Γ-glutamylcysteine Synthetase Activity Research Articles

Cysteine improves boar sperm quality via glutathione biosynthesis during the liquid storage.

ObjectiveSperm is particularly susceptible to reactive oxygen species (ROS) stress. Glutathione (GSH) is an endogenous antioxidant that regulates sperm redox homeostasis. However, it is not clear whether boar sperm could utilize cysteine for synthesis GSH to protect sperm quality from ROS damage. Therefore, the present study was undertaken to elucidate the mechanism of how cysteine is involved in protecting boar sperm quality during liquid storage.MethodsSperm motility, membrane integrity, lipid peroxidation, 4-hydroxyIlonenal (4-HNE) modifications, mitochondrial membrane potential, as well as the levels of ROS, GSH, and, ATP were evaluated. Moreover, the enzymes (GCLC: glutamate cysteine ligase; GSS: glutathione synthetase) that are involved in glutathione synthesis from cysteine precursor were detected by western blotting.ResultsCompared to the control, addition of 1.25 mM cysteine to the liquid storage significantly increased boar sperm progressive motility, straight-line velocity, curvilinear velocity, beat-cross frequency, membrane integrity, mitochondrial membrane potential, ATP level, acrosome integrity, activities of superoxide dismutase and catalase, and GSH level, while reducing the ROS level, lipid peroxidation and 4-HNE modifications. It was also observed that the GCLC and GSS were expressed in boar sperm. Interestingly, when we used menadione to induce sperm with ROS stress, the menadione associated damages were observed to be reduced by the cysteine supplementation. Moreover, compared to the cysteine treatment, the γ-glutamylcysteine synthetase (γ-GCS) activity, GSH level, mitochondrial membrane potential, ATP level, membrane integrity and progressive motility in boar sperm were decreased by supplementing with an inhibitor of GSH synthesis, buthionine sulfoximine.ConclusionThese data suggest that boar sperm could biosynthesize the GSH from cysteine in vitro. Therefore, during storage, addition of cysteine improves boar sperm quality via enhancing the GSH synthesis to resist ROS stress.

N-acetylcysteine provides protection against the toxicity of dietary T-2 toxin in juvenile Chinese mitten crab (Eriocheir sinensis)

The present research aimed to investigate the protection of N-acetylcysteine (NAC) on T-2 toxin-induced toxicity in Chinese mitten crab (Eriocheir sinensis). Crabs (0.27 ± 0.00 g) were fed four different diets (Control, T-2, 0.05% NAC + T-2 and 0.1% NAC + T-2) for 8 weeks. The results showed that T-2 toxin decreased growth performance, and induced oxidative stress, immunodepression and apoptosis in juvenile crabs. Crabs with NAC had higher weight gain, specific growth rate and hepatosomatic index than those in the T-2 group. The 0.1% NAC elevated crab survival and oxygen consumption rate and decreased alanine transaminase and aspartate transaminase in the serum of crabs fed T-2 toxin. Reactive oxygen species and malonaldehyde levels of hemolymph and hepatopancreas in crabs fed NAC were significantly lower than those in the T-2 group, while glutathione peroxidase, total antioxidant capacity, γ-glutamylcysteine synthetase activities and glutathione content significantly increased with 0.1% NAC. Total hemocyte count, respiratory burst activity, total protein, acid phosphatase (ACP) in hemolymph and ACP, alkaline phosphatase (AKP) in hepatopancreas were significantly enhanced with 0.1% NAC. Phagocytosis and AKP in hemolymph were significantly enhanced with 0.05% NAC supplementation. Furthermore, the mRNA expressions of antimicrobial peptides (ALF1, ALF2, crustin-1 and crustin-2) and peroxinectin were up-regulated with NAC. Besides NAC down-regulated the mRNA expressions of Relish, lipopolysaccharide-induced TNF-alpha factor, caspase-3, caspase-8 and p53 genes, and the ratio of Bax to Bcl-2 in the hepatopancreas. Our findings indicate that dietary NAC could improve the growth performance and survival of juvenile Chinese mitten crab fed T-2 toxin. The 0.1% NAC could effectively alleviate oxidative stress, immunodepression and apoptosis induced by T-2 toxin.

