A rice cadmium （Cd） sensitive mutant cadB-1 was obtained using Agrobacterium tumefaciens mediated system.After exposure of cadB-1 and wild type （WT） rice seedlings to a range of Cd concentrations for 10 d,Cd accum...A rice cadmium （Cd） sensitive mutant cadB-1 was obtained using Agrobacterium tumefaciens mediated system.After exposure of cadB-1 and wild type （WT） rice seedlings to a range of Cd concentrations for 10 d,Cd accumulated to higher levels in roots,stems and leaves of both cadB-1 and WT with increasing external Cd concentrations,and the inhibition of seedling growth in cadB-1 was more serious than in WT.Hydrogen peroxide accumulation was higher in leaves and roots of cadB-1.The ratios of reduced glutathione （GSH）/oxidized glutathione （GSSG）,ascorbate （ASC）/dehydroascorbate （DHA） and reduced nicotinamide adenine dinucleotide phosphate （NADPH）/oxidized nicotinamide adenine dinucleotide phosphate （NADP＋） were lower in cadB-1 than in WT both in leaves and roots under high Cd levels.The activities of ascorbate peroxidase （APX）,glutathione peroxidase （GR）,dehydroascorbate reductase （DHAR） and monodehydroascorbate reductase （MDHAR） were also lower in cadB-1 than in WT both in leaves and roots under the treatment of high levels of Cd.Our results suggest that under Cd stress,the ASC-GSH cycle was more seriously inhibited in cadB-1 than in WT,indicating that the mutant cadB-1 is less able to scavenge reactive oxygen species and sensitive to Cd.展开更多
Nonylphenol( NP) is a stable metabolic product of nonylphenol ethoxylates,which is widely used as an industrial surfactant. NP has been classified as an endocrine disrupter,and its toxicity to organisms can be biomagn...Nonylphenol( NP) is a stable metabolic product of nonylphenol ethoxylates,which is widely used as an industrial surfactant. NP has been classified as an endocrine disrupter,and its toxicity to organisms can be biomagnified through the food chain. As compared with the endocrine disrupting effect,the toxicity of NP to organisms has not been studied intensively,and the toxicity mechanisms have often been ignored. In the present study,Microcystis aeruginosa,a freshwater alga belonging to the first level of the trophic chain,was chosen to detect the toxicity of NP. The mechanisms of toxicity mediated by the AsA-GSH cycle were explored. The acute toxicity of NP to M. aeruginosa within 96 h was studied and an EC_(50) concentration of 3. 45 mg/L was found. Further,the results showed that the toxicity of NP increased with the increase in concentration and exposure time. As compared with that in the control,the APX and MDHAR activities mostly increased,whereas DHAR activity fluctuated.However,the AsA content elevated at first,but decreased significantly after 72 h. For the GSH system,GR activity was always higher than that in the control. Nevertheless,the reduced GSH content was mostly inhibited. Therefore,the performance of AsA-GSH antioxidant defense system could explain the results of NP toxicity: the enzyme activities and antioxidant molecules increased initially,but an overall decline appeared after exposure for 24 h. This research is helpful for estimating the toxicity of NP integrally and improves people's understanding of mechanisms of NP toxicity in algae.展开更多
As the most northerly mangrove species in China, Kandelia obovata may undergo extreme cold event stress. Enhancing the cold tolerance of this species is crucial to its successful afforestation. This study aimed to det...As the most northerly mangrove species in China, Kandelia obovata may undergo extreme cold event stress. Enhancing the cold tolerance of this species is crucial to its successful afforestation. This study aimed to determine the resistance of K. obovata seedlings to low temperature stress by cold acclimation and to explain the mechanisms for alleviating cold injury. To understand these mechanisms, seedlings that were acclimatized and not acclimatized were exposed to 5℃/- 2℃(day/night)for 48 h.Results showed that low temperature stress reduced leaf photosynthesis of non-acclimatized seedlings by