Glutathione in adaptation of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> to cadmium stress

The role of glutathione (GSH) in the adaptation of wild type Arabidopsis thaliana plants to Cd stress was investigated. The nutrient solution (control or containing 50 or 100 μM Cd) was supplemented with buthionine sulfoximine (BSO; 50, 100, 500 μM, to decrease the GSH content in plants) or GSH (50, 100, 500 μM, to increase its content in plants) in order to find how GSH content could regulate Cd stress responses. BSO application did not influence plant biomass, while exogenous GSH (especially 500 μM) reduced root biomass. BSO (500μM) in combination with Cd (100 μM) increased Cd toxicity on root growth (by over 50 %), most probably due to reduced GSH content and phytochelatin (PC) accumulation (by over 96 %). On the other hand, combination of exogenous GSH (500 μM) with Cd (100 μM) was also more toxic to plants than Cd alone despite a significant increase in GSH and PC accumulation (up to 2.7 fold in the roots). This fact could indicate that the natural content of endogenous GSH in wild type A. thaliana plants is sufficient for Cd-tolerance. A decrease in this GSH content led to decreased Cd-tolerance of the plants but an increase in GSH content did not enhance Cd-tolerance, and it showed even toxic effect on the plants.     

Influence of glutathione chemical effectors in the response of maize to arsenic exposure

To support the key role of glutathione (GSH) in the mechanisms of tolerance and accumulation of arsenic in plants, this work examines the impact of several effectors of GSH synthesis or action in the response of maize (Zea mays L.) to arsenic. Maize was exposed in hydroponics to iso-toxic rates of 150 μM arsenate or 75 μM arsenite for 9 days and GSH effectors, flurazole (an herbicide safener), l-buthionine-sulfoximine (BSO, a known inhibitor of GSH biosynthesis), and dimercaptosuccinate (DMS) and dimercaptopropanesulfonate (DMPS) (two thiols able to displace GSH from arsenite-GSH complexes) were assayed. The main responses of plants to arsenic exposure consisted of a biomass reduction (fresh weight basis) of about 50%, an increase of non-protein thiol (NPTs) levels (especially in the GSH precursor γ-glutamylcysteine and the phytochelatins PC? and PC?) in roots, with little effect in shoots, and an accumulation of between 600 and 1000 ppm of As (dry weight basis) in roots with very little translocation to shoots. Growth inhibition caused by arsenic was partially or completely reversed in plants co-treated with flurazole and arsenate or arsenite, respectively, highly exacerbated in plants co-treated with BSO, and not modified in plants co-treated with DMS or DMPS. These responses correlated well with an increase of both NPTs levels in roots and glutathione transferase activity in roots and shoots due to flurazole treatment, the decrease of NPTs levels in roots caused by BSO and the lack of effect on NPT levels caused by both DMS and DMPS. Regarding to arsenic accumulation in roots, it was not modified by flurazole, highly reduced by BSO, and increased between 2.5- and 4.0-fold by DMS and DMPS. Therefore, tolerance and accumulation of arsenic by maize could be manipulated pharmacologically by chemical effectors of GSH.     

Inhibition of glutathione synthesis reduces chilling tolerance in maize

 The role of glutathione (GSH) in protecting plants from chilling injury was analyzed in seedlings of a chilling-tolerant maize (Zea mays L.) genotype using buthionine sulfoximine (BSO), a specific inhibitor of γ-glutamylcysteine (γEC) synthetase, the first enzyme of GSH synthesis. At 25 °C, 1 mM BSO significantly increased cysteine and reduced GSH content and GSH reductase (GR: EC 1.6.4.2) activity, but interestingly affected neither fresh weight nor dry weight nor relative injury. Application of BSO up to 1 mM during chilling at 5 °C reduced the fresh and dry weights of shoots and roots and increased relative injury from 10 to almost 40%. Buthionine sulfoximine also induced a decrease in GR activity of 90 and 40% in roots and shoots, respectively. Addition of GSH or γEC together with BSO to the nutrient solution protected the seedlings from the BSO effect by increasing the levels of GSH and GR activity in roots and shoots. During chilling, the level of abscisic acid increased both in controls and BSO-treated seedlings and decreased after chilling in roots and shoots of the controls and in the roots of BSO-treated seedlings, but increased in their shoots. Taken together, our results show that BSO did not reduce chilling tolerance of the maize genotype analyzed by inhibiting abscisic acid accumulation but by establishing a low level of GSH, which also induced a decrease in GR activity. Received: 9 November 1999 / Accepted: 17 February 2000     

