Ascorbate GSH Research Articles

SpHsfA4c from Sedum plumbizincicola Enhances Cd Tolerance by the AsA–GSH Pathway in Transgenic Populus × canescens

The ascorbate (AsA)–glutathione (GSH) metabolism pathway is an important antioxidant system in cadmium (Cd) detoxification; the AsA–GSHpathway is generally regulated by a specific set of functional genes. However, transcription factors involved in AsA–GSH pathway have yet to be identified. Herein, we transformed a heat shock transcription factor SpHsfA4c from Sedum plumbizincicola into Populus. × canescens. Under 100 μM CdCl2 stress for 30 d, the leaf chlorosis of wild-type poplars (WT) is more serious than that in transgenic poplars. The root biomass, shoot biomass and tolerance index (TIs) of transgenic poplars were higher than those in WT. In addition, transgenic poplars have higher Cd2+ uptake and Cd content. Compared with WT, the contents of hydrogen peroxide (H2O2) and superoxide anion (O2•−) in transgenic poplars were significantly reduced in leaves under Cd treatment. The expression levels of five enzymes (ascorbate peroxidase (APX), catalases (CAT), superoxide dismutase (SOD), peroxidase (POD) and glutathione S-transferase (GST)) were higher in transgenic poplars than those in WT. Transgenic poplars contained higher concentrations of intermediate metabolites, including GSH, AsA and phytochelatins (PCs), and a higher GSH/GSSG ratio in the AsA–GSH metabolism pathway. In Fourier transform infrared (FTIR) spectra, the characteristic peaks indicated that the contents of cysteine, GSH and AsA in transgenic poplars were exceeded compared to those in WT. These results suggested that SpHsfA4c can activate the AsA–GSH metabolism pathway to reduce Cd-associated oxidative stress. Therefore, overexpressing SpHsfA4c in P. × canescens can give rise to a superior Cd tolerance. Our results provide a theoretical significance for breeding potential new germplasm resources with high biomass and high Cd tolerance for remediation of soil heavy metal pollution.

Comprehensive physiological, transcriptomic, and metabolomic analyses reveal the synergistic mechanism of Bacillus pumilus G5 combined with silicon alleviate oxidative stress in drought-stressed Glycyrrhiza uralensis Fisch.

Glycyrrhiza uralensis Fisch. is often cultivated in arid, semi-arid, and salt-affected regions that suffer from drought stress, which leads to the accumulation of reactive oxygen species (ROS), thus causing oxidative stress. Plant growth-promoting bacteria (PGPB) and silicon (Si) have been widely reported to be beneficial in improving the tolerance of plants to drought stress by maintaining plant ROS homeostasis. Herein, combining physiological, transcriptomic, and metabolomic analyses, we investigated the response of the antioxidant system of G. uralensis seedlings under drought stress to Bacillus pumilus (G5) and/or Si treatment. The results showed that drought stress caused the overproduction of ROS, accompanied by the low efficiency of antioxidants [i.e., superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), the ascorbate (AsA)-glutathione (GSH) pool, total carotenoids, and total flavonoids]. Inversely, supplementation with G5 and/or Si enhanced the antioxidant defense system in drought-stressed G. uralensis seedlings, and the complex regulation of the combination of G5 and Si differed from that of G5 or Si alone. The combination of G5 and Si enhanced the antioxidant enzyme system, accelerated the AsA-GSH cycle, and triggered the carotenoid and flavonoid metabolism, which acted in combination via different pathways to eliminate the excess ROS induced by drought stress, thereby alleviating oxidative stress. These findings provide new insights into the comparative and synergistic roles of PGPB and Si in the antioxidant system of plants exposed to drought and a guide for the application of PGPB combined with Si to modulate the tolerance of plants to stress.

Insight into the Mechanism of Salt-Induced Oxidative Stress Tolerance in Soybean by the Application of Bacillus subtilis: Coordinated Actions of Osmoregulation, Ion Homeostasis, Antioxidant Defense, and Methylglyoxal Detoxification.