inducing oxidative stress and structural damage to chloroplasts. These phenomena were shown by increasing levels of malondialdehyde (MDA), O2-and H2O2, as well as decreasing enzyme activities in the ascorbate–glutathione (AsA-GSH) cycle. However, cold-acclimatized seedlings had improved photosynthetic rates and efficiency of photosystem II (PSII) under low temperature stress. Compared with non-acclimatized seedlings, leaves of coldacclimatized seedlings under low temperature stress for 48 h exhibited higher anti-oxidative enzyme activities, lower levels of O2^- and H2O2, less damage to chloroplast structure, and removed 33.7% of MDA at low temperature stress for 48 h. The data indicate that cold acclimation enhances photosynthetic capacity by effectively regulating activation in the PSII electron transport and the AsA–GSH cycle to scavenge excess ROS in chloroplasts, while the latter is more important.展开更多
Glutathione peroxidase, the first example of selenoproteins identified in mammals, was subjected to force field calculations and molecular dynamics in order to enable a clearer comprehension of enzymatic selenium cata...Glutathione peroxidase, the first example of selenoproteins identified in mammals, was subjected to force field calculations and molecular dynamics in order to enable a clearer comprehension of enzymatic selenium catalysis. Starting from the established X-ray structure of bovine GPX, all kinetically defined intermediates and enzyme substrate complexes were modelled. The models thus obtained support the hypothesis that the essential steps of the catalysis are three distinct redox changes of the active site selenium which, in the ground state, presents itself at the surface of selenoperoxidases as the center of a characteristic triad built by selenocysteine, glutarnine and tryptophan. In GPX, four arginine residues and a lysine residue provide an electrostatic architecture which, in each reductive step, directs the donor substrate GSH towards the catalytic center in such a way that 1ts sulfhydryl group must react with the selenium moiety. To this end, different equally efficient modes of substrate binding appear possible. The models are consistent with substrate specificity data, kinetic pattern and other functional characteristics of the enzyme. Comparison of molecular models of GPX with those of other members of the GPX superfamily reveals that the cosubstrate binding mechanisrns are unique for the classical type of cytosolic glutathione peroxidases but cannot operate e. g. in plasma GPX and phospholipid hydroperoxide GPX. The structural differences between the selenoperoxidases, shown to be relevant to their specificities, are discussed in terms of functional diversification within the GPX superfamily展开更多
This research provides, to the authors’ knowledge, the first integrative model of oxidative stress and C1 metabolism in plants. Increased oxidative stress can cause irreversible damage to photosynthetic components an...This research provides, to the authors’ knowledge, the first integrative model of oxidative stress and C1 metabolism in plants. Increased oxidative stress can cause irreversible damage to photosynthetic components and is harmful to plants. Perturbations at the genetic level may increase oxidative stress and upregulate antioxidant systems in plants. One of the key mechanisms involved in oxidative stress regulation is the ascorbate-glutathione cycle which operates in chloroplasts as well as the mitochondria and is responsible for removal of reactive oxygen species (ROS) generated during photosynthetic operations and respiration. In this research, the complexity of molecular pathway systems of oxidative stress is modeled and then integrated with a previously developed in silico model of C1 metabolism system. This molecular systems integration provides two important results: 1) demonstration of the scalability of the CytoSolve®?Collaboratory™, a computational systems biology platform that allows for modular integration of molecular pathway models, by coupling the in silico model of oxidative stress