Cross-talk between calcium and reactive oxygen species originated from NADPH oxidase in abscisic acid-induced antioxidant defence in leaves of maize seedlings

The signal interactions between calcium (Ca2+) and reactive oxygen species (ROS) originated from plasma membrane NADPH oxidase in abscisic acid (ABA)-induced antioxidant defence were investigated in leaves of maize (Zea mays L.) seedlings. Treatment with ABA led to significant increases in the activity of plasma membrane NADPH oxidase, the production of leaf O2-, and the activities of several antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR). However, such increases were blocked by the pretreatment with Ca2+ chelator EGTA or Ca2+ channel blockers La3+ and verapamil, and NADPH oxidase inhibitors such as diphenylene iodonium (DPI), imidazole and pyridine. Treatment with Ca2+ also significantly induced the increases in NADPH oxidase activity, O2- production and the activities of antioxidant enzymes, and the increases were arrested by pretreatment with the NADPH oxidase inhibitors. Treatment with oxidative stress induced by paraquat, which generates O2-, led to the induction of antioxidant defence enzymes, and the up-regulation was suppressed by the pretreatment of Ca2+ chelator and Ca2+ channel blockers. Our data suggest that a cross-talk between Ca2+ and ROS originated from plasma membrane-bound NADPH oxidase is involved in the ABA signal transduction pathway leading to the induction of antioxidant enzyme activity, and Ca2+ functions upstream as well as downstream of ROS production in the signal transduction event in plants.     

Tomato tos1 mutation identifies a gene essential for osmotic tolerance and abscisic acid sensitivity

Osmotic stress severely limits plant growth and agricultural productivity. We have used mutagenesis to identify plant genes that are required for osmotic stress tolerance in tomato. As a result, we have isolated a novel mutant in tomato (tos1) caused by a single recessive nuclear mutation that is hypersensitive to general osmotic stress. Growth measurements demonstrated that the tos1 mutant is less sensitive to intracellular abscisic acid (ABA) and this decreased ABA sensitivity of tos1 is a basic cellular trait expressed by the mutant at all developmental stages analysed. It is not caused by a deficiency in the synthesis of ABA because the tos1 seedlings accumulated more ABA than the wild type (WT) after osmotic stress. In contrast, the tss2 tomato mutant, which is also hypersensitive to osmotic stress, is hypersensitive to exogenous ABA. Comparative analysis of tos1 and tss2 indicates that appropriate ABA perception and signalling is essential for osmotic tolerance.     

Abscisic acid negatively modulates plant defence against rice black‐streaked dwarf virus infection by suppressing the jasmonate pathway and regulating reactive oxygen species levels in rice

Abscisic acid (ABA) plays a multifaceted role in plant immunity and can either increase resistance or increase susceptibility to some bacterial and fungal pathogens depending on the pathosystem. ABA is also known to mediate plant defence to some viruses. In this study, the relationship between the ABA pathway and rice black‐streaked dwarf virus (RBSDV) was investigated in rice. The expression of ABA pathway genes was significantly reduced upon RBSDV infection. Application of exogenous hormones and various ABA pathway mutants revealed that the ABA pathway plays a negative role in rice defence against RBSDV. Exogenous hormone treatment and virus inoculation showed that ABA inhibits the jasmonate‐mediated resistance to RBSDV. ABA treatment also suppressed accumulation of reactive oxygen species by inducing the expression of superoxidase dismutases and catalases. Thus, ABA modulates the rice–RBSDV interaction by suppressing the jasmonate pathway and regulating reactive oxygen species levels. This is the first example of ABA increasing susceptibility to a plant virus.     