Considering the growth-promoting potential and other regulatory roles of bacteria, we investigated the possible mechanism of the role of Bacillus subtilis in conferring salt tolerance in soybean. Soybean (Glycine max cv. BARI Soybean-5) seeds were inoculated with B. subtilis, either through a presoaking with seeds or a direct application with pot soil. After 20 days of sowing, both the seed- and soil-inoculated plants were exposed to 50, 100, and 150 mM of NaCl for 30 days. A clear sign of oxidative stress was evident through a remarkable increase in lipid peroxidation, hydrogen peroxide, methylglyoxal, and electrolyte leakage in the salt treated plants. Moreover, the efficiency of the ascorbate (AsA)–glutathione (GSH) pathways was declined. Consequently, the plant growth, biomass accumulation, water relations, and content of the photosynthetic pigments were decreased. Salt stress also caused an increased Na+/K+ ratio and decreased Ca2+. On the contrary, the B. subtilis inoculated plants showed increased levels of AsA and GSH, their redox balance, and the activities of the AsA–GSH pathway enzymes, superoxide dismutase, catalase, glutathione peroxidase, glutathione S-transferase, and peroxidase. The B. subtilis inoculated plants also enhanced the activities of glyoxalase enzymes, which mitigated methylglyoxal toxicity in coordination with ROS homeostasis. Besides this, the accumulation of K+ and Ca2+ was increased to maintain the ion homeostasis in the B. subtilis inoculated plants under salinity. Furthermore, the plant water status was uplifted in the salt treated soybean plants with B. subtilis inoculation. This investigation reveals the potential of B. subtilis in mitigating salt-induced oxidative stress in soybean plants through modulating the antioxidant defense and glyoxalase systems along with maintaining ion homeostasis and osmotic adjustments. In addition, it was evident that the soil inoculation performed better than the seed inoculation in mitigating salt-induced oxidative damages in soybean.

Determination of some physiological parameters in spinach (Spinacia oleracea L.) cultivated in Kosovo

Summary The objective of this work was to investigate the levels of photosynthetic pigments, antioxidant enzyme activities (superoxide dismutase [SOD], catalase [CAT], peroxidase [POX], ascorbate peroxidase [APX], glutathione reductase [GR], and MDA), total glutathione (GSH), total ascorbate (AsA), and lipid peroxidation of two spinach cultivars (Clipper and Matador) sampled in different locations in Kosovo (Prizren, Ferizaj, and Obiliq). The photosynthetic pigments, chlorophyll a and b as well as carotenoids, were extracted by 80% acetone, and concentrations were calculated based on the absorbance values at 663, 645, and 480 nm. There was a significant difference in SOD, APX, and GR activities between varieties at each location. The MDA and AsA contents in Obiliq were the highest, compared to other locations. From the increase in lipid peroxidation level, it can be assumed that the degradation in pigment content was caused by the decrease in total AsA content and APX and GR enzyme activities, which are the main enzymes of the AsA–GSH cycle in chloroplasts.

A Computational Study of the Role of Secondary Metabolites for Mitigation of Acid Soil Stress in Cereals Using Dehydroascorbate and Mono-Dehydroascorbate Reductases

The present study investigates the potential ameliorative role of seven secondary metabolites, viz., ascorbate (AsA), reduced glutathione (GSH), jasmonic acid (JA), salicylic acid (SA), serotonin (5-HT), indole–3–acetic acid (IAA) and gibberellic acid (GA3), for mitigation of aluminium (Al3+) and manganese (Mn2+) stress associated with acidic soils in rice, maize and wheat. The dehydroascorbate reductase (DHAR) and mono-dehydroascorbate reductase (MDHAR) of the cereals were used as model targets, and the analysis was performed using computational tools. Molecular docking approach was employed to evaluate the interaction of these ions (Al3+ and Mn2+) and the metabolites at the active sites of the two target enzymes. The results indicate that the ions potentially interact with the active sites of these enzymes and conceivably influence the AsA–GSH cycle. The metabolites showed strong interactions at the active sites of the enzymes. When the electrostatic surfaces of the metabolites and the ions were generated, it revealed that the surfaces overlap in the case of DHAR of rice and wheat, and MDHAR of rice. Thus, it was hypothesized that the metabolites may prevent the interaction of ions with the enzymes. This is an interesting approach to decipher the mechanism of action of secondary metabolites against the metal or metalloid - induced stress responses in cereals by aiming at specific targets. The findings of the present study are reasonably significant and may be the beginning of an interesting and useful approach towards comprehending the role of secondary metabolites for stress amelioration and mitigation in cereals grown under acidic soil conditions.