with the in silico model of C1 metabolism, and 2) derivation of new insights on the effects of oxidative stress on C1 metabolism relative to formaldehyde (HCHO), a toxic molecule, and glutathione (GSH), an important indicator of oxidative homeostasis in living systems. Previous in silico modeling of C1 metabolism, without oxidative stress, observed complete removal of formaldehyde via formaldehyde detoxification pathway and no change in glutathione concentrations. The results from this research of integrative oxidative stress with C1 metabolism, however, demonstrate significant upregulation of formaldehyde concentrations, with concomitant downregulation and depletion of glutathione. Sensitivity analysis indicates that kGSH-HCHO, the rate constant of GSH-HCHO binding, VSHMT, the rate of formation of sarcosine from glycine, and , the rate of superoxide formation significantly affect formaldehyde homeostasis in the C1 metabolism. Future research may employ this integrative model to explore which conditions initiate oxidative stress and the resultant upregulation and downregulation of formaldehyde and glutathione.展开更多
To explore whether glutathione regulates diapause determination and termina tion in the bivoltine silkworm Bombyx mori, we monitored the changes in glutathione redox cycle in the ovary of both diapanse and nondiapause...To explore whether glutathione regulates diapause determination and termina tion in the bivoltine silkworm Bombyx mori, we monitored the changes in glutathione redox cycle in the ovary of both diapanse and nondiapauseegg producers, as well as those in dia pause eggs incubated at different temperatures. The activity ofthioredoxin reductase （TrxR） was detected in ovaries but not in eggs, while neither ovaries nor eggs showed activity of glutathione peroxidase. A lower reduced glutathione/oxidized glutathione （GSH/GSSG） ratio was observed in the ovary of diapauseegg producers, due to weaker reduction of oxidized glutathione （GSSG） to the reduced glutathione （GSH） catalyzed by glutathione reductase （GR） and TrxR. This indicates an oxidative shift in the glutathione redox cy cle during diapause determination. Compared with the 25℃treated diapause eggs, the 5℃treated diapause eggs showed lower GSH/GSSG ratio, a result of stronger oxidation of GSH catalyzed by thioredoxin peroxidase and weaker reduction of GSSG catalyzed by GR. Our study demonstrated the important regulatory role of glutathione in diapause determination and termination of the bivoltine silkworm.展开更多
Pyridine nucleotides, ascorbate and glutathione are major redox metabolites in plant cells, with specific roles in cellular redox homeostasis and the regulation of the cell cycle. However, the regulation of these meta...Pyridine nucleotides, ascorbate and glutathione are major redox metabolites in plant cells, with specific roles in cellular redox homeostasis and the regulation of the cell cycle. However, the regulation of these metabolite pools during exponential growth and their precise functions in the cell cycle remain to be characterized. The present analysis of the abundance of ascorbate, glutathione, and pyridine nucleotides during exponential growth of Arabidopsis cells in culture provides evidence for the differential regulation of each of these redox pools. Ascorbate was most abundant early in the growth cycle, but glutathione was low at this point. The cellular ascorbate to dehydroascorbate and reduced glutathione （GSH） to glutathione disulphide ratios were high and constant but the pyridine nucleotide pools were largely oxidized over the period of exponential growth and only became more reduced once growth had ceased. The glutathione pool increased in parallel with poly （ADP-ribose） polymerase （PARP） activities and with increases in the abundance of PARP1 and PARP2 mRNAs at a time of high cell cycle activity as indicated by transcriptome information. Marked changes in the intracellular partitioning of GSH between the cytoplasm and nucleus were observed. Extension of the exponential growth phase by dilution or changing the media led