镉诱导油菜叶片氧化胁迫及硫化物的络合作用

研究了不同浓度Cd(0、10、20、30、40μmol／L)对油菜(Brassica napus L．)“油研—8号”叶片H2O2积累、膜脂过氧化作用以及与硫代谢有关的抗镉机制。结果表明,随着Cd处理浓度的增加,叶片中H2O2及MDA的积累量均较对照显著增加,但对缺硫处理组( Cd—S)与未缺硫处理组( Cd S)无显著差异。10～30μmol／L Cd诱导过氧化氢酶(Catalase,CdT)、过氧化物酶(Guaiacol peroxidase,POD)的活力提高,40μmol／L Cd对CdT活力有抑制作用。 Cd—S组的POD活力显著高于 Cd S组。在Cd的诱导下,根部非蛋白巯基及酸不稳定性硫的含量大幅度增加。缺硫处理可明显阻碍上述含硫化合物的合成,从而加重Cd对叶片的伤害。上述结果表明,Cd对活性氧清除酶活性的诱导提高并非是抗镉关键途径,而与硫代谢有关的对Cd的络合作用才是油菜抗镉的重要机制。     

人谷胱甘肽硫转化酶基因在链霉菌的高效表达

谷胱甘肽硫转化酶(GST)是一个具有广泛底物专一性,与一些化学致癌物代谢有关的酶的一个家族.它既有谷胱甘肽硫转化酶的活性,也有过氧化物酶的活性,能使谷胱甘肽分子上的巯基(SH)与广泛的亚硝基本类化合物结合,使这类致癌物转化成无毒的化合物.它亦能与细胞内源的一些阴离子化合物(如胆红素和亚铁血红素)结合并参与运输.在人体中,GST的缺乏常与新生儿非溶血性的胆红素积累而导致的病因有关,同时GST在保护生物体过氧化反应的损伤中也起着重要的作用.     

硫和6-苄氨基腺嘌呤对水稻幼苗硝酸还原酶活性的影响

缺硫培养6天的水稻幼苗,其叶片和根中的硝酸还原酶(NR)活性明显下降。用1pPm 的6-苄氨基腺嘌呤(6-BA)处理培养了10天的水稻幼苗根系,24小时后缺硫培养的水稻幼苗叶片和根系的 NR 活性升高,加硫培养的水稻幼苗叶片和根中的 NR 活性下降。用~(35)S示踪发现,6-BA 可降低加硫幼苗对~(35)S 的吸收和转化,但促进缺硫幼苗对~(35)S 的转化。     

外源一氧化氮对镉胁迫下绿豆幼苗根尖抗氧化酶的影响

采用水培法研究外源一氧化氮对镉(Cd)胁迫下绿豆幼苗根尖抗氧化酶活性的影响。结果表明:0.01mmol/L和0.1mmol/L一氧化氮供体硝普钠(sodium nitroprusside,SNP)显著促进上胚轴生长,1mmol/LSNP则抑制绿豆幼苗生长。Cd单独处理抑制根尖抗坏血酸过氧化物酶(ascorbate peroxidase,APX)和超氧化物歧化酶(superoxide dismutase,SOD)活性而刺激脂氧合酶(lipoxygenase,LOX)、谷胱甘肽转硫酶(glutathione S-transferase,GST)、谷胱甘肽还原酶(glutathione reductase,GR)和过氧化物酶(guaiacol peroxidase,POD)活性上升。0.1mmol/LSNP预处理能够明显缓解Cd对根生长的抑制,降低根尖中MDA含量,提高根尖APX和SOD活性,降低LOX和POD活性,但不影响GST和GR活性。     

谷胱甘肽硫转移酶基因表达的调控

催化内源性或外源性亲电子化合物与谷胱甘肽(GSH)结合的谷胱甘肽硫转移酶(GST)超基因家族是一族解毒功能蛋白.其基因的表达通过不同的机制受多种物质的调控.根据最近文献资料,对调控谷胱甘肽硫转移酶基因表达的基因结构、调控机制及氧化应激对谷胱甘肽硫转移酶基因表达的调控作用等作一简要综述.     