Ascorbate-Mediated Modulation of Cadmium Stress Responses: Reactive Oxygen Species and Redox Status in Brassica napus.

The role of ascorbate (AsA) in antioxidant defense system-associated resistance to cadmium (Cd) in oilseed rape plants has not yet been clearly demonstrated. The present study investigated the critical role of exogenous AsA on the physiological and biochemical responses of reactive oxygen species (ROS) and antioxidant scavenging defense systems in oilseed rape (Brassica napus L. cv. Tammi) seedlings exposed to Cd. Cd (10 μM) treatment led to significant reductions in plant growth; increases in the levels of superoxide anion radical, hydrogen peroxide, and malondialdehyde; and increases in Cd uptake and accumulation by the roots and shoots in hydroponically grown 10-day-old seedlings. Moreover, it reduced AsA content and AsA redox ratios, which have been correlated with reductions in glutathione (GSH) and/or nicotinamide adenine dinucleotide phosphate (NADPH) redox status. However, exogenously applying AsA to Cd-exposed seedlings decreased Cd-induced ROS, improved antioxidant defense systems by increasing AsA, GSH, and NADPH contents, and increased Cd uptake and accumulation in both roots and shoots of the plants. These results provided evidence that the enhancement in AsA redox status can be linked to an increase in the GSH and/or NADPH redox ratios through the induction of the AsA–GSH–NADPH cycle. Thus, these results suggest that exogenous AsA application to oilseed rape seedlings under Cd stress might alleviate the overall Cd toxicity by regulating the homeostasis of the AsA–GSH–NADPH cycle, which reestablishes the steady-state cellular redox status.

Exogenous Glutathione Increases Arsenic Translocation Into Shoots and Alleviates Arsenic-Induced Oxidative Stress by Sustaining Ascorbate-Glutathione Homeostasis in Rice Seedlings.

Glutathione (GSH) plays diverse roles in the physiological processes, stress defense, growth, and development of plants. This study investigated the effects of exogenous GSH on the biochemical responses of reactive oxygen species and antioxidant levels in rice (Oryza sativa L. cv. Dasan) seedlings under arsenic (As) stress. As treatment inhibited growth; increased the level of superoxide, hydrogen peroxide, and malondialdehyde; and enhanced the uptake of As by the roots and shoots in hydroponically grown 14-day-old seedlings. Furthermore, it reduced GSH content and GSH redox ratios, which have been correlated with the decrease in ascorbate (AsA) redox state. Whereas the exogenous application of GSH in As-treated seedlings reduced As-induced oxidative stress, improved antioxidant defense systems by maintaining antioxidant and/or redox enzyme homeostasis, and increased the AsA and GSH contents, the GSH application also increased the As translocation from the roots to the shoots. These results indicated that the increase in GSH redox state can be linked to an increase in the AsA redox ratio via the induction of the AsA–GSH cycle. Therefore, the results suggest that exogenous GSH application should be a promising approach to enhance As stress resistance in rice plants.

Exogenous Melatonin Counteracts NaCl-Induced Damage by Regulating the Antioxidant System, Proline and Carbohydrates Metabolism in Tomato Seedlings.