Effect of stationary magnetic field strengths of 150 and 200 mT on reactive oxygen species production in soybean

Our previous investigation reported the beneficial effect of pre-sowing magnetic treatment for improving germination parameters and biomass accumulation in soybean. In this study, soybean seeds treated with static magnetic fields of 150 and 200 mT for 1 h were evaluated for reactive oxygen species (ROS) and activity of antioxidant enzymes. Superoxide and hydroxyl radicals were measured in embryos and hypocotyls of germinating seeds by electron paramagnetic resonance spectroscopy and kinetics of superoxide production; hydrogen peroxide and antioxidant activities were estimated spectrophotometrically. Magnetic field treatment resulted in enhanced production of ROS mediated by cell wall peroxidase while ascorbic acid content, superoxide dismutase and ascorbate peroxidase activity decreased in the hypocotyl of germinating seeds. An increase in the cytosolic peroxidase activity indicated that this antioxidant enzyme had a vital role in scavenging the increased H(2)O(2) produced in seedlings from the magnetically treated seeds. Hence, these studies contribute to our first report on the biochemical basis of enhanced germination and seedling growth in magnetically treated seeds of soybean in relation to increased production of ROS.     

拟南芥CEO1基因在氯化镉诱导的氧化胁迫中的作用

以拟南芥ceo1、突变体为材料,研究CEO1（clone eight-one）在镉胁迫条件下作用的结果表明,与野生型植株相比,150μmol·L^-1的CdCl2处理10d后,拟南芥ceo1突变体表现为植株生长矮小,叶片卷曲发黄,根系短小。镉处理后,拟南芥突变体幼苗叶中H2O2的积累较多;镉处理1h后的突变体中抗坏血酸过氧化物酶（APX）活性明显上升,至2h时又开始下降,而镉处理2h后,野生型APX活性才开始增加。镉处理2h后的野生型的谷胱甘肽还原酶（GR）显著增加,而突变体无明显变化。两种类型拟南芥的超氧化物歧化酶（SOD）与过氧化氢酶（CAT）的活性没有明显差异。     

ITN1, a novel gene encoding an ankyrin-repeat protein that affects the ABA-mediated production of reactive oxygen species and is involved in salt-stress tolerance in Arabidopsis thaliana

Salt stress and abscisic acid (ABA) induce accumulation of reactive oxygen species (ROS) in plant cells. ROS not only act as second messengers for the activation of salt-stress responses, but also have deleterious effects on plant growth due to their cytotoxicity. Therefore, the timing and degree of activation of ROS-producing or ROS-scavenging enzymes must be tightly regulated under salt-stress conditions. We identified a novel locus of Arabidopsis, designated itn1 (increased tolerance to NaCl1), whose disruption leads to increased salt-stress tolerance in vegetative tissues. ITN1 encodes a transmembrane protein with an ankyrin-repeat motif that has been implicated in diverse cellular processes such as signal transduction. Comparative microarray analysis between wild-type and the itn1 mutant revealed that induction of genes encoding the ROS-producing NADPH oxidases (RBOHC and RBOHD) under salt-stress conditions was suppressed in the mutant. This suppression was accompanied by a corresponding reduction in ROS accumulation. The ABA-induced expression of RBOHC and RBOHD was also suppressed in the mutant, as was the case for RD29A, an ABA-inducible marker gene. However, the ABA-induced expression of another marker gene, RD22, was not impaired in the mutant. These results suggest that the itn1 mutation partially impairs ABA signaling pathways, possibly leading to the reduction in ROS accumulation under salt-stress conditions. We discuss the possible mechanisms underlying the salt-tolerant phenotype of the itn1 mutant.     

The effects of dopamine on antioxidant enzymes activities and reactive oxygen species levels in soybean roots

In the current work, we investigated the effects of dopamine, an neurotransmitter found in several plant species on antioxidant enzyme activities and ROS in soybean (Glycine max L. Merrill) roots. The effects of dopamine on SOD, CAT and POD activities, as well as H₂O₂, O₂^•−, melanin contents and lipid peroxidation were evaluated. Three-day-old seedlings were cultivated in half-strength Hoagland nutrient solution (pH 6.0), without or with 0.1 to 1.0 mM dopamine, in a growth chamber (25°C, 12 h photoperiod, irradiance of 280 μmol m⁻² s⁻¹) for 24 h. Significant increases in melanin content were observed. The levels of ROS and lipid peroxidation decreased at all concentrations of dopamine tested. The SOD activity increased significantly under the action of dopamine, while CT activity was inhibited and POD activity was unaffected. The results suggest a close relationship between a possible antioxidant activity of dopamine and melanin and activation of SOD, reducing the levels of ROS and damage on membranes of soybean roots.     