to increases in the glutathione and nicotinamide adenine dinucleotide, oxidized form （NAD）-plus-nicotinamide adenine dinucleotide, reduced form （NADH） pools and to higher NAD/NADH ratios but the nicotinamide adenine dinucleotide phosphate, oxidized form （NADP）-plus-nicotinamide adenine dinucleotide phosphate, reduced form （NADPH） pool sizes, and NAPD/NADPH ratios were much less affected. The ascorbate, glutathione, and pyridine nucleotide pools and PARP activity decreased before the exponential growth phase ended. We conclude that there are marked changes in intracellular redox state during the growth cycle but that redox homeostasis is maintained by interplay of the major redox pyridine nucleotides, glutathione, and ascorbate pools. The correlation between PARP expression and activity and GSH accumulation and the finding that GSH can be recruited to the nucleus suggest a relationship between redox regulation and nuclear enzyme activity.展开更多
基金supported by the grants from Zhejiang Gongshang University,China(Grant No.1110KU111008)the National Science Foundation of China(Grant No.20977084)
文摘A rice cadmium （Cd） sensitive mutant cadB-1 was obtained using Agrobacterium tumefaciens mediated system.After exposure of cadB-1 and wild type （WT） rice seedlings to a range of Cd concentrations for 10 d,Cd accumulated to higher levels in roots,stems and leaves of both cadB-1 and WT with increasing external Cd concentrations,and the inhibition of seedling growth in cadB-1 was more serious than in WT.Hydrogen peroxide accumulation was higher in leaves and roots of cadB-1.The ratios of reduced glutathione （GSH）/oxidized glutathione （GSSG）,ascorbate （ASC）/dehydroascorbate （DHA） and reduced nicotinamide adenine dinucleotide phosphate （NADPH）/oxidized nicotinamide adenine dinucleotide phosphate （NADP＋） were lower in cadB-1 than in WT both in leaves and roots under high Cd levels.The activities of ascorbate peroxidase （APX）,glutathione peroxidase （GR）,dehydroascorbate reductase （DHAR） and monodehydroascorbate reductase （MDHAR） were also lower in cadB-1 than in WT both in leaves and roots under the treatment of high levels of Cd.Our results suggest that under Cd stress,the ASC-GSH cycle was more seriously inhibited in cadB-1 than in WT,indicating that the mutant cadB-1 is less able to scavenge reactive oxygen species and sensitive to Cd.
基金Support by Natural Science Foundation of Shandong Province,China(ZR2017LEE023,BS2014HZ011)Scientific Research Starting Fund of Binzhou University(2013Y16)
文摘Nonylphenol( NP) is a stable metabolic product of nonylphenol ethoxylates,which is widely used as an industrial surfactant. NP has been classified as an endocrine disrupter,and its toxicity to organisms can be biomagnified through the food chain. As compared with the endocrine disrupting effect,the toxicity of NP to organisms has not been studied intensively,and the toxicity mechanisms have often been ignored. In the present study,Microcystis aeruginosa,a freshwater alga belonging to the first level of the trophic chain,was chosen to detect the toxicity of NP. The mechanisms of toxicity mediated by the AsA-GSH cycle were explored. The acute toxicity of NP to M. aeruginosa within 96 h was studied and an EC_(50) concentration of 3. 45 mg/L was found. Further,the results showed that the toxicity of NP increased with the increase in concentration and exposure time. As compared with that in the control,the APX and MDHAR activities mostly increased,whereas DHAR activity fluctuated.However,the AsA content elevated at first,but decreased significantly after 72 h. For the GSH system,GR activity was always higher than that in the control. Nevertheless,the reduced GSH content was mostly inhibited. Therefore,the performance of AsA-GSH antioxidant defense system could explain the results of NP toxicity: the enzyme activities and antioxidant molecules increased initially,but an overall decline appeared after exposure for 24 h. This research is helpful for estimating the toxicity of NP integrally and improves people's understanding of mechanisms of NP toxicity in algae.