Melatonin, a natural agent, has multiple functions in animals as well as in plants. However, its possible roles in plants under abiotic stress are not clear. Nowadays, soil salinity is a major threat to global agriculture because a high soil salt content causes multiple stresses (hyperosmotic, ionic, and oxidative). Therefore, the aim of the present study was to explore: (1) the involvement of melatonin in biosynthesis of photosynthetic pigments and in regulation of photosynthetic enzymes, such as carbonic anhydrase (CA) and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco); (2) the role of melatonin in osmoregulation by proline and carbohydrate metabolism; and (3) the function of melatonin in the antioxidant defense system under salinity. Outcomes of the study reveal that under non-saline conditions, application of melatonin (20 and 50 µM) improved plant growth, viz. shoot length, root length, shoot fresh weight (FW), root FW, shoot dry weight (DW), root DW and leaf area and physio-biochemical parameters [chlorophyll (Chl) a and b, proline (Pro) and total soluble carbohydrates (TSC) content, and increased the activity of CA and Rubisco]. However, tomato seedlings treated with NaCl exhibited enhanced Chl degradation, electrolyte leakage (EL), malondialdehyde (MDA) and reactive oxygen species (ROS; superoxide and hydrogen peroxide). ROS were detected in leaf and root. Interestingly, application of melatonin improved plant growth and reduced EL, MDA and ROS levels through upregulation of photosynthesis enzymes (CA, Rubisco), antioxidant enzymes (superoxide dismutase, catalase, glutathione reductase and ascorbate reductase) and levels of non-enzymatic antioxidants [ascorbate (ASC) and reduced glutathione (GSH)], as well as by affecting the ASC—GSH cycle. Additionally, exogenous melatonin also improved osmoregulation by increasing the content of TSC, Pro and Δ1-pyrroline-5-carboxylate synthetase activity. These results suggest that melatonin has beneficial effects on tomato seedlings growth under both stress and non-stress conditions. Melatonin’s role in tolerance to salt stress may be associated with the regulation of enzymes involved in photosynthesis, the antioxidant system, metabolism of proline and carbohydrate, and the ASC—GSH cycle. Also, melatonin could be responsible for maintaining the high ratios of GSH/GSSG and ASC/DHA.

Cytosolic monodehydroascorbate reductase gene affects stress adaptation and grain yield under paddy field conditions in Oryza sativa L. japonica

Monodehydroascorbate reductase (MDHAR), which is responsible for growth, development and stress response in plants, is a key enzyme in the maintenance of the ascorbate (AsA) pool through the AsA–glutathione (AsA–GSH) cycle and is induced by abiotic stresses. It has highly conserved regions containing FAD- and NAD(P)H-binding domains. In particular, NAD(P)H is a significant electron donor in the AsA–GSH pathway. In this context, we introduced RNA interference (RNAi) to determine the functional role of Oryza sativa L. japonica MDHAR isoform 3 (OsMDHAR3) and developed transgenic (mdhar3) rice plants in which the NAD(P)H domain was silenced. The mdhar3 rice plants were more sensitive to salt stress than the wild-type (WT) plants. In addition, the mdhar3 rice plants showed decreased ability for environmental adaptation because of an imbalance in the redox homeostasis and reduced AsA pool. These plants showed increased hydroperoxide levels and ion leakage, and decreased chlorophyll content and ascorbate/dehydroascorbate ratio under the paddy field conditions; they also exhibited a reduction in the total biomass and grain yield. Furthermore, the activity of a purified E196A mutant of the OsMDHAR protein decreased to approximately 70% of the activity of the WT protein. These results suggest that OsMDHAR3 plays a critical role in the intrinsic resistance, as well as in the sensitivity of seed maturation and productivity, of rice plants to environmental stresses, thereby indicating the functional importance of NADH in MDHAR activity, in vivo and in vitro.

Lanthanum improves the cadmium tolerance of Zea mays seedlings by the regulation of ascorbate and glutathione metabolism

The effect of lanthanum on the metabolism of ascorbate (AsA) and glutathione (GSH) in the leaves of maize seedlings under cadmium stress was investigated. The findings showed that Cd remarkably increased electrolyte leakage (EL), the activities of ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase (MDHAR), glutathione reductase, L-galactono-1,4-lactone dehydrogenase, and γ-glutamylcysteine synthetase, and the content of reduced AsA, reduced GSH, total AsA, total GSH, malondialdehyde (MDA), and Cd, compared with control. However, Cd significantly decreased the dry biomass of roots and shoots. Treatment with La + Cd evidently increased the activities of above enzymes except MDHAR, the content of reduced AsA, reduced GSH, total AsA and total GSH, and the dry biomass of roots and shoots, compared with Cd stress alone. Meanwhile, treatment with La + Cd remarkably decreased EL and the content of Cd and MDA compared with Cd stress alone. Our results suggested that La could be used as a regulator to improve the Cd tolerance of maize for its role in the alleviation of Cd-induced oxidative damage by regulating the metabolism of AsA and GSH.

