:1.Appropriate concentration of EBR can effectively increase the content of photosynthetic pigments in the leaves of Atropa belladonna L.under salt stress,and alleviate the damage of PS?100 mmol·L-11 NaCl stress significantly reduced the content of photosynthetic pigments,the PS?response center was damaged,and photosynthesis was inhibited.After the addition of exogenous EBR,the chlorophyll a,chlorophyll b,total chlorophyll content and carotenoid content increased to varying degrees;maximum photochemical efficiency?Fv/Fm?,photochemical quenching coefficient?qP?,actual photochemical quantum yield?Yield?showed an upward trend;initial fluorescence?F0?and non-photochemical quenching coefficient?NPQ?gradually decreased.It shows that EBR can effectively alleviate the photoinhibition caused by NaCl stress,facilitate the photochemical efficiency,and enhance the resistance to NaCl stress.On the whole,0.1 mg·L-1 EBR has the best treatment effect.2.Exogenous EBR can maintain the balance of osmotic potential and reduce the damage of NaCl stress to Atropa belladonna L.'s antioxidant system.With the increase of the number of NaCl stress days,the content of proline and soluble sugar increased significantly;SOD and POD activities increased first and then decreased,and CAT activity continued to decrease;the content of membrane lipid peroxidation product MDA increased remarkably.It indicates that Atropa belladonna L.has been subjected to osmotic stress,and membrane lipid peroxidation is intensified.After EBR treatment,the content of proline and soluble sugar was further increased,boosted the activity of the three antioxidant enzymes,and the content of MDA decreased significantly.It shows that EBR can slow down the inhibitory effect of NaCl stress on seedling growth,increase antioxidant enzyme activity and osmoregulation substance content under NaCl stress,reduce the degree of membrane lipid peroxidation,and enhance NaCl tolerance of Atropa belladonna L..3.Exogenous EBR can effectively enhance the nitrogen metabolism of Atropa belladonna L.under NaCl stress.The results showed that with the NaCl stress time prolonged,the nitrate nitrogen content decreased notably,and ammonium nitrogen accumulated in large quantities;GDH activity increased first and then decreased.The free amino acid,soluble protein,nitrate reductase?NR?activity and glutamine synthetase?GS?activity continued to decrease.In addition,the content of ornithine and arginine was decreased notably.It was shown that NaCl stress reduced the absorption of nitrate nitrogen,caused a large accumulation of ammonium nitrogen to poison cells,reduced the activity of key enzymes,and disordered the nitrogen metabolism pathway.0.1 mg·L-1and 0.2 mg·L-1 EBR both had better effects on reduce ammonium nitrogen content,increase nitrate nitrogen,free amino acids,soluble proteins,and ornithine and arginine content;meanwhile,the activity of NR,GS and GDH were facilitated,which shows that exogenous EBR can promote the absorption and utilization of NO3--N by increasing the activity of key enzymes in nitrogen metabolism,accelerate the assimilation of ammonia,increase the content of nitrogenous organic compounds,and provide more material basis for secondary metabolism,validly regulated nitrogen metabolism of Atropa belladonna L..4.Under NaCl stress,the application of 0.1 mg·L-1EBR can effectively increase the polyamine content,and increase the content of scopolamine and scopolamine by up-regulating the expression of key enzyme genes in the TAs pathway.Investigating the effects of tropane alkaloid metabolism under salt stress.The results showed that the scopolamine content increased first and then decreased,while the scopolamine content have been steadily decreasing.Then the content of scopolamine and scopolamine have risen,which showed that when the EBR concentration was 0.1 mg·L-1,the effect of increasing the content of scopolamine and scopolamine was the best.In order to further explore its regulatory mechanism,using Atropa belladonna L.seedlings at 20 days as the material,the polyamine?Put,Spd,Spm?content,Put synthesis key enzyme activity,and the relative expression of key enzyme gene?PMT,TR?,H6H and PPAR?in TAs synthesis pathway were detected.We found that NaCl stress significantly reduced the polyamine content and the activity of put synthesis key enzymes,increased the expression of TRI gene,and suppressed the expression levels of PMT?H6H and PPAR.0.1 mg·L-1 exogenous EBR treatment significantly increased the content of polyamines and the total amount of polyamines,increased the ratio of?Spd+Spm?/Put,and reduced the ratio of Put/PAs.And the relative gene expression levels of TR?and H6H up-regulation notably.It is suggested that EBR can effectively alleviate the inhibitory effect of NaCl stress on Atropa belladonna L.secondary metabolism,increase the content of precursor substances and key enzyme activities in the TAs synthesis pathway,and up-regulate the expression of key enzyme gene,then promote the accumulation of scopolamine and scopolamine.In summary,NaCl stress has a certain degree of effect on the primary and secondary metabolism of Atropa belladonna L,which inhibits the progress of photosynthesis,nitrogen metabolism,and tropane alkaloid synthesis pathways,exacerbate the degree of membrane lipid peroxidation.under NaCl stress,the addition of exogenous EBR can alleviate NaCl stress on photosynthesis and enhance resistance Oxidation system activity,increase nitrogen metabolism capacity,promote the accumulation of tropane alkaloids,and improve the NaCl tolerance of Atropa belladonna L.. |
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