Methylglyoxal-Scavenging Enzyme Activities Trigger Erythroascorbate Peroxidase and Cytochrome c Peroxidase in Glutathione-Depleted Candida albicans.

γ-Glutamylcysteine synthetase (Gcs1) and glutathione reductase (Glr1) activity maintains minimal levels of cellular methylglyoxal in Candida albicans. In glutathione-depleted Δgcs1, we previously saw that NAD(H)-linked methylglyoxal oxidoreductase (Mgd1) and alcohol dehydrogenase (Adh1) are the most active methylglyoxal scavengers. With methylglyoxal accumulation, disruptants lacking MGD1 or ADH1 exhibit a poor redox state. However, there is little convincing evidence for a reciprocal relationship between methylglyoxal scavenger genes-disrupted mutants and changes in glutathione-(in)dependent redox regulation. Herein, we attempt to demonstrate a functional role for methylglyoxal scavengers, modeled on a triple disruptant (Δmgd1/Δadh1/Δgcs1), to link between antioxidative enzyme activities and their metabolites in glutathione-depleted conditions. Despite seeing elevated methylglyoxal in all of the disruptants, the result saw a decrease in pyruvate content in Δmgd1/Δadh1/Δgcs1 which was not observed in double gene-disrupted strains such as Δmgd1/Δgcs1 and Δadh1/Δgcs1. Interestingly, Δmgd1/Δadh1/Δgcs1 exhibited a significantly decrease in H2O2 and superoxide which was also unobserved in Δmgd1/Δgcs1 and Δadh1/Δgcs1. The activities of the antioxidative enzymes erythroascorbate peroxidase and cytochrome c peroxidase were noticeably higher in Δmgd1/Δadh1/Δgcs1 than in the other disruptants. Meanwhile, Glr1 activity severely diminished in Δmgd1/Δadh1/Δgcs1. Monitoring complementary gene transcripts between double gene-disrupted Δmgd1/Δgcs1 and Δadh1/Δgcs1 supported the concept of an unbalanced redox state independent of the Glr1 activity for Δmgd1/Δadh1/Δgcs1. Our data demonstrate the reciprocal use of Eapx1 and Ccp1 in the absence of both methylglyoxal scavengers; that being pivotal for viability in non-filamentous budding yeast.

Glutathione ethyl ester supplementation prevents airway hyper-responsiveness in mice.

BackgroundOxidative stress plays an important role in the pathogenesis of asthma. Glutathione (GSH) is considered to be one of the most important antioxidants. Our study systematically investigated the effect of the GSH alternative, glutathione ethyl ester (GSH-EE), on airway hyper-responsiveness (AHR) in mice.MethodsSixty-three male specific pathogen-free mice were used. Asthma was induced using a single dose of ovalbumin (OVA). The normal group (n=15) received vehicle only [Al(OH)3 in saline]. Then, 48 mice were divided into two groups, including a control group who received sodium phosphate buffer (pH =7.4), and the GSH-EE group who received 0.1% GSH-EE. AHR was measured 2, 6, and 12 hours after exposure to nebulized OVA (0.01%). The animals were then sacrificed, and lung tissue and the bronchi-alveolar lavage fluid (BALF) were harvested. Factors involved in the antioxidant response to asthma were then measured in these tissues, including thiol content (from GSH and protein), γ-glutamylcysteine synthetase (γ-GCS) activity and expression, and nuclear factor-erythroid-2-related factor (Nrf2) expression.ResultsThe GSH-EE group showed a significant attenuation of AHR (P<0.01) 2 hours after OVA challenge, and significantly enhanced thiol contents by approximately 45% (P<0.05) at 2 and 6 hours after the last OVA challenge, compared to the control group. γ-GCS activity was also higher in the GSH-EE group compared to the control group at different time points (P<0.01). γ-GCSh and Nrf2 protein expression increased in the GSH-EE group and the control group compared with the normal group, but there was no statistically significant difference (P>0.05) between the GSH-EE group and the control group.ConclusionsGSH-EE supplementation can prevent AHR in asthmatic mice during the early stages. It may function by serving as a precursor for GSH biosynthesis and by protecting sulfhydryl groups from oxidation.