基金supported by Zhejiang Provincial Natural Science Foundation of China(Grant Nos.LY18C030001 and LQ13C030002)National Natural Science Foundation of China(Grant No.41776097)+4 种基金Special Funding for Research of National Oceanic Public Service Industry of China(Grant No.201505028)National Science and Technology Basic Resources Survey Special of China(Grant No.2017FY100700)Zhejiang Province Science and Technology Plan Project of China(Grant Nos.2013C25096 and2014F50003)Zhejiang Province Foundation of the Nonprofit Technology Research Projects of China(Grant No.2015C33227)Wenzhou Municipal Science and Technology Plan Project of China(Grant Nos.N20140046,N20170008 and S20160004)
文摘As the most northerly mangrove species in China, Kandelia obovata may undergo extreme cold event stress. Enhancing the cold tolerance of this species is crucial to its successful afforestation. This study aimed to determine the resistance of K. obovata seedlings to low temperature stress by cold acclimation and to explain the mechanisms for alleviating cold injury. To understand these mechanisms, seedlings that were acclimatized and not acclimatized were exposed to 5℃/- 2℃(day/night)for 48 h.Results showed that low temperature stress reduced leaf photosynthesis of non-acclimatized seedlings by inducing oxidative stress and structural damage to chloroplasts. These phenomena were shown by increasing levels of malondialdehyde (MDA), O2-and H2O2, as well as decreasing enzyme activities in the ascorbate–glutathione (AsA-GSH) cycle. However, cold-acclimatized seedlings had improved photosynthetic rates and efficiency of photosystem II (PSII) under low temperature stress. Compared with non-acclimatized seedlings, leaves of coldacclimatized seedlings under low temperature stress for 48 h exhibited higher anti-oxidative enzyme activities, lower levels of O2^- and H2O2, less damage to chloroplast structure, and removed 33.7% of MDA at low temperature stress for 48 h. The data indicate that cold acclimation enhances photosynthetic capacity by effectively regulating activation in the PSII electron transport and the AsA–GSH cycle to scavenge excess ROS in chloroplasts, while the latter is more important.
文摘Glutathione peroxidase, the first example of selenoproteins identified in mammals, was subjected to force field calculations and molecular dynamics in order to enable a clearer comprehension of enzymatic selenium catalysis. Starting from the established X-ray structure of bovine GPX, all kinetically defined intermediates and enzyme substrate complexes were modelled. The models thus obtained support the hypothesis that the essential steps of the catalysis are three distinct redox changes of the active site selenium which, in the ground state, presents itself at the surface of selenoperoxidases as the center of a characteristic triad built by selenocysteine, glutarnine and tryptophan. In GPX, four arginine residues and a lysine residue provide an electrostatic architecture which, in each reductive step, directs the donor substrate GSH towards the catalytic center in such a way that 1ts sulfhydryl group must react with the selenium moiety. To this end, different equally efficient modes of substrate binding appear possible. The models are consistent with substrate specificity data, kinetic pattern and other functional characteristics of the enzyme. Comparison of molecular models of GPX with those of other members of the GPX superfamily reveals that the cosubstrate binding mechanisrns are unique for the classical type of cytosolic glutathione peroxidases but cannot operate e. g. in plasma GPX and phospholipid hydroperoxide GPX. The structural differences between the selenoperoxidases, shown to be relevant to their specificities, are discussed in terms of functional diversification within the GPX superfamily
文摘This research provides, to the authors’ knowledge, the first integrative model of oxidative stress and C1 metabolism in plants. Increased oxidative stress can cause irreversible damage to photosynthetic components and is harmful to plants. Perturbations at the genetic level may increase oxidative stress and upregulate antioxidant systems in plants. One of the key mechanisms involved in oxidative stress regulation is the ascorbate-glutathione cycle which operates in chloroplasts as well as the mitochondria and is responsible for removal of reactive oxygen species (ROS) generated during photosynthetic operations and respiration. In this research, the complexity of molecular pathway systems of oxidative stress is modeled and then integrated with a previously developed in silico model of C1 metabolism system. This molecular systems integration provides two important results: 1) demonstration of the scalability of the CytoSolve®?Collaboratory™, a computational systems biology platform that allows for modular integration of molecular pathway models, by coupling the in silico model of oxidative stress with the in silico model of C1 metabolism, and 2) derivation of new insights on the effects of oxidative stress on C1 metabolism relative to formaldehyde (HCHO), a toxic molecule, and glutathione (GSH), an important indicator of oxidative homeostasis in living systems. Previous in silico modeling of C1 metabolism, without oxidative stress, observed complete removal of formaldehyde via formaldehyde detoxification pathway and no change in glutathione concentrations. The results from this research of integrative oxidative stress with C1 metabolism, however, demonstrate significant upregulation of formaldehyde concentrations, with concomitant downregulation and depletion of glutathione. Sensitivity analysis indicates that kGSH-HCHO, the rate constant of GSH-HCHO binding, VSHMT, the rate of formation of sarcosine from glycine, and , the rate of superoxide formation significantly affect formaldehyde homeostasis in the C1 metabolism. Future research may employ this integrative model to explore which conditions initiate oxidative stress and the resultant upregulation and downregulation of formaldehyde and glutathione.