Cysteine does not repress adenosine-5′-phosphosulfate reductase through its conversion to either sulfate or glutathione

Cysteine (Cys) represses the activity of several key regulatory enzymes in the plant sulfate assimilatory pathway. However, it is not clear whether this effect arises from Cys itself or through its conversion to either sulfate or glutathione (GSH). Therefore, we examined this phenomenon by analyzing the activity of adenosine-5′-phosphosulfate (APS) reductase. Both APS reductase (AR) activity and mRNA levels were decreased by treatingArabidopsis thaliana roots with 1 mM Cys. The intracellular sulfate concentration was not affected, whereas enzymatic activity and, to some extent, the mRNA level, declined. Cys treatment in sulfur-starved plants also diminished both parameters. However, this response to Cys was more efficient than when plants were treated with an equal amount of sulfate. When Cys was removed from both Cys-and sulfate-fed plants, AR activity was recovered; the same removal of sulfate was not so effective. Moreover, buthionine sulfoximine (BSO), an inhibitor of GSH synthesis, did not influence the repression of AR by Cys. Finally the AR enzyme was inhibited by cysteinein vitro. These results indicate that Cys represses AR by inhibiting mRNA expression and by directly repressing enzymatic activity, rather than through its conversion to either sulfate or GSH.     

Overexpression of<Emphasis Type="Italic">Arabidopsis</Emphasis> phytochelatin synthase (<Emphasis Type="Italic">atpcsi</Emphasis>) does not change the maximum capacity for non-protein thiol production induced by Cadmium

Phytochelatins (PCs) play an important role in heavy-metal homeostasis and detoxification. However, we previously reported that the overexpression of PC synthase inArabidopsis does not lead to increased tolerance of cadmium but, rather, plants show higher Cd sensitivity. Here, we compared the maximum capacity for non-protein thiol (NPT) production at various concentrations of Cd in order to estimate PC synthesis indirectly for both transgenic (pcs9) and wild-type plants. The pcs9 line produced the highest level of NPT when treated with 200 p.M Cd for 3 d. In comparison, the maximum productivity by the wild type was in response to 500 μM Cd. Nevertheless, the absolute amounts of NPT produced did not differ significantly between those two genotypes. Furthermore, exogenous application of 1 mM GSH did not dramatically change the capacity for either pcs9 or wild-type plants. These results suggest that Cd hypersensitivity in the transgenic pcs9 may not be caused by supraoptimal intracellular concentrations of PC, but may, instead, be due to overexpressed PC synthase itself because that enzyme can bind metals. This action, therefore, may lead to some unknown disruption in cellular metal homeostasis under Cd stress.     

Phytochelatin synthesis and cadmium localization in wild type of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

AlthoughArabidopsis thaliana is known as a model plant, in molecular studies, as well as heavy metal tolerance of higher plants, there have been no detailed studies of its cadmium accumulation, tolerance and cellular distribution in a wild type of this species. In hydroponic experiments the wild type of A. thaliana (L.) Heynh cv. Columbia plants grew at cadmium concentrations varying from 5 to 100 M with phytotoxicity symptoms depending on the concentration and time of application. The concentration of cadmium in roots and shoots increased from 0.28 and 0.08 mg g^–1 d.wt at 5 M Cd treatment after 7 days to 0.82 and 0.85 mg g^–1 d.wt at 100 M Cd treatment after 14 days, respectively. Most of the cadmium (69–88% of its total pool) was found in shoot. Cd application induced the biosynthesis of phytochelatins (PCs) in root and shoot tissues. Studies with buthionine sulfoximine [BSO, specific inhibitor of glutathione (GSH) synthesis] supported the presence of Cd–phytochelatin complexes and their role in Cd detoxification and tolerance in wild type of A. thaliana. Cellular distribution of cadmium was examined using energy-dispersive X-ray micro-analysis. Particularly interesting was the observation of cadmium localized in the root pericycle.     