Nitric oxide interferes with plant photo-oxidative stress by detoxifying reactive oxygen species

Oxidative stress within chloroplasts is originated due to light‐dependent O₂ reduction. This may be exacerbated by bipyridinium herbicides, which act at photosystem I as artificial electron acceptors. Their oxidation produces a superoxide anion that further dismutates to H₂O₂ and then, by the Fenton reaction, H₂O₂ may be reduced to the hydroxyl radical (OH^?). Reactive oxygen species (ROS), when produced in high amounts, provoke severe damage to the plant cell. Herein it is reported that two nitric oxide (NO) donors, sodium nitroprusside (100 µm ) and S‐nitroso‐N‐acetylpenicillamine (200 µm ), greatly reduced lipid peroxidation and the protein loss caused by the application of a high dose of the bipyridinium herbicide diquat to potato leaf pieces or isolated chloroplasts. Nitric oxide donors also protected the RNA against oxidative damage. Photo‐oxidative toxicity was correlated with an increase in photosynthetic electron transport and ROS production, but the rate of electron transport was restored and the ROS free amount was markedly reduced in the presence of NO. The specific activity of superoxide dismutase was not affected by diquat or NO donors, whereas just a small increase in catalase activity was observed after 24 h of treatment. These results provide strong evidence that NO is a potent antioxidant in plants and that its action may, at least in part, be explained by its ability to directly scavenge ROS.     

Zooming into sub-organellar localization of reactive oxygen species in guard cell chloroplasts during abscisic acid and methyl jasmonate treatments

Regulation of stomata movements is crucial for plants ability to cope with their changing environment. Guard cell’s (GC) water potential directs water flux inside/outside this cell, which eventually is causing the stoma to open or close, respectively. Some of the osmolytes which accumulates in the GC cytoplasm and are known to play a role in stomata opening are sugars, arising from chloroplast starch degradation. During stomata closure, the accumulated osmolytes are removed from the GC cytoplasm. Surprisingly little is known about prevention of starch degradation and forming additional sugars which may interfere with osmotic changes that are necessary for correct closure of stomata.   One of the early events leading to stomata closure is production of reactive oxygen species (ROS) in various sub-cellular sites and organelles of the stoma. Here we report that ROS production during abscisic acid (ABA) and methyl jasmonate (MJ) stimuli in Arabidopsis GC chloroplasts were more than tripled. Moreover, ROS were detected on the sub-organelle level in compartments that are typically occupied by starch grains. This observation leads us to suspect that ROS function in that particular location is necessary for stomata closure. We therefore hypothesize that these ROS are involved in redox control that lead to the inactivation of starch degradation that takes place in these compartments, thus contributing to the stoma closure in an additional way.     

Changes in expression of genes encoding antioxidant enzymes, heme oxygenase-1, Bcl-2, and Bcl-xl and in level of reactive oxygen species in tumor cells resistant to doxorubicin

The relationship between expression of genes encoding key antioxidant enzymes, heme oxygenase-1, Bcl-2, and Bcl-xl and change in production of reactive oxygen species (ROS) resulting from development of resistance of cancer cells K562, MCF-7, and SKOV-3 to the prooxidant chemotherapeutic agent doxorubicin (DOX) has been studied. Significant increase in mRNA level and activity of Mn-superoxide dismutase (Mn-SOD), catalase, and selenium-dependent glutathione peroxidase-1 (GPx-1) and reduced ROS level was found in resistant K562/DOX and SKVLB cells. In contrast, no change in ROS level was observed in MCF-7/DOX cells in parallel with decrease in Mn-SOD and catalase mRNAs and corresponding activities concurrently with high increase in GPx-1 mRNA and activity. As a result of the development of resistance, a similarity was found between the change in ROS level and the change in ho-1 and bcl-2 gene expression, whereas elevation of bcl-xl gene expression was observed in all three types of resistant cells. Particular features of development of adaptive antioxidant response as well as redox-dependent change in bcl-2 gene expression under formation of DOX resistance of cancer cells of different genesis are discussed.     