Triacontanol Reduces Transplanting Shock in Machine-Transplanted Rice by Improving the Growth and Antioxidant Systems.

Machine transplantation results in serious transplant shock in seedlings and results in a longer recover stage, which negatively impacts the growth of low-position tillers and the yield of machine-transplanted rice. A barrel experiment was conducted to examine the effect of the foliar application of triacontanol (TRIA) on machine-transplanted rice during the recovery stage. TRIA (0, 1, 5, and 10 μM) was sprayed over leaves 2 days before transplanting. The chlorophyll content, sucrose content, oxidative damage, antioxidant enzyme levels, glutathione (GSH), and ascorbate (ASA) redox states, tiller dynamics and yield components of the plants were investigated. The results show that foliar-applied TRIA significantly alleviates the growth inhibition and oxidative damage caused by transplant shock. Furthermore, the application of TRIA increased the chlorophyll and sucrose contents of the plants. Importantly, TRIA not only significantly improved the activity of catalase (CAT) and guaiacol peroxidase (POD), demonstrating that POD can play an important role in scavenging H2O2 during the recovery stage, but it also enhanced the redox states of ASA and GSH by regulating the activities of enzymes involved in the ASA–GSH cycle, such as ascorbate peroxidase (APX) and glutathione reductase (GR). A dose of 10 μM TRIA was the most efficient in reducing the negative effects of transplant shock, increasing the panicles, grain filling, and grain yield per hill by 17.80, 5.86, and 16.49%, respectively. These results suggest that TRIA acts to reduce transplant shock in association with the regulation of the redox states of ASA and GSH and antioxidant enzymes and serves as an effective antioxidant to maintain photosynthetic capacity and promote the occurrence of low tillers.

Gibberellin-like effects of KAR1 on dormancy release of Avena fatua caryopses include participation of non-enzymatic antioxidants and cell cycle activation in embryos.

Main conclusionThe induction of dormancy release and germination ofAvena fatuacaryopses by KAR1involves ABA degradation to phaseic acid. Both, KAR1and GA3, control the AsA–GSH cycle, DNA replication and accumulation of β-tubulin in embryos before caryopses germination.Avena fatua caryopses cannot germinate in darkness at 20 °C because of dormancy, but karrikinolide-1 (KAR1), a compound in plant-derived smoke, and gibberellic acid (GA3) induced an almost complete germination. The radicle protrusion through the coleorhiza was preceded by increased water uptake, rupture of coat, increased embryo size and coleorhiza length as well as coleorhiza protrusion through covering structures. The stimulatory effect of KAR1 was correlated with the reduced content of abscisic acid (ABA) and an increase in phaseic acid (PA) in embryos from caryopses before coleorhiza protrusion. Two non-enzymatic antioxidants, ascorbate (AsA) and reduced glutathione (GSH), did not affect the germination of dormant caryopses, but in the presence of KAR1 or GA3 they only slightly delayed the germination. The stimulatory effect of KAR1 or GA3 on the final germination percentage was markedly antagonized by lycorine, an AsA biosynthesis inhibitor. KAR1 and GA3 applied during caryopses imbibition resulted in increases of AsA, dehydroascorbate (DHA) and GSH, but reduced the embryos’ oxidized glutathione (GSSG) content. Furthermore, both KAR1 and GA3 induced an additional ascorbate peroxidase (APX) isoenzyme and increased the glutathione reductase (GR) activity. Both compounds stimulated β-tubulin accumulation in radicle+coleorhiza (RC) and plumule+coleoptile (PC), and enhanced the transition from G1 to S and also from S to G2 phases. The comparison of the effects produced by KAR1 and GA3 shows a similar action; thus the KAR1 effect may not be specific. The study provides new data regarding the mechanism with which KAR1, a representative of a novel class of plant growth regulators, regulates dormancy and germination of caryopses.