外源硫与乙烯缓解马齿苋镉胁迫的生理机制研究

解析镉(Cd)胁迫下外源乙烯和硫(S)对马齿苋(Portulaca oleracea L.)生理响应的机制。在Cd胁迫下,检测乙烯利(乙烯供体)和(NH4)2SO4对马齿苋叶片氧化胁迫、S同化吸收、葡萄糖含量、乙烯合成、光合作用的影响。结果显示,外源S和乙烯处理,可降低Cd胁迫下马齿苋叶片与根中的Cd含量;增强ATP硫酸化酶(ATP-S)和谷胱甘肽还原酶(GR)活性,提升马齿苋叶片还原型谷胱甘肽(GSH)含量,从而降低叶片H2O2与丙二醛(MDA)含量;增强1-氨基环丙烷羧酸合酶(ACS)活性,提升叶片乙烯含量,从而降低葡萄糖含量,提升Rubisco酶活性和光合作用。同时添加外源S和乙烯,对上述指标的促进或者抑制效果加强,而添加乙烯活性专一性抑制剂二环庚二烯(Norbornadiene,NBD)则对上述各指标具有反向作用。外源S和乙烯可以通过降低马齿苋对Cd的吸收,增强体内乙烯信号传导途径,促进GSH的合成,降低葡萄糖含量,从而缓解Cd胁迫诱导的氧化胁迫和葡萄糖介导的光合抑制作用。     

2种黄芪幼苗对缺硫胁迫的生理响应机制

以药用植物膜荚黄芪（Astragalus membranaceus（Fisch.）Bge.）及其变种蒙古黄芪（Astragalus membranaceus Bge.var.mongholicus Hsiao）幼苗为材料，在水培条件下研究了两种黄芪幼苗在不同程度缺硫条件下的生物量积累、光合参数、次生代谢产物积累等的变化规律。结果显示：与供应Hoagland全营养液相比，低硫和无硫供应处理均显著抑制了两种黄芪幼苗根系和地上部的伸长生长以及生物量的累积，并且引起了植株叶片失绿变黄、根系褐变等一系列缺硫症状。此外，缺硫还导致了两种黄芪幼苗叶片叶绿素含量降低以及光合参数下降、全株各部位细胞膜脂过氧化程度增加。相对膜荚黄芪而言，不同程度缺硫对蒙古黄芪生长发育的影响较小。进一步研究发现，在低硫或无硫条件下，膜荚黄芪植株各部位总酚含量出现了明显下降，而蒙古黄芪根系和叶片总酚含量却因缺硫处理而显著升高。同时我们还发现，硫营养缺乏诱导了蒙古黄芪叶片非光化学能量耗散的升高以及植株各部位主要异黄酮物质的大量积累。我们推断，营养液中硫元素供应的减少引起了黄芪幼苗硫营养不良，影响了植株叶绿素合成，降低了植株的光合能力，并同时引起了全株的氧化胁迫，最终使得黄芪幼苗生长发育受到抑制；蒙古黄芪在缺硫胁迫下提高了光保护能力，合成了较多的异黄酮类物质，有效缓解了缺硫胁迫对其生长的影响，从而在面对缺硫胁迫时比膜荚黄芪表现出了更强的耐性。     

干旱高温胁迫下转基因水稻的生理变化

用PEG-6000、38℃及PEG-6000+38℃胁迫处理携带谷胱甘肽转移酶(GST)和过氧化氢酶(CAT1)的转基因水稻和非转基因水稻(Oryza sativa L.cv中花11), 并比较分析了二者在胁迫下的生理指标和抗氧化酶活性变化.结果显示,在单一PEG及PEG和高温复合胁迫下,植株生长、光合参数和相对含水量的降低幅度及H2O2和MDA(malon dialdehyde)的积累量在非转基因水稻与转基因水稻之间都有显著差异.在这2种胁迫下,转基因水稻的可溶性糖含量及CAT和POD (Peroxidase) 活性与非转基因水稻也有显著差异.这些结果表明,谷胱甘肽转移酶(GST)和过氧化氢酶(CAT1)的表达减轻了转基因水稻在单一干旱及干旱+高温复合胁迫下的伤害.     