The effect of short-term methionine restriction on glutathione synthetic capacity and antioxidant responses at the whole tissue and mitochondrial level in the rat liver.

Dietary methionine restriction (MR) where methionine is the sole source of sulfur amino acid increases lifespan in diverse species. Methionine restricted rodents experience a decrease in glutathione (GSH), a major antioxidant, in several tissues, which is paradoxical to longevity interventions because tissues with low GSH might experience more oxidative damage. Liver plays a key role in GSH synthesis and here we examined how MR influences GSH metabolism in the liver. We also hypothesised that low GSH might be subsidized by compensatory pathway(s) in the liver. To investigate GSH synthesis and antioxidant responses, Fischer-344 rats were given either a MR diet or a control diet for 8 weeks. Based on γ-glutamylcysteine synthetase activity, GSH synthetic capacity did not respond to low dietary methionine availability. Tissue level protein and lipid oxidation markers do not support elevated oxidative damage, despite low GSH availability. Whole tissue and mitochondrial level responses to MR differed. Specifically, the activity of glutathione reductase and thioredoxin reductase increase in whole liver tissue which might offset the effects of declined GSH availability whereas mitochondrial GSH levels were unperturbed by MR. Moreover, enhanced proton leak in liver mitochondria by MR (4 week) presumably diminishes ROS production. Taken together, we suggest that the effect of low GSH in liver tissue is subsidized, at least in part, by increased antioxidant activity and possibly by enhanced mitochondrial proton leak.

Nitric oxide regulates glutathione synthesis and cold tolerance in forage legumes

The role of nitric oxide (NO) in cold acclimation through regulation on glutathione synthesis was investigated in Medicago falcata, Medicago truncatula and NITRATE REDUCTASE 1 (NIA1) mutants nia1-1 and nia1-2 of M. truncatula. M. falcata is an important forage legume with great cold tolerance, while M. truncatua is a model legume with lower cold tolerance than M. falcata. Glutathione synthesis and redox were promoted in both species in response to cold treatment, which showed an association with cold induced γ-glutamylcysteine synthetase (γ-ECS) and glutathione reductase (GR) activities with increased transcript levels of γ-ECS1, glutathione synthetase1 (GSHS1), GSHS2, GR1 and GR2 in both species. The cold induced glutathione concentration, glutathione redox, activities of γ-ECS and GR, and transcripts of γ-ECS1, GSHS1, GSHS2, GR1 and GR2 were suppressed by pretreatment with either NR inhibitor or NO scavenger in M. falcata and M. truncatula or by knock-out of NIA1 in nia1-1 and nia1-2 mutants. Moreover, treatment with exogenous NO resulted in the similar alterations in glutathione synthesis and redox as well as levels in enzyme activity and transcripts as cold treatment. The results suggested that NR-derived NO production is involved in cold regulated glutathione synthesis and redox by regulating gene expression. Higher levels of glutathione concentration, glutathione redox, activities of γ-ECS and GR, and transcripts of γ-ECS1, GSHS1, GSHS2, GR1 and GR2 were observed in M. falcata than in M. truncatula after cold treatment, suggesting that the difference in glutathione synthesis and redox is associated with the differential cold tolerance between the two species.