Sulfur decreases cadmium translocation and enhances cadmium tolerance by promoting sulfur assimilation and glutathione metabolism in Brassica chinensis L.

We investigated the ameliorative role of sulfur (S) in protecting plants against cadmium (Cd) toxicity by using two pakchoi (Brassica chinensis L.) cultivars with different Cd tolerance levels. The exposure of pakchoi seedlings to 100μM Cd inhibited plant growth, increased superoxide content, enhanced membrane lipid peroxidation, and induced Cd accumulation in the roots and shoots. Application of S to Cd-stressed plants alleviated Cd-induced oxidative stress by promoting the capacity of the ascorbate (AsA)–glutathione (GSH) cycle, enhanced S assimilation by increasing the activity of ATP sulfurylase (ATPS) and o-acetylserine(thiol)lyase (OASTL), and decreased Cd translocation from the roots to the shoots by enhancing phytochelatins (PCs) biosynthesis. Results suggested that S reversed Cd-induced growth inhibition and oxidative stress by restraining Cd translocation from the roots to the shoots and upregulating S assimilation and GSH metabolism, including the AsA–GSH cycle and PCs synthesis.

Silicon Application Alleviates Drought Stress in Wheat Through Transcriptional Regulation of Multiple Antioxidant Defense Pathways

Silicon (Si) has been shown to increase plant stress tolerance. However, the molecular mechanisms underlying the effects of Si in alleviating drought stress in winter wheat are unknown. The aim of this study was to investigate the effects of Si on photosynthetic pigments, antioxidant contents, and transcription of several genes involved in the antioxidant defense pathway in drought-stressed wheat plants. Si application resulted in higher leaf chlorophyll content, and lower lipid peroxidation levels and H2O2 contents in wheat flag leaves compared with untreated plants. In addition, Si increased the contents of ascorbate (ASC), reduced glutathione (GSH), and the total phenolic and total flavonoid contents in wheat flag leaves. Quantitative real-time PCR was used to analyze gene expression patterns for three antioxidant enzyme genes, four ASC–GSH cycle genes, and five flavonoid biosynthesis pathway genes. Si application increased the expression of these genes in drought-stressed wheat compared with the −Si treatments. Elevated levels of gene expression and low malondialdehyde and H2O2 contents indicate that Si plays a pivotal role in the coordinated transcriptional regulation of multiple antioxidant defenses in response to drought. Our work also suggests that Si may be important in the metabolic or physiological activities for growth and development of winter wheat.

Biofortification with potassium: antioxidant responses during postharvest of cherry tomato fruits in cold storage

Tomato fruits are sensitive to storage at low temperatures after harvest. Under these conditions, the main mechanism induced in fruits is oxidative stress, which can translate as lipid peroxidation and in turn deteriorate fruit quality. The aim of the present work was to investigate whether the effect of a biofortification program with potassium (K) improves the postharvest storage of cherry tomato fruits at 4 °C, through a better antioxidant response. Three K treatments were applied during the crop cycle of the plants: 5, 10, and 15 mM of KCl. The parameters in fruits on the day of harvest and after 21 days of postharvest cold storage at 4 °C, such as activity of lipoxygenase, malondialdehyde, catalase, superoxide dismutase, and the enzymes involved in the AsA–GSH cycle as well as the forms of ascorbate (AsA) and glutathione (GSH), were analyzed. The tomato fruits harvested from plants treated with 15 mM of KCl after 21 days of postharvest at 4 °C showed a lower degree of lipid peroxidation, an effective regeneration of AsA, and the highest pool of this compound in comparison with the other treatments. This response was because it presented the highest ascorbate peroxidase and monodehydroascorbate reductase activity. In addition, the treatments of 10 and 15 mM KCl presented the highest GSH pool, as well as a satisfactory regeneration of this tripeptide. All these results lead to the conclusion that the rate of 15 mM of KCl applied to this tomato variety (Solanum lycopersicum L. cv AsHiari grafted on cv. Maxifort rootstock) is adequate to mitigate the negative effects of postharvest chilling.