镉胁迫对旱柳细胞膜透性和抗氧化酶活性的影响

以‘Fuyang 3’旱柳为材料,通过水培方法研究了中、高剂量(5、25μmol.L-1)镉胁迫下旱柳器官的镉(Cd2 )积累、叶和根细胞膜渗透及抗氧化酶活性的变化。结果表明:Cd2 主要积累在旱柳的根部(最高达1 297.71μg.g-1),其次是枝条(最高为163.13μg.g-1)。中、高剂量Cd2 胁迫下,旱柳叶相对电导率、根K 渗透以及根和叶丙二醛(MDA)含量均未发生显著变化。中、高剂量Cd2 胁迫使旱柳根系的超氧歧化酶(SOD)、抗坏血酸过氧化物酶(APX)和谷胱甘肽转移酶(GST)活性,以及叶片的SOD、愈创木酚过氧化酶(POD)、谷胱甘肽过氧化物酶(GPX)和GST活性均比对照显著增强;且高剂量胁迫旱柳根系的SOD和GST活性,及其叶片的POD、GPX和GST活性均显著高于相应中剂量胁迫。研究发现,旱柳在不同浓度镉胁迫条件下,其根是Cd2 主要积累器官,其叶和根细胞质膜能保持相对稳定性,其根和叶各抗氧化酶活性发生不同程度改变,从而使旱柳对Cd2 胁迫表现出一定的忍耐性。     

镉污染对不同生境拟水狼蛛氧化酶和金属硫蛋白应激的影响

为探讨镉(Cd)对机体抗氧化功能及金属硫蛋白(MT)的影响,在室内分别用不添加Cd2 和添加浓度为20 mgkg-1 Cd2 培养基培养的黑腹果蝇来饲喂4种不同生境下(S1,S2,S3和S4)拟水狼蛛,于饲喂5d、10d和20d后,分别测定其体内MT和丙二醛 (MDA)的含量及超抗氧化酶(GST、SOD和CAT)活性。结果表明：1,不同生境拟水狼蛛用不添加Cd2 培养基培养的黑腹果蝇饲喂后,不添加Cd2 对照组拟水狼蛛镉的积累量和MT含量无显著变化,但均显著低于添加Cd2 污染组。添加Cd2 污染组拟水狼蛛镉的积累量和MT含量都显著高于对照组,且均随着饲喂时间的延长而显著升高,具有明显的时间–效应关系(p<0.05)。2,在饲喂5d和10d后,不添加Cd饲喂的拟水狼蛛MDA含量和抗氧化酶系差异都不显著。添加Cd2 污染组(S1,S2和S3)MDA含量显著高于对照组(S4),MDA含量与饲喂时间呈显著正相关(p<0.05);GST、SOD和CAT等抗氧化酶活性污染组显著低于对照组,与饲喂时间呈显著负相关(p<0.05);饲喂20d后,污染组MDA含量和抗氧化酶（SOD和CAT）活性均与对照组无显著差异,但GST活性差异显著。     

Effects of a root-colonized dark septate endophyte on the glutathione metabolism in maize plants under cadmium stress