A novel compound AB-38b improves diabetes-associated cognitive decline in mice via activation of Nrf2/ARE pathway

ObjectiveDiabetes-associated cognitive decline (DACD) is increasingly being concerned, and oxidative stress plays a vital role in the pathological process. AB-38b is a novel synthetic compound with two specific active groups of biphenyl dicarboxylate and α, β unsaturated ketone, showing good antioxidant activity. The aim of this study was to investigate the ameliorative effects of AB-38b on DACD in mice, and to explore the possible mechanisms from glyoxylase 1 (Glo-1) enhancement and NF-E2-related factor-2 (Nrf2) activation. MethodsExperimental type 2 mouse model of diabetes with C57BL/6 mice was made through high-fat diet combining with intraperitoneal streptozotocin. Diabetic mice were treated by gavage with AB-38b (0, 10, 20 and 40 mg/kg) or resveratrol (40 mg/kg), a typical inducer of Nrf2, for 8 weeks. Cognitive performances were evaluated by the novel object recognition task. Then brain tissues were collected to assess hippocampal damages, protein glycation, Glo-1 functions and protein expression, and the classic Nrf2/ARE pathway. ResultAB-38b markedly increased the preference index to novel object and the number of neurons in hippocampal CA1 area of diabetic mice. AB-38b significantly elevated the activity and protein of Glo-1, while reduced the levels of advanced glycation end products (AGEs) and protein expression of its receptor RAGE. Moreover, AB-38b raised Nrf2 expression and phosphorylation, as well as the protein expression and enzymatic activity of γ-glutamylcysteine synthetase, a well-known gene of Nrf2/ARE pathway, in hippocampus of the diabetic mice. ConclusionAB-38b improved the cognitive performances of diabetic mice, which was achieved via up-regulation of Glo-1 and activation of Nrf2/ARE pathway.

Ameliorative role of boron to toxicity of aluminum in trifoliate orange roots

Aluminum (Al) toxicity is a major limiting factor for plant productivity. Boron (B) could mitigate Al toxicity in many plant species. However, information about the mechanisms of B alleviating Al toxicity in citrus is lacking. Trifoliate orange rootstock (Poncirus trifoliate L. Raf.) seedlings were irrigated with a nutrient solution containing two B and two Al levels. Results showed that exposure to Al severely impeded plant growth-related parameters. However, B supply improved plant biomass, root activity and relative root elongation under Al stress. Furthermore, B reduced the Al-induced H2O2 accumulation in roots as evidenced by lower fluorescence intensity of H2O2 staining. Boron decreased the Al-stimulated ascorbate (AsA) synthesis by down-regulated AsA synthesis-related metabolites in the L-galactose pathway. Boron alleviated some of the toxic effects of Al by decreasing redox states of AsA and enzyme activities involved in ascorbate-glutathione (AsA-GSH) cycle, ascorbate peroxidase, dehydroascorbate reductase, glutathione reductase and glutathione peroxidase while increased glutathione (GSH) content and γ-glutamylcysteine synthetase (γ-GCS) activity. Overall, our results suggest that B protects roots against Al-induced oxidative stress possibly by reducing metabolites accumulation in the L-galactose pathway of AsA synthesis and regulating AsA-GSH cycle.

Involvement of glutathione metabolism in Eichhornia crassipes tolerance to arsenic.

Aquatic macrophytes are potentially useful for phytoremediation programmes in environments contaminated by arsenic (As). Biochemical and physiological modification analyses in different plant parts are important to understand As tolerance mechanisms. The objective was to evaluate glutathione metabolism in leaves and roots of Eichhornia crassipes (Mart.) Solms treated to As. Specimens of E.crassipes were cultured for 3days in Clark's nutrient solution containing 7μm As. The enzymes ATP sulphurylase (ATPS), glutathione reductase (GR), glutathione peroxidase (GSH-Px), glutathione sulphotransferase (GST) and γ-glutamylcysteine synthetase (γ-ECS) activity, glutathione content, total protein and non-protein thiols were evaluated. The ATPS activity increased in roots. GR activity in leaves and GSH-Px in roots were lower. GST activity was higher in roots and lower in leaves, and γ-ECS activity was higher in leaves. Glutathione levels were lower, total thiol levels were higher and non-protein levels did not change in E.crassipes leaves and roots. Exposure to As increased enzyme activity involved with sulphur metabolism, such as ATPS. Higher GR activity and lower GSH-Px indicate increased glutathione conjugation to As due to increased GSH availability. The higher GST activity indicates its participation in As detoxification and accumulation through As GSH conjugation. Changes in glutathione and thiol levels suggest high phytochelatin synthesis. In conclusion, the increments in ATPS, GR, GST and γ-ECS activity indicate that these enzymes are involved in GSH metabolism and are part of the E.crassipes As detoxification mechanism.