Salicylic acid increases the contents of glutathione and ascorbate and temporally regulates the related gene expression in salt-stressed wheat seedlings

Exogenous salicylic acid (SA) significantly improved abiotic tolerance in higher plants, and ascorbate (ASA) and glutathione (GSH) play important roles in abiotic tolerance. In this study, SA (0.5mM) markedly increased the contents of ASA and GSH in SA-treated plants during salt stress (250mM NaCl). The transcript levels of the genes encoding ASA and GSH cycle enzymes were measured using quantitative real-time PCR. The results indicated that, during salt stress, exogenous SA significantly enhanced the transcripts of glutathione peroxidase (GPX1), phospholipid hydroperoxide glutathione peroxidase (GPX2) and dehydroascorbate reductase (DHAR) genes at 12h, glutathione reductase (GR) at 24h, 48h and 72h, glutathione-S-transferase 1 (GST1), 2 (GST2), monodehydroascorbate reductase (MDHAR) and glutathione synthetase (GS) at the 48h and 72h after salt stress, respectively. The results implied that SA temporally regulated the transcript levels of the genes encoding ASA–GSH cycle enzymes, resulting in the increased contents of GSH and ASA and enhanced salt tolerance.

24-Epibrassinolide maintains elevated redox state of AsA and GSH in radish (Raphanus sativus L.) seedlings under zinc stress

Exogenous-applied 24-epibrassinolide (EBR) increased the seedling growth of radish (Raphanus sativus L.) in terms of seedling length, fresh weight and dry weight both in zinc (Zn2+)-stressed and unstressed conditions. Moreover, EBR lowered the Zn2+ uptake and bioaccumulation. Increased oxidation of ascorbate (AsA) and glutathione (GSH) pools to dehydroascorbate and glutathione disulfide respectively was observed in Zn2+-stressed seedlings, a clear indication of oxidative stress. However, exogenous application of EBR to stressed seedlings inhibited the oxidation of ascorbate and glutathione, maintaining redox molecules in reduced form. Under Zn2+ stress, enzymatic activities of ascorbate–glutathione cycle such as ascorbate peroxidase, monodehydroascorbate reductase increased but the dehydroascorbate reductase, glutathione reductase decreased. Zn2+ stress induced the gamma-glutamylcysteine synthetase, and glutathione-s-transferase activities in radish seedlings were further enhanced with EBR application. Zn2+ toxicity decreased the thiol content but, EBR supplementation resulted in restoration of thiol pool. The results of present study clearly demonstrated that external application of EBR modulates the AsA and GSH redox status to combat the oxidative stress of Zn2+ in seedlings via the AsA–GSH cycle and glutathione metabolism as an antioxidant defense system.

Effect of Selenium on Ascorbate–Glutathione Metabolism During PEG-induced Water Deficit in Trifolium repens L.

To elucidate the effect of selenium (Se) on the ascorbate–glutathione (ASC–GSH) cycle under drought stress, the activities of antioxidant enzymes and the levels of molecules involved in ASC–GSH metabolism were studied in Trifolium repens seedlings subjected to polyethylene glycol (PEG)-induced water deficit alone or combined with 5 μM Na2SeO4. Compared to the control, H2O2, thiobarbituric acid reactive substances (TBARS), ascorbate (ASC), dehydroascorbate (DHA), and glutathione disulfide (GSSG) contents increased, whereas a constant content of glutathione (GSH) and decreases in ASC/DHA and GSH/GSSG ratios were observed in the presence of PEG. The activities of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) were upregulated, except for monodehydroascorbate reductase (MDHAR) activity during PEG-induced water deficit. Se application decreased the contents of H2O2, TBARS, DHA, and GSSG, increased the levels of GSH and ASC, and inhibited the decreases of ASC/DHA and GSH/GSSG ratios. Although it did not affect APX activity significantly, Se addition improved the activities of MDHAR, DHAR, and GR. Furthermore, GR activity showed the highest increase followed by that of DHAR and MDHAR in decreasing order. These data indicated that fluctuations in ASC–GSH metabolism resulting from Se may have a positive effect on drought stress mitigation, and the regulation in the ASC–GSH cycle can be attributed mainly to GR and DHAR in PEG + Se-treated T. repens seedlings.