A high Cd-tolerant dark septate endophyte (DSE), Exophiala pisciphila, was inoculated into maize (Zea mays L.) roots under Cd stress. The Cd content, enzymes activity and thiol compound content relevant to glutathione (GSH) metabolism in maize leaves were analyzed. The Cd content in maize shoots increased with increasing Cd stress, but the DSE significantly reduced the Cd content at the 40?mg/kg Cd treatment. Cd stress increased the enzyme activity of glutathione reductase (GR), glutathione S-transferase (GST) and glutathione peroxidase (GSH-Px) as well as the thiol compound contents of sulfur, thiols (-SH) and oxidized glutathione (GSSG). The content of reduced GSH and the GSH/GSSG ratio reached a peak at the 5?mg/kg Cd treatment but then decreased with increasing Cd stress. Furthermore, the DSE significantly enhanced the GR and GSH-Px activity and increased the contents of -SH and GSH under low Cd stress (5 and 10?mg/kg), but decreased the γ-glutamylcysteine synthetase and GST activity under high Cd stress (20 and 40?mg/kg). Highly positive correlations between the Cd content with enzymes activity and enzymes activity with thiol compound content were observed. Results indicated that DSE played a role in activating GSH metabolism in maize leaves under Cd stress.     

Responses of Glutathione and Glutathione S-Transferase to Cadmium and Mercury Exposure in Pedunculate Oak (Quercus robur) Leaf Discs

Changes in chlorophyll, non-protein thiol and glutathione (GSH) levels, and the activity of glutathione S-transferase (GST) were investigated in cadmium(ll) and mercury(ll) cchloride treated leaf discs of mature pedunculate oak trees (Quercus robur). Both heavy metals caused decreases in chlorophyll content, but mercury was more toxic than cadmium. Cadmium treatments (30–250μiM) resulted in increasing non-protein thiol levels after 3d, but GSH contents decreased. Mercury (1–20μM) led to a concentration-dependent decline in both non-protein thiol and GSH levels. GST activities were not modified significantly by cadmium, but mercury treatments caused a dose- and time-dependent enzyme induction. Both the phytotoxic- and GST-inducing effect of mercury could be prevented by the cysteine precursor L-2-oxo-4-thiazolidinecarboxylic acid.     

Responses of two kidney bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis>) cultivars to the combined stress of sulfur deficiency and cadmium toxicity

Plants suffer from combined stress of sulfur deficiency and cadmium toxicity in some agricultural lands. However, little is known about the reaction in plants, such as responses in antioxidant enzymes and non-protein thiol compounds, to such combined stress. Therefore, in this study, four treatments, S-sufficiency (T_S?Cd), S-deficiency (T_?S?Cd), Cd stress (T_S+Cd) and combined stress of S-deficiency and Cd stress (T_?S+Cd), were set up to investigate (1) the effects of sulfur deficiency or sulfur sufficiency on Cd toxicity to kidney bean cultivar seedlings and the related mechanisms, and (2) the responses of two kidney bean cultivars to combined stress of S-deficiency and Cd-tolerance. The results showed significant increases in hydrogen peroxide (H₂O₂) and malondialdehyde contents and significant increases in antioxidant enzyme (superoxide dismutase, catalase, peroxidase, and glutathione S-transferase) activities and non-protein thiol compounds (non-protein thiols, reduced glutathione, phytochelatins) synthesis in the plants in T_S+Cd and T_?S+Cd. On the tissue level, higher proportion of Cd was found to be immobilized/deposited in roots, while on the sub-cell level, higher proportion of Cd was located in cell walls and vacuole fractions with lower in cell organelles. Taken together, the results indicated that Cd detoxification was achieved by the two kidney bean cultivars through antioxidant enzyme activation, non-protein thiol compound synthesis and sub-cellular compartmentalization. In addition, the results indicated that sufficient S supply helped to relieve Cd toxicity, which is of special significance for remediation or utilization of Cd-contaminated soils as S is a plant essential nutrient.     