Curcumin inhibits the growth via Wnt/β-catenin pathway in non-small-cell lung cancer cells.

In recent decades, the death rate from lung cancer appears to be an increasing yearly trend, particularly for non-small-cell lung cancer (NSCLC). Curcumin is a yellow pigment found in turmeric rhizomes, reported to exhibit various anti-inflammatory, anti-angiogenic, anti-proliferative, and antioxidant properties. Many reports have suggested that curcumin could induce apoptosis in malignant cells, and therefore, has great potential in tumor treatment. However, little is known about the effect of curcumin on NSCLC or its associated mode of action. Therefore, in this study, we explored curcumin's effect on NSCLC and investigated its associated mechanism. The non-small-cell lung cancer (NSCLC) cell line A549 was cultured and subjected to MTT and clonogenic survival assays to assess cell proliferation. Reactive oxygen species (ROS) levels were measured using a Fluostar Omega Spectrofluorimeter. Superoxide dismutase (SOD) and γ-glutamyl cysteine synthetase (γ-GCS) activity in A549 cells were both determined by a commercial determination kit. Expression levels of p-GSK3β (Ser9), c-Myc, cyclin D1, β-catenin α-tubulin, and proliferating cell nuclear antigen (PCNA) were analyzed by Western blot. Results of the MTT and clonogenic survival assay indicated that curcumin reduced A549 proliferation. ROS levels and SOD and γ-GCS activities were detected. Curcumin decreased intracellular ROS levels and increased SOD and γ-GCS activity. Meanwhile, the ROS inhibitor N-Acetylcysteine (NAC) reversed the decrease in ROS levels and the increase in SOD and γ-GCS activity. These results indicate that oxidative stress is involved in the curcumin-induced reduction of A549 viability. Curcumin also strongly inhibited β-catenin and p-GSK3β (Ser9) protein expression, as well as the expression of downstream cyclin D1 and c-Myc. Similarly, NAC reversed the inhibition of β-catenin and p-GSK3β (Ser9) protein expression, as well as the expression of downstream cyclin D1 and c-Myc. We showed that curcumin inhibits NSCLC proliferation via the Wnt/β-catenin pathway.

The effect of a doubled glutathione level on parameters affecting the germinability of recalcitrant Acer saccharinum seeds during drying

Approximately 20% of plant species, including silver maple (Acer saccharinum L.), produce seeds that are sensitive to desiccation, which is reflected in their poor storage potential and viability. In the search for a compound that can improve seed recalcitrance, freshly harvested seeds were soaked in either 2.5 mM reduced glutathione (GSH) or water and desiccated to comparable water levels of 55–20%. We examined the impact of a doubled endogenous level of glutathione on the seed germination capacity, the activity of enzymes involved in glutathione metabolism, the cell membrane components and integrity, reactive oxygen species, and ascorbate levels. GSH treatment resulted in slower dehydration and a higher germination capacity. The increased glutathione was mainly consumed by glutathione S-transferase, leading to more efficient detoxification, and by dehydroascorbate reductase (DHAR), accelerating the ascorbate regeneration. As a result, the cellular environment became more reduced, and protection of the membrane structures was enhanced. The ameliorated membrane integrity was manifested via a lower electrolyte leakage and a lower lipid peroxide level despite the higher level of hydrogen peroxide (H2O2) detected in the GSH-treated seeds. The degradation of phospholipids (PLs) was less intense and related to the phosphatidylinositol (PI) level, which is the precursor of the phospholipase D cofactor, whereas in water-soaked seeds, PL degradation was promoted by H2O2. The germination capacity of the dehydrated seeds depended primarily on the level of H2O2, lipid hydroxyperoxides, electrolyte leakage, GSH, the half-cell reduction potential of glutathione, PI, and the activity of DHAR and γ-glutamylcysteine synthetase. Interestingly, H2O2 affected all of the parameters. The germination of GSH-boosted seeds was strongly impacted by the pool of ascorbate, the half-cell reduction potential of ascorbate, and the glutathione peroxidase activity. In general, germination was DHAR activity-dependent. A strong negative correlation was detected in the water-soaked seeds, whereas a strong positive correlation was detected in the GSH-treated seeds. The enhanced level of glutathione likely improved the efficiency of the ascorbate–glutathione cycle, confirming its effect on seed germinability after dehydration.