Developmental and tissue-specific changes in oxidative parameters and antioxidant systems in tomato fruits grown under salt stress

In tomato plants, salt stress has been induced to improve the quality of fruit. In general, plants under salt stress produce reactive oxygen species (ROS). Plants have their own ROS scavenging systems (antioxidant systems). In tomato plants, salt-induced changes in antioxidant systems have been examined extensively in leaves and roots; however, detailed information about salt-stressed fruits is not available. We examined the salt-induced changes in the antioxidant systems of the pericarp (containing epidermis) and pulp (containing seeds, placenta, and locule) during fruit ripening. Salt treatments were applied by adding 100 mM NaCl to the nutrient solution. In the pericarp and pulp, lipid peroxidation and hydrogen peroxide content were not increased by salt stress during ripening, indicating the absence of salt-induced oxidative stress. In the pericarp, the activities of superoxide dismutase (EC 1.15.1.1) and ascorbate (ASA)–glutathione (GSH) cycle-related enzymes increased with salt stress at the turning stage. Thus, at the turning stage, the antioxidant system may contribute to the enzymatic reaction involved ASA–GSH cycle. However, during the red and over-ripe fruit stages, salt stress produces little effect on antioxidant enzymes. In addition, the concentrations of antioxidants, such as the reduced form of ASA and GSH, increased during ripening in the control fruit, but those in the salt-stressed fruit remained unchanged. Therefore, the antioxidant system may contribute to the nonenzymatic reactions such as ASA and GSH taking place during the red and over-ripe fruit stages. In contrast, in the pulp, salt stress produces little effect on antioxidants and antioxidant enzymes. These results indicate that the salt-stressed fruit has protection mechanisms against salt-induced oxidative stress during ripening in both the pericarp and pulp. Salt-induced changes in antioxidant systems differed between the pericarp and pulp.

Cadmium responses in Arabidopsis thaliana: glutathione metabolism and antioxidative defence system

We investigated the effect of cadmium (Cd) on leaves of 3‐week‐old Arabidopsis thaliana with a particular interest on glutathione (GSH) production and consumption, and antioxidative defence. Plants were exposed to either 1 or 10 μM Cd for 1 week. Several genes and correspondent proteins known to participate in the regulation of redox states in plants were analysed in Cd‐stressed and unstressed plants. Cd induced a significant increase in the messenger RNA level of genes involved in GSH synthesis (gsh1 and gsh2) and phytochelatin synthase (pcs1). We observed a significant decrease of reduced GSH in Cd‐treated plants. In parallel, an increase of phytochelatin (PC), predominantly PC2 was observed. Cd treatment increased the accumulation of glutathione disulphide (GSSG), keeping the GSH/GSSG ratio lower than in control plants. The accumulation of GSSG was accompanied by a decrease of the glutathione reductase (gr) transcript level, while the activities of GR and nicotinamide nucleotide phosphate‐reducing enzymes were significantly enhanced. The antioxidative defence mechanism related to the ascorbate (AsA)–GSH cycle was studied in parallel. A general increase of reactive oxygen species scavenging enzymes such as ascorbate peroxidase, catalase or superoxide dismutase was observed. Our data suggest that the plants respond to Cd stress by the AsA–GSH defence network at both transcriptional and enzymatic level. Taken together, it appears that Arabidopsis plants exposed to 1 μM Cd were able to adopt a new metabolic equilibrium, allowing them to cope with this metal. However, when exposed to 10 μM Cd, loss of cellular redox homeostasis resulted in oxidative stress and toxicity.