Glutathione and glutathione disulfide affect adventitious root formation and growth in tomato seedling cuttings

To study the relationship between glutathione and rooting, tomato seedling cuttings, grown on basal- or on auxin-supplemented media, were treated with the reduced (GSH) or oxidized (GSSG) form of this antioxidant. In turn, the consequences of the depletion of GSH pool on rooting were tested using l-buthionine sulfoximine (BSO), a specific inhibitor of GSH biosynthesis. Effects of the aforementioned treatments on rooting response were assessed. GSH treatment promoted root formation on cuttings grown on both basal- and auxin-supplemented media. Whereas GSSG did not affect the number of roots formed by cuttings grown on basal medium, it strongly enhanced the rooting stimulatory effect of auxin treatment. GSH depletion resulting from BSO application did not change the number of roots formed. All the tested compounds, namely GSH, GSSG, BSO and auxin, had a strong inhibitory effect on the elongation of regenerated roots. Supplementing the rooting medium with glutathione efficiently increased the GSH level in the rooting zones, while addition of BSO led to a strong decrease in endogenous GSH level. Neither of the treatments affected the level of GSSG. Exogenous auxin affect neither GSH nor GSSG levels in rooting zones; however, in the regenerated roots, GSH level was significantly higher when the organs were formed on auxin-supplemented medium. Patterns of GSH distribution in the roots regenerated on basal- and auxin-enriched media were studied using the GSH-specific dye monochlorobimane and confocal laser scanning microscopy. GSH was found in the root apical meristem and in the elongation zone. Auxin did not change the GSH distribution; however, the number of fluorescent cells was higher when roots were regenerated on auxin-supplemented medium.     

铜、镉胁迫下外源NO介导的番茄解毒途径

一氧化氮(NO)作为信号分子,在抵御重金属胁迫中起重要作用,但对不同离子胁迫下的解毒机制尚缺乏研究.本研究采用营养液培养法,研究了铜(Cu)、镉(Cd)单一或复合胁迫下,番茄幼苗对Cu、Cd的吸收转运特性及对外源NO的响应机制.结果表明: 50 μmol·L^-1的Cu²⁺、Cd²⁺均显著抑制番茄植株的生长,其中Cd胁迫对生长的抑制效应远高于Cu胁迫.Cu、Cd单一或复合胁迫均使番茄根系Cu、Cd含量显著升高,但根系对Cu、Cd吸收存在严格选择性.根细胞对必需元素Cu表现出“奢侈吸收”的现象,而对毒性较强的Cd则吸收相对较少,胞内Cd浓度仅为Cu的1/10左右.外源NO处理可不同程度地缓解Cu、Cd胁迫,其中缓解Cd胁迫的效能更强.番茄对被动进入细胞的Cu、Cd具有相似的解毒机制:一方面,Cu、Cd胁迫诱导细胞质中产生谷胱甘肽(GSH)、植物螯合肽(PCs)和金属硫蛋白(MTs),络合过多的Cu、Cd离子,降低其生物毒性;另一方面,过多的Cu、Cd离子或螯合物被转运至液泡区隔化.外源NO通过调控GSH-GSSG(氧化型谷胱甘肽)氧化还原状态及GSH-PCs代谢方向的改变,促进Cu、Cd离子转运至液泡区隔化来缓解胁迫抑制;NO还可诱导植株叶片或根系表达更多的金属硫蛋白、GSH和PCs,而且上述响应普遍存在叠加效应.这可能是NO介导番茄对Cu、Cd胁迫的另一主要解毒途径.     

Responses of non-protein thiols to Cd exposure in Cd hyperaccumulator Arabis paniculata Franch

We investigated the responses of phytochelatins (PCs), glutathione (GSH) and other non-protein thiols in Cd hyperaccumulator Arabis paniculata after Cd exposure. Applying γ-glutamylcysteine synthetase (γ-ECS) inhibitor, l-buthionine-sulfoximine (BSO), the roles of PCs in Cd tolerance and Cd accumulation in A. paniculata were evaluated. Plants were exposed to four Cd concentrations (0, 50, 100 and 250 μM) for different times (2w or 3w) with and without BSO. Overall, Cd exposure had little impact on plant biomass after 2w or 3w of growth except at the highest Cd level. A. paniculata tolerated ≤100 μM Cd with up to 1127 mg kg^?1 Cd in the shoots and 5624 mg kg^?1 Cd in the roots after 3w of Cd exposure. Cd exposure induced formation of PCs and three unknown thiols in the roots, but none were detected in the shoots. BSO had no significant effect on Cd sensitivity in plants though it reduced Cd accumulation in the roots. In addition, the molar ratio of PCs:Cd, which ranged from 0.7 to 1.3 after exposing to 50–100 μM Cd without BSO in the roots, was close to the value expected for PC-mediated Cd sequestration in plants. Those data indicate that GSH and PCs did not contribute to Cd tolerance in the shoots and Cd transport from the root to shoot in A. paniculata, but they may play an important role in Cd accumulation and Cd complexation in the roots of A. paniculata.     