Effects of L-glutamine on boar sperm quality during liquid storage at 17°C

The quality of boar spermatozoa is affected by oxidative stress during preservation in vitro. It has been demonstrated that L-Glutamine (Gln) can effectively protect cells from oxidative stress-induced injury. There are, however, no reports to date evaluating the effects of Gln on boar semen liquid preservation at 17 °C. The aims of the present study were to elucidate whether the addition of Gln to the extender BTS could improve the quality of boar spermatozoa when stored at 17 °C and to determine the mechanism underlying Gln protection of spermatozoa against preservation-induced damage. Boar semen samples were collected and diluted with Beltsville Thawing Solution (BTS) containing different concentrations (0, 10, 20, 40 or 80 mM) of Gln. The results indicated the addition of 20 mM Gln to the BTS improved (P < 0.05) the motility, acrosome integrity and membrane integrity of boar sperm during liquid preservation. Interestingly, treatment of spermatozoa with Gln addition to the extender resulted in ROS quenching, while enhancing γ-glutamyl cysteine synthetase (γ-GCS) activity, and glutathione (GSH) content of spermatozoa. These results suggest that BTS supplemented with Gln can provide greater protective capacity to boar sperm against oxidative stress by enhancing GSH synthesis during liquid preservation.

Glutathione-induced alleviation of cadmium toxicity in Zea mays

Glutathione (GSH) is known to alleviate cadmium (Cd) stress in many plant species. However, the comprehensive mechanisms responsible for this effect in maize are still need more investigation. Here, a combination of physiological and molecular approaches was utilized in GSH-Cd treated maize seedlings, which revealed that GSH reversed the adverse effects of Cd, as reflected by plant growth, plant hormones, vacuole, stoma development, gene expression, etc. Plant growth, root cell viability, photosynthetic capacity, redox equilibrium, and cell ultrastructure recovery following GSH treatment, coupled with the strong up-regulation of Cd tolerance-related genes (e.g., phytochelatin synthetase-like protein, MYB and WRKY transcription factors, and CYP450), demonstrated the efficient activation of cellular defense against Cd toxicity. The addition of GSH significantly elevated GSH/GSSG ratio and the activity of γ-glutamylcysteine synthetase in both shoots and roots and markedly reduced Cd concentration in shoots. Ethylene emission rate and abscisic acid (ABA) content were significantly reduced after GSH application in the presence of Cd, except ABA content in leaves. These findings highlighted the significance of GSH in alleviating Cd-stress in maize and indicate a promising strategy for safe food production.

Effects of dietary conjugated linoleic acid on lipid peroxidation in breast and thigh muscles of broiler chickens

The effect of dietary supplementation with conjugated linoleic acid (CLA) on lipid peroxidation in the breast muscles (Pectoralis major and Pectoralis minor) and thigh muscles of broiler chickens was investigated. A total of ninety-six 21-day-old Arbor Acres male broiler chickens were assigned to 2 dietary treatments (0 and 1.5% CLA) with 8 replicates per 6 chickens each. The left breast and thigh muscles of broiler chickens were excised on day 42. The broilers receiving the CLA diet exhibited lower malondialdehyde content and reactive oxygen species production (P &amp;lt; 0.01) in the breast and thigh muscles than the broilers receiving the control diet. After the dietary CLA supplementation, the glutathione (GSH) content in the breast and thigh muscles of the broiler chickens fed the CLA diet increased by 21.89 and 21.56%, respectively (P &amp;lt; 0.05) while the γ-glutamylcysteine synthetase（γ-GCS）activity increased by 28.57 and 25.80% (P &amp;lt; 0.05), respectively. Dietary CLA significantly increased the CLA content (P &amp;lt; 0.01) and saturated fatty acid content (P &amp;lt; 0.05), and decreased the monounsaturated fatty acid content (P &amp;lt; 0.01) in the breast and thigh muscles. These results showed that dietary CLA may decrease the lipid peroxidation level in the breast and thigh muscles of broiler chickens perhaps through increasing the γ-GCS activity to induce GSH synthesis and changing the fatty acid composition to increase oxidative stability.

