NO对盐胁迫下苜蓿根系生长抑制及氧化损伤的缓解效应

Nitric oxide protection of alfalfa seedling roots against salt-induced inhibition of growth and oxidative damage

以"甘农4号"苜蓿品种为材料,采用水培法,用NO供体硝普钠(SNP)、硝普钠类似物亚铁氰化钠(不产生NO)、NO特异清除剂c-PTIO、一氧化氮合酶(NOS)活性抑制剂N-硝基-L-精氨酸甲脂(L-NAME)、硝酸还原酶(NR)活性抑制剂钨酸盐处理苜蓿植株,研究NO对盐胁迫下苜蓿幼苗根系生长、根系活力、根系中渗透调节物质、膜脂过氧化、活性氧含量及抗氧化酶活性等的影响,探讨NO调控苜蓿幼苗根系耐盐性的生理机制。结果表明:盐胁迫下SNP处理提高了根系活力,促进了苜蓿幼苗根系生长,降低游离脯氨酸含量,促进可溶性蛋白含量增加;增强超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、愈创木酚过氧化物酶(GPX)、抗坏血酸过氧化物酶(APX)和谷胱甘肽还原酶(GR)活性,提高还原型抗坏血酸(As A)和还原型谷胱甘肽(GSH)含量,降低过氧化氢(H2O2)、羟自由基(OH·)含量、超氧阴离子(O·-2)产生速率和膜脂过氧化产物丙二醛(MDA)含量;同时,SNP处理显著促进了苜蓿幼苗根系内源NO的积累。NO供体SNP的类似物亚铁氰化钠对盐胁迫下苜蓿根系各项生理生化指标无明显影响;盐胁迫下添加c-PTIO、L-NAME和钨酸盐进一步降低了苜蓿幼苗根系活力和根系生长,抑制了根系抗氧化系统活性,加剧了根系膜脂过氧化作用,降低了内源NO积累,添加SNP则能缓解该抑制效应;表明外源SNP处理能明显缓解盐胁迫对苜蓿幼苗根系生长的抑制和氧化损伤,且通过NOS和NR途径产生的内源NO也可能在苜蓿根系适应盐胁迫的调节中起关键作用;该研究结果为苜蓿耐盐机制及NO在苜蓿耐盐育种、化学调控和盐碱地栽培利用等提供了理论依据。

Roots play important roles in anchoring plants in the soil and absorbing water and nutrients from the soil. Soil salinity is one of the major abiotic stresses that adversely affect root growth and development. The exogenous application of some bioactive substances is a simple and effective approach to improve plant stress tolerance in agricultural production.Nitric oxide（ NO） is a bioactive,gaseous,multifunctional molecule which plays a central role and mediates a variety of physiological processes and responses to biotic and abiotic stresses,including salt. However,information on the effects of exogenous SNP on alfalfa root systems under salt stress conditions and its physiological mechanisms is lacking. In this study,Medicago sativa L. cv. Gannong No. 4 was used as the experimental material and NO-donor sodium nitroprusside（SNP）,NO-scavenger 2-（4-carboxyphenyl）-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide（ c-PTIO）,tungstate,the nitrate reductase（ NR） inhibitor,nitric oxide synthase（ NOS） inhibitor NG-nitro-L-Arg-methyl ester（ L-NAME） and sodium ferrocyanide（ SNP analogue,does not release NO） were used to investigate the effects of NO on growth,root vigor,osmoregulatory molecules,membrane lipid peroxidation,reactive oxygen species（ ROS） accumulation and the activity of antioxidant enzyme in alfalfa roots under Na Cl stress. Results showed that when 100 μmol / L SNP was applied in the nutrient solution under Na Cl stress,the growth of alfalfa roots was significantly improved for eight days,the root vigor value and the content of free proline were increased and the content of soluble protein was decreased throughout the treatment period. SNP also caused an increase in the activities of ascorbate peroxidase（ APX）,catalase（ CAT）,glutathione reductase（ GR）,guaiacol peroxidase（ GPX）,superoxide dismutase（ SOD）,and the contents of reduced ascorbic acid（As A） and reduced glutathion（GSH） under salt stress. Whereas,the content of H2O2,OH^·,production rate of O2^·- and MDA level were decreased in alfalfa roots. Meanwhile,SNP pretreatment improved the endogenous NO accumulation.Sodium ferrocyanide,the NO-donor SNP analogue,has no noticeable effect on the damage caused by Na Cl stress for alfalfa roots. When applied with the NO-scavenger c-PTIO,under Na Cl stress,the NR inhibitor tungstate and NOS inhibitor L-NAME both reduced the accumulation of endogenous NO in roots,further inhibited the activity of the antioxidant system,aggravated membrane lipid peroxidation and root growth,however,their harmful effects on growth,antioxidant system and membrane lipid peroxidation could be reversed by the addition of SNP. Thus,the results showed that application of SNP significantly increased root vigor,activated the antioxidant system,alleviated the oxidative root damage induced by salt stress and promoted root growth under Na Cl stress. Endogenous NO may also take part in the regulation of salt-tolerance of alfalfa roots under Na Cl stress,and the release of NO according to the NOS and NR pathways may play an important role in alleviating the inhibitive effect in alfalfa root growth. These results may provide a theoretical basis for the mechanisms of salt resistance,help research into NO in the chemical regulation of alfalfa salt tolerance,the breeding of salt tolerant alfalfa cultivars,and the utilization of saline land in the future.