Selective and Synergistic Activity of L-S,R-Buthionine Sulfoximine on Malignant Melanoma Is Accompanied by Decreased Expression of Glutathione-S-Transferase

L-buthionine-S,R-sulfoximine (BSO) selectively inhibits glutathione (GSH) synthesis. Malignant melanoma may be uniquely dependent on GSH and its linked enzymes, glutathione S-transferase (GST) and GSH-peroxidase, for metabolism of reactive orthoquinones and peroxides produced during melanin synthesis. We compared the in vitro effects of BSO on melanoma cell lines and fresh melanoma specimens (n = 118) with breast and ovarian cell lines and solid tumors (n = 244). IC₅₀ values (μM) for BSO on melanoma, breast and ovarian tumor specimens were 1.9, 8.6, and 29, respectively. The IC₉₀ for melanoma was 25.5 μM, a level 20-fold lower than steady state levels achieved clinically. The sensitivity of individual specimens of melanoma correlated with their melanin content (r = 0.63). BSO synergistically enhanced BCNU activity against melanoma cell lines and human tumors. We followed GSH levels, GST enzyme activity, GST isoenzyme profiles and mRNA levels after BSO. BSO (50 μM) treatment for 48 hr resulted in a 95% decrease in ZAZ and M14 melanoma cell line GSH levels, and a 60% decrease in GST enzyme activity. GST-μ. protein and mRNA levels were significantly reduced in both cell lines. GST expression was unaffected. These data suggest that BSO action on melanoma may be related to GSH depletion, diminishing the capacity to scavenge toxic metabolites produced during melanin synthesis. We report here for the first time that BSO enhancement of alkylator action may be related in part to down regulation of GST. BSO may be a clinically useful adjunct in the treatment of malignant melanoma.     

Response of Wild Type of Arabidopsis thaliana to Copper Stress

Wild type of Arabidopsis thaliana plants were cultivated hydroponically in Hoagland and Arnon nutrient solution and treated with copper (5 – 100 µM) for 2, 4, 7 and 14 d. A progressive decrease of the root length and biomass was observed at increasing Cu concentration in the nutrient solution. Roots accumulated higher amounts of Cu than shoots at all Cu treatments. Changes of cell and chloroplast ultrastructure of Cu-treated plants were also observed. Cu application did not induce formation of Cu-phytochelatin complexes. Changes in glutathione and glutathione disulfide content observed in roots and shoots of Cu-treated plants suggest their participation in amelioration of metal-induced oxidative stress.     

Altered differentiation in rat and rabbit limb bud micromass cultures by glutathione modulating agents

Glutathione (GSH) is the primary source of reducing equivalents in most cells, contributes significantly to the cellular redox potential and can control differentiation, proliferation, and apoptosis. Using limb bud micromass cultures from Sprague Dawley rats and New Zealand White rabbits, GSH modulating agents, L-buthionine-S,R-sulfoximine (BSO) and diethyl maleate (DEM) altered the formation of Alcian blue positive chondrogenic foci. Limb bud micromass cultures were treated for 5 d with BSO (50 or 100 μM) or DEM (5–25 μM). GSH content was determined by HPLC analysis. In rat cultures, BSO treatment did not affect differentiation but did show GSH depletion. In rabbit cultures, BSO completely inhibited differentiation and significantly depleted GSH. Treatment of rat cultures with DEM resulted in the dose-dependent decrease of chondrogenic foci, which correlated with a dose-dependent depletion of GSH. DEM completely inhibited rabbit limb bud cell differentiation and depleted GSH by 44%. Inhibition of differentiation was confirmed in rabbit cultures by the reduction in BMP-4 content. Addition of N-acetylcysteine to rabbit micromass cultures restored chondrogenic foci differentiation seen following treatment with both DEM and BSO. These results show species differences in GSH depletion in rat vs. rabbit limb bud cells and implicate GSH and cysteine in affecting pathways involved in chondrocyte differentiation.     