Glutamine protects rabbit spermatozoa against oxidative stress via glutathione synthesis during cryopreservation.

Mammalian spermatozoa are extremely susceptible to high doses of reactive oxygen species (ROS). The aim of the present study was to investigate the potential role of glutamine in protecting rabbit spermatozoa against ROS stress during cryopreservation and post-thaw incubation. Freshly ejaculated semen was diluted with Tris-citrate-glucose extender supplemented with glutamine. The addition of 20mM glutamine significantly improved sperm motility, acrosome integrity, membrane integrity and mitochondrial activity. Meanwhile, 20mM glutamine addition decreased lipid peroxidation and DNA damage in frozen-thawed spermatozoa. Interestingly, supplementation with 20mM glutamine led to increases in glutathione content and γ-glutamyl cysteine synthetase and glutathione peroxidase activity, with concomitant decreases in ROS levels during cryopreservation and post-thaw incubation. In conclusion, the addition of glutamine to extender solutions protects rabbit spermatozoa from ROS attack by enhancing glutathione synthesis.

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

ABSTRACTA 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.

The mechanism of improved intracellular organic selenium and glutathione contents in selenium-enriched Candida utilis by acid stress.

Batch culture of Candida utilis CCTCC M 209298 for the preparation of selenium (Se)-enriched yeast was carried out under different pH conditions, and maximal intracellular organic Se and glutathione (GSH) contents were obtained in a moderate acid stress environment (pH3.5). In order to elucidate the physiological mechanism of improved performance of Se-enriched yeast by acid stress, assays of the key enzymes involved in GSH biosynthesis and determinations of energy supply and regeneration were performed. The results indicated that moderate acid stress increased the activity of γ-glutamylcysteine synthetase and the ratios of NADH/NAD+ and ATP/ADP, although no significant changes in intracellular pH were observed. In addition, the molecular mechanism of moderate acid stress favoring the improvement of Se-yeast performance was revealed by comparing whole transcriptomes of yeast cells cultured at pH3.5 and 5.5. Comparative analysis of RNA-Seq data indicated that 882 genes were significantly up-regulated by moderate acid stress. Functional annotation of the up-regulated genes based on gene ontology and the Kyoto Encyclopedia of Genes and Genome (KEGG) pathway showed that these genes are involved in ATP synthesis and sulfur metabolism, including the biosynthesis of methionine, cysteine, and GSH in yeast cells. Increased intracellular ATP supply and more amounts of sulfur-containing substances in turn contributed to Na2SeO3 assimilation and biotransformation, which ultimately improved the performance of the Se-enriched C. utilis.

Aluminium Induced Glutathione is Essential for Developing Resistance Against Fusarium Infection in Wheat

Pre-treatment of wheat seedlings with 50 μM concentration of aluminium sulphate (Al) induces enhanced accumulation of total glutathione. After 48 h the abundance in total glutathione in Al pre-treated seedlings increased by 60 % as compared to untreated control seedlings. The abundance in total glutathione was proportional to the increase in the reduced form of glutathione (GSH). The abundance in glutathione content was linked with the enhancement of γ-glutamylcysteine synthetase (γ-ECS) activity. To assess the role of over-expressed glutathione during plant-pathogen interaction, Al primed seedlings were infected with Fusarium oxysporum. Studies on growth parameters, disease severity index (DSI) and reactive oxygen species (ROS) activity suggested over-expressed glutathione as a key player in mitigating Fusarium induced oxidative stress in wheat seedlings. To emphasize the role of Al induced glutathione in restricting fungal infection, wheat seedlings were treated with 1 mM concentration of Dl-buthionine-SR-sulfoximine (BSO) for 48 h. BSO treated seedlings showed 30 % decline in total glutathione content coupled with decline in γ-ECS activity. BSO treated seedlings infected with Fusarium were rapidly and highly susceptible to the fungus as compared to untreated control seedlings when infected with the Fusarium. Thus, Al pre-treated seedlings with enhanced concentration of glutathione maintained the cellular redox balance and protected wheat against Fusarium induced oxidative stress.