Differential response of wheat roots to Cu, Ni and Cd treatment: oxidative stress and defense reactions

Wheat seedlings cv. Zyta were treated with Cu, Ni and Cd at the concentrations causing approximately 50 % root growth inhibition, i.e. 12.5, 50 and 60 μM, respectively. Tissue metal accumulation, membrane permeability, lipid peroxidation, protein oxidation, concentration of thiol compounds as well as protease, glutathione S-transferase (GST) and peroxidase (POD) activities were studied in roots after 7 days of metal exposure. The metals showed different concentrations in root tissues with Cu and Cd being accumulated to the smallest and to the greatest extent, respectively. Membrane permeability was significantly enhanced by Cu and Ni but not by Cd treatment. All metals induced similar increase in protein oxidation, while significant enhancement of lipid peroxidation was observed only in the case of Cu treatment. The detected thiol compounds: cysteine (Cys), homocysteine (Hcy), γ-glutamylcysteine (γ-GluCys) and glutathione (GSH) were differently influenced by the metal treatment. Ni appeared to be the most effective inductor of GSH accumulation while both Cu and Ni similarly increased Cys content in the roots. Accumulation of γ-GluCys was found in response to Cu and Cd applications. Concentration of Hcy was enhanced by Cd treatment but exposure to Ni decreased its content below the level of detection. The activity of GST was considerably elevated by Cd and Ni treatments, while POD activity was increased only in response to Cu application. Our study showed that wheat roots differently responded to treatment with metals used at the concentrations having similar impact on growth.     

Cadmium stress tolerance in crop plants: Probing the role of sulfur

Plants can''t move away and are therefore continuously confronted with unfavorable environmental conditions (such as soil salinity, drought, heat, cold, flooding and heavy metal contamination). Among heavy metals, cadmium (Cd) is a non-essential and toxic metal, rapidly taken up by roots and accumulated in various plant tissues which hamper the crop growth and productivity worldwide. Plants employ various strategies to counteract the inhibitory effect of Cd, among which nutrient management is one of a possible way to overcome Cd toxicity. Sulfur (S) uptake and assimilation are crucial for determining crop yield and resistance to Cd stress. Cd affects S assimilation pathway which leads to the activation of pathway responsible for the synthesis of cysteine (Cys), a precursor of glutathione (GSH) biosynthesis. GSH, a non-protein thiol acts as an important antioxidant in mitigating Cd-induced oxidative stress. It also plays an important role in phytochelatins (PCs) synthesis, which has a proven role in Cd detoxification. Therefore, S assimilation is considered a crucial step for plant survival under Cd stress. The aim of this review is to discuss the regulatory mechanism of S uptake and assimilation, GSH and PC synthesis for Cd stress tolerance in crop plants.Key words: cadmium, cysteine, glutathione, phytochelatins, stress tolerance, sulfur     

Stress tolerance in transgenic tobacco seedlings that overexpress glutathione S-transferase/glutathione peroxidase

Overexpression of a tobacco glutathione S-transferase with glutathione peroxidase activity (GST/GPX) in transgenic tobacco (Nicotiana tabacum L.) enhanced seedling growth under a variety of stressful conditions. In addition to increased GST and GPX activity, transgenic GST/GPX-expressing (GST+) seedlings had elevated levels of monodehydroascorbate reductase activity. GST+ seedlings also contained higher levels of glutathione and ascorbate than wild-type seedlings and the glutathione pools were more oxidized. Thermal or salt-stress treatments that inhibited the growth of wild-type seedlings also caused increased levels of lipid peroxidation. These treatments had less effect on the growth of GST+ seedling growth and did not lead to increased lipid peroxidation. Stress-induced damage resulted in reduced metabolic activity in wild-type seedlings while GST+ seedlings maintained metabolic activity levels comparable to seedlings grown under control conditions. These results indicate that overexpression of GST/GPX in transgenic tobacco seedlings provides increased glutathione-dependent peroxide scavenging and alterations in glutathione and ascorbate metabolism that lead to reduced oxidative damage. We conclude that this protective effect is primarily responsible for the ability of GST+ seedlings to maintain growth under stressful conditions.     