Thiols of Cu-treated maize plants inoculated with the arbuscular-mycorrhizal fungus Glomus intraradices

Mycorrhizal colonization of roots, fresh weight, content of cysteine, γ-glutamylcysteine (γEC). glutathione (GSH), thiol groups in Cu-binding peptides (CuBP), and the uptake of Cu were measured in roots and shoots of maize ( Zea mays L., cv. Honeycomb F-1) grown in quartz sand, with Cu at 0, 4.5, 9, 15 and 30 μg g⁻¹ added with or without inoculum of the arbuscular-mycorrhizal fungus (AMF) Glomus intraradices . In control plants (no Cu added) AMF significantly reduced shoot growth, but did not affect root growth. At an external Cu supply of 9 μg (g quartz sand)⁻¹ or higher, both mycorrhizal colonization and growth of roots and shoots of mycorrhizal and non-mycorrhizal plants were significantly reduced.
With up to 9 μg Cu g⁻¹, mycorrhizal colonization increased the content of cysteine, γEC and GSH in the roots. However, the amount of thiols in CuBPs was not increased by mycorrhizal colonization in Cu-treated plants and no differences in Cu uptake were detected between non-mycorrhizal and mycorrhizal plants. A CuBP-complex with a relative molecular mass of 7300 and a SH:Cu ratio of 1.77:1 was separated on a Sephadex G-50 column from both non-inoculated and inoculated roots of Cu-treated plants. HPLC chromatography of the CuBPs of both non-inoculated and inoculated roots resulted in a similar peak pattern, indicating that no additional CuBPs were formed by the fungus. In conclusion, our results do not support the idea that AMF protects maize from Cu-toxicity.     

外源NO对铜胁迫下番茄幼苗根系抗坏血酸谷胱甘肽循环的影响

采用营养液培养方法,研究外源NO对铜胁迫下番茄(Lycopersicon esculentum Mill.)幼苗根系抗坏血酸(AsA)-谷胱甘肽(GSH)循环中抗氧化物质和抗氧化酶系的影响.结果表明：外施适量NO(硝普钠)可提高铜胁迫下番茄幼苗根系AsA、GSH含量和AsA/DHA(氧化型抗坏血酸)、GSH/GSSG(氧化型谷胱甘肽),降低DHA和GSSG含量.添加100 μmol·L^-1 BSO(谷胱甘肽合成酶抑制剂)处理下,外源NO可提高铜胁迫下番茄幼苗根系的AsA含量、AsA/DHA及抗坏血酸酶(AAO)、单脱氢抗坏血酸还原酶(MDHAR)和脱氢抗坏血酸还原酶(DHAR)比活性,降低DHA、GSH、GSSG含量及抗坏血酸过氧化物酶(APX)、谷胱甘肽还原酶(GR)比活性;添加250 μmol·L^-1 BSO处理下,外源NO提高了铜胁迫下番茄幼苗根系的AsA、GSH、GSSG含量、AsA/DHA及APX和GR比活性,降低了DHA含量及AAO、DHAR和MDHAR比活性.说明外源NO影响了铜胁迫下番茄根系的AsA-GSH代谢循环,并通过调节AsA/DHA、GSH/GSSG的变化来减轻氧化胁迫,从而缓解铜胁迫对番茄根系的伤害.     

Differential accumulation of soluble proteins in roots of metallicolous and nonmetallicolous populations of Agrostis capillaris L. exposed to Cu

Differential expression of soluble proteins was explored in roots of metallicolous (M) and non‐M (NM) plants of Agrostis capillaris L. exposed to increasing Cu to partially identify molecular mechanisms underlying higher Cu tolerance in M plants. Plants were cultivated for 2 months on perlite with a CuSO₄ (1–30 μM) spiked‐nutrient solution. Soluble proteins extracted by the trichloroacetic acid/acetone procedure were separated with 2DE (linear 4–7 pH gradient). After Coomassie Blue staining and image analysis, 19 proteins differentially expressed were identified using LC‐MS/MS and Expressed Sequence Tag (ESTs) databases. At supra‐optimal Cu exposure (15–30 μM), glycolysis was likely altered in NM roots with increased production of glycerone‐P and methylglyoxal based on overexpression of triosephosphate isomerase and fructose bisphosphate aldolase. Changes in tubulins and higher expressions of 5‐methyltetrahydropteroyltriglutamatehomocysteine methyltransferase and S‐adenosylmethionine synthase underpinned impacts on the cytoskeleton and stimulation of ethylene metabolism. Increased l ‐methionine and S‐adenosylmethionine amounts may also facilitate production of nicotianamine, which complexes Cu, and of l ‐cysteine, needed for metallothioneins and GSH. In M roots, the increase of [Cu/Zn] superoxide dismutase suggested a better detoxification of superoxide, when Cu exposure rose. Higher Cu‐tolerance of M plants would rather result from simultaneous cooperation of various processes than from a specific mechanism.     