Mangiferin Upregulates Glyoxalase 1 Through Activation of Nrf2/ARE Signaling in Central Neurons Cultured with High Glucose.

Mangiferin, a natural C-glucoside xanthone, has anti-inflammatory, anti-oxidative, neuroprotective actions. Our previous study showed that mangiferin could attenuate diabetes-associated cognitive impairment of rats by enhancing the function of glyoxalase 1 (Glo-1) in brain. The aim of this study was to investigate whether Glo-1 upregulation by mangiferin in central neurons exposed to chronic high glucose may be related to activation of Nrf2/ARE pathway. Compared with normal glucose (25mmol/L) culture, Glo-1 protein, mRNA, and activity levels were markedly decreased in primary hippocampal and cerebral cortical neurons cultured with high glucose (50mmol/L) for 72h, accompanied by the declined Nrf2 nuclear translocation and protein expression of Nrf2 in cell nucleus, as well as protein expression and mRNA level of γ-glutamylcysteine synthetase (γ-GCS) and superoxide dismutase activity, target genes of Nrf2/ARE signaling. Nonetheless, high glucose cotreating with mangiferin or sulforaphane, a typical inducer of Nrf2 activation, attenuated the above changes in both central neurons. In addition, mangiferin and sulforaphane significantly prevented the formation of advanced glycation end-products (AGEs) reflecting Glo-1 activity, while elevated the level of glutathione, a cofactor of Glo-1 activity and production of γ-GCS, in high glucose cultured central neurons. These findings demonstrated that Glo-1 was greatly downregulated in central neurons exposed to chronic high glucose, which is expected to lead the formation of AGEs and oxidative stress damages. We also proved that mangiferin enhanced the function of Glo-1 under high glucose condition by inducing activation of Nrf2/ARE signaling pathway.

Early Abscisic Acid Accumulation Regulates Ascorbate and Glutathione Metabolism in Soybean Leaves Under Progressive Water Stress

Ascorbate (AsA) and glutathione (GSH) play an important role in improving the tolerance of plants to water stress. The objective of this study was to investigate the effect of early abscisic acid (ABA) accumulation on AsA and GSH metabolism in soybean plants after 24 h of exposure to progressive water stress. The results showed that AsA, total AsA, GSH and total GSH content, and ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), GSH reductase (GR), GSH peroxidase (GPX), l-galactono-1,4-lactone dehydrogenase (GLDH), and γ-glutamylcysteine synthetase (γ-GCS) activities were increased by progressive water stress. The above increases, except for total GSH content and the activities of GLDH and γ-GCS, were blocked by pretreatment with tungstate, an ABA biosynthesis inhibitor, which significantly suppressed the early increase in ABA and reactive oxygen species (ROS) in stressed plants. Application of ABA reversed the effects of tungstate. Pretreatments with several ROS scavengers, such as Tiron and dimethylthiourea (DMTU), and an inhibitor of NADPH oxidase, diphenyleneiodonium (DPI), significantly arrested the early accumulation of ROS but not ABA in stressed plants. Furthermore, the above-mentioned pretreatments remarkably prevented any increase in APX, MDHAR, DHAR, GR, and GPX activities, as well as AsA, total AsA and GSH levels in stressed plants. Our results indicated that early ABA accumulation caused by progressive water stress triggers an early rise in ROS levels, which, in turn, leads to regulation of AsA and GSH metabolism.