Postnuclear Supernatant: An In Vitro Model for Assessing Cadmium-Induced Neurotoxicity

Cadmium (Cd) is a toxic heavy metal commonly found in industrial workplaces, a food contaminant and a major constituent of cigarette smoke. Most of the organs are susceptible to Cd-induced toxicity, including brain. Postnuclear supernatant (PNS) has been accepted as an in vitro model for assessing xenobiotic induced toxicity. The goal of the present study was to validate PNS as an in vitro model for investigating the effect of Cd-induced neurotoxicity. Neurotoxic induction by Cd was established in a dose-dependent manner in PNS in vitro. Enzymatic and non-enzymatic antioxidants were used as biomarkers of exposure. Antioxidant enzymatic activity was measured as a significant increase in activities of catalase, superoxide dismutase, and glutathione S-transferase. On exposure to Cd, a significant increase in acetylcholinesterase and decrease in sodium-potassium ATPase activity was also observed. Non-enzymatic effect was also demonstrated as a significant elevation in reduced glutathione and non-protein thiol activity, but there was no significant increase or decrease in the concentrations of protein thiol. In accordance with the toxicity of Cd towards the studied brain structure, Cd-induced oxidative stress has been a focus of toxicological research as a possible mechanism of neurotoxicity. Our results suggest that PNS preparations can be used as a model for future investigation of xenobiotic-induced neurotoxicity under in vitro conditions.     

Engineering stress tolerance in transgenic plants

Research in our laboratory has focused on the analysis of the functions of a variety of enzymes that are involved in the scavenging of reactive oxygen intermediates (ROI) such as superoxide radicals (·O ₂ ⁻ ) and hydrogen peroxide (H₂O₂). Recent work has been on transgenic plants that over-express glutathione S-transferases (GST) that also have glutathione peroxidase activity. Transgenic tobacco plants that contain gene constructs that encode two different tobacco GST’s had elevated levels of both GST and GPX activity. Analysis of mature vegetative transgenic tobacco plants that over-express GST/GPX failed to show any increase in paraquat tolerance or protection from photooxidative stress. However, seeds of these GST/GPX-expressing tobacco lines are capable of more rapid germination and seedling growth at low temperatures and at elevated salt concentrations. Reduced levels of lipid peroxidation were noted in GST/GPX-expressing seedling compared to control seedlings under both stressful and non-stressful conditions. In addition, GST/GPX-expressing seedlings significantly accumulated more oxidized glutathione (GSSG) than control seedlings during stress. These characteristics clearly indicate that over-expression of GST/GPX in transgenic seedlings can have substantial effects on their stress tolerance. Furthermore, it appears that this effect is due primarily to the elevated levels of GPX activity.     

Induction of glutathione S-transferase activity and glutathione level in plants exposed to glyphosate

Treatment with the herbicide glyphosate led to significantly increased activities of the enzyme gluiathione S-transferase (GST, EC 2.5.1.18) in wheat ( Triticum aestivum L. cv. Kadett and cv. Satu), pea ( Pisum sativum L. ev. Debreceni Világoszöld) and in maize ( Zea mays. L. Pioneer 3839 hybrid) tissues. GST activities in wheat seedlings (cv. Kadett) exposed to 960 μM glyphosate for 4 days were ca 6-fold and 3-fold higher in shoots and roots, respectively, than in the controls. Glyphosate increased the GST activity to a lesser extent in pea and maize than in wheat. In wheat seedlings (cv. Satu) exposed let 120 μM glyphosate gradual increases in the content of non-protein thiols were observed. After 7 days exposure to glyphosate the thiol levels rose to about 360% and 220% of the controls in wheal shoots and roots, respectively. The elevation of thiol content in glyphosate-treated plants was shown to be primarily due increases of glutathione level. These results suggest that the enhanced glutathione metabolism may have a role in the mode of action or degradation of this herbicide.     

The effects of methylglyoxal on glutathione S-transferase from Nicotiana tabacum

Methylglyoxal (MG) is one of the aldehydes that accumulate in plants under environmental stress. Glutathione S-transferases (GSTs) play important roles, including detoxification, in the stress tolerance systems of plants. To determine the effects of MG, we characterized recombinant GST. MG decreased GST activity and thiol contents with increasing K(m). GST can serve as a target of MG modification, which is suppressed by application of reduced glutathione.