Adaptive Copper Tolerance in <Emphasis Type="Italic">Elsholtzia haichowensis</Emphasis> Involves Production of Cu-induced Thiol Peptides

Copper (Cu) accumulation and tolerance mechanisms in Elsholtzia haichowensis, an indicator plant of Cu mines, were investigated under hydroponics supplied with different concentrations (0.32, 50.0, 100.0 and 200.0 μM) of Cu for 8 days. Cu at 100 and 200 μM significantly decreased the root dry weight, but had no significant effect on shoot dry weight. The plants grown in the presence of 200 μM Cu accumulated 288 and 7626 μg g⁻¹ DW total Cu in the shoots and roots, respectively. A greater proportion of accumulated Cu was water-soluble accounting for 42–93% of the total Cu content in the shoots. The concentrations of reduced glutathione (GSH) and protein thiols were significantly enhanced under excess Cu supply. However, the concentrations of these compounds, particularly protein thiols, were much higher in the leaves than that in the roots. Three UV-absorbing peaks could be eluted out through gel filtration chromatography on Sephadex G-50. A large amount of Cu was detected in the UV-absorbing peaks in 40–50 and 70–90 ml elution fractions of the root extract, and in 40–50 and 120–140 ml elution fractions of the leaf extract. The results suggested that the adaptive Cu tolerance mechanism in E. haichowensis might involve the active participation of protein thiols which had a more important role in the leaves than in the roots.     

Physiological responses of <Emphasis Type="Italic">Lupinus luteus</Emphasis> to different copper concentrations

Yellow lupin (Lupinus luteus L.) plants were grown in hydroponic solution for 15 d under different copper concentrations (0.1, 0.5, 1.0, 10, 25 and 50 μM). With increasing Cu concentration total biomass was not affected, leaf area slightly decreased, while chlorophyll content decreased considerably. Cu content increased significantly both in roots and in leaves, but the contents of other ions were only slightly affected at the highest Cu concentration (Mn content decreased both in roots and in leaves, P content decreased only in leaves and Zn content increased in roots). Superoxide dismutase (SOD) activity increased up to day 7 after copper application. Peroxidase (GPOD) and polyphenol oxidase (PPO) activities also increased, while catalase (CAT) activity remained constant.     

Interactive effects of zinc and nickel on the glutathione system state in <Emphasis Type="Italic">Mimulus guttatus</Emphasis> plants

To determine whether the enhanced stress tolerance of ZnSO₄ with NiSO₄-treated Mimulus guttatus Fischer ex DC. plants was associated with the glutathione (GR-GSH) system, we investigated the changes in glutathione redox state (reduced (GSH), oxidized (GSSG) forms, total reduced (GSHt) glutathione, and GSH/GSSG ratio) and in the enzymatic activities of glutathione reductase (GR) and peroxidatic glutathione S-transferases (GST). The 6-week-old plants were grown in water culture during 4 weeks on a modified Rorison’s medium with ZnSO₄ (50, 100, and 200 μM) and NiSO₄ (20 and 80 μM) in a condition of separate or simultaneous supply of the components. Dry biomass accumulations of roots and shoots were not influenced by the examined treatments. The positive correlations between the total external concentrations of ZnSO₄ and NiSO₄ and the total Zn and Ni contents in roots and leaves were found. It was determined that the MDA content was higher in the ZnSO₄-treated plants than in the NiSO₄-treated ones. The supplementation of the ZnSO₄-treated plants with varied concentrations of NiSO₄ decreased the Zn-induced increase in the MDA levels. The inverse proportionality between the MDA and pigment levels in leaves was found. The Zn-Ni interactions were shown to induce the decreases in the GR activity, the total peroxidatic GST activity, and the GSH/GSSG ratio in roots. However, in leaves, the GR activity and the GSH/GSSG ratio were significantly increased and the total peroxidatic GST activity was decreased. The supplementation of the ZnSO₄-treated plants with varied concentrations of NiSO₄ restored the Zn-induced reduction in the GSHt levels in roots and decreased the Zn-induced increase in the GSSG levels in leaves, which resulted in more reduced state of the intracellular environment. It was likely to cause a decrease of the MDA level. Thus, our studies on the Zn?Ni interactions identified the antagonizing role of Ni in Zn toxicity by the GR-GSH system.