文摘To explore whether glutathione regulates diapause determination and termina tion in the bivoltine silkworm Bombyx mori, we monitored the changes in glutathione redox cycle in the ovary of both diapanse and nondiapauseegg producers, as well as those in dia pause eggs incubated at different temperatures. The activity ofthioredoxin reductase （TrxR） was detected in ovaries but not in eggs, while neither ovaries nor eggs showed activity of glutathione peroxidase. A lower reduced glutathione/oxidized glutathione （GSH/GSSG） ratio was observed in the ovary of diapauseegg producers, due to weaker reduction of oxidized glutathione （GSSG） to the reduced glutathione （GSH） catalyzed by glutathione reductase （GR） and TrxR. This indicates an oxidative shift in the glutathione redox cy cle during diapause determination. Compared with the 25℃treated diapause eggs, the 5℃treated diapause eggs showed lower GSH/GSSG ratio, a result of stronger oxidation of GSH catalyzed by thioredoxin peroxidase and weaker reduction of GSSG catalyzed by GR. Our study demonstrated the important regulatory role of glutathione in diapause determination and termination of the bivoltine silkworm.
文摘Pyridine nucleotides, ascorbate and glutathione are major redox metabolites in plant cells, with specific roles in cellular redox homeostasis and the regulation of the cell cycle. However, the regulation of these metabolite pools during exponential growth and their precise functions in the cell cycle remain to be characterized. The present analysis of the abundance of ascorbate, glutathione, and pyridine nucleotides during exponential growth of Arabidopsis cells in culture provides evidence for the differential regulation of each of these redox pools. Ascorbate was most abundant early in the growth cycle, but glutathione was low at this point. The cellular ascorbate to dehydroascorbate and reduced glutathione （GSH） to glutathione disulphide ratios were high and constant but the pyridine nucleotide pools were largely oxidized over the period of exponential growth and only became more reduced once growth had ceased. The glutathione pool increased in parallel with poly （ADP-ribose） polymerase （PARP） activities and with increases in the abundance of PARP1 and PARP2 mRNAs at a time of high cell cycle activity as indicated by transcriptome information. Marked changes in the intracellular partitioning of GSH between the cytoplasm and nucleus were observed. Extension of the exponential growth phase by dilution or changing the media led to increases in the glutathione and nicotinamide adenine dinucleotide, oxidized form （NAD）-plus-nicotinamide adenine dinucleotide, reduced form （NADH） pools and to higher NAD/NADH ratios but the nicotinamide adenine dinucleotide phosphate, oxidized form （NADP）-plus-nicotinamide adenine dinucleotide phosphate, reduced form （NADPH） pool sizes, and NAPD/NADPH ratios were much less affected. The ascorbate, glutathione, and pyridine nucleotide pools and PARP activity decreased before the exponential growth phase ended. We conclude that there are marked changes in intracellular redox state during the growth cycle but that redox homeostasis is maintained by interplay of the major redox pyridine nucleotides, glutathione, and ascorbate pools. The correlation between PARP expression and activity and GSH accumulation and the finding that GSH can be recruited to the nucleus suggest a relationship between redox regulation and nuclear enzyme activity.