Analysis of reactive oxygen species and antioxidant defenses in complex I deficient patients revealed a specific increase in superoxide dismutase activity

The mechanism of free radical production by complex I deficiency is ill-defined, although it is of significant contemporary interest. This study studied the ROS production and antioxidant defenses in children with mitochondrial NADH dehydrogenase deficiency. ROS production has remained significantly elevated in patients compared to controls. The expression of all antioxidant enzymes significantly increased at mRNA level. However, the enzyme activities did not correlate with high mRNA or protein expression. Only the activity of superoxide dismutase (SOD) was found to correlate with higher mRNA expression in patient derived cell lines. The activities of the enzymes such as glutathione peroxidase (GPx), Catalase (CAT) and glutathione-S-transferase (GST) were significantly reduced in patients (p<0.05 or p<0.01). Glutathione reductase (GR) activity and intracellular glutathione (GSH) levels were not changed. Decreased enzyme activities could be due to post-translational or oxidative modification of ROS scavenging enzymes. The information on the status of ROS and marking the alteration of ROS scavenging enzymes in peripheral lymphocytes or lymphoblast cell lines will provide a better way to design antioxidant therapies for such disorders.     

ABA-deficient (aba1) and ABA-insensitive (abi1-1, abi2-1) mutants of Arabidopsis have a wild-type stomatal response to humidity

In most plant species, a decrease in atmospheric humidity at the leaf surface triggers a decrease in stomatal conductance. While guard cells appear to respond to humidity‐induced changes in transpiration rate, as opposed to relative humidity or vapour pressure difference, the underlying cellular mechanisms for this response remain unknown. In the present set of experiments, abscisic acid (ABA)‐deficient (aba1) and ABA‐insensitive (abi1‐1 and abi2‐1) mutants of Arabidopsis thaliana were used to test the hypothesis that the humidity signal is transduced by changes in the flux or concentration of ABA delivered to the stomatal complex in the transpiration stream. In gas exchange experiments, stomatal conductance was as sensitive to changes in vapour pressure difference in aba1, abi1‐1 and abi2‐1 mutant plants as in wild‐type plants. These experiments appear to rule out an obligate role for either the concentration or flux of ABA or ABA conjugates as mediators of the guard cell response to atmospheric water potential. The results stand in contrast to the well‐established role of ABA in mediating guard cell responses to decreases in soil water potential.     

根系渗透胁迫时杨树光合作用光抑制与活性氧的关系

为更多地了解自然条件下活体叶片的光抑制,研究了渗透胁迫时杨树无性系幼苗叶片的光抑制与活性氧代谢的关系．结果表明,随胁迫时间的延长和胁迫强度的增大,杨树叶片O2^-生成加快,H2O2和丙二醛(MDA)含量增多,超氧物歧化酶(SOD)活性升高,过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性降低,活性氧代谢失衡,光合作用的光抑制加剧．用二乙基二硫代氨基甲酸铜盐抑制SOD活性,或用甲基紫精加速O2^-的生成,亦可使杨树叶片发生光抑制．渗透胁迫时杨树无性系幼苗清除H2O2能力降低,限制了叶片通过Mehler反应耗散过剩光能,防御光破坏作用的发挥;光抑制的发生与活性氧的积累有关．     

Interferon-gamma induces reactive oxygen species and endoplasmic reticulum stress at the hepatic apoptosis

Interferon-gamma (IFN-gamma) induces cell-cycle arrest and p53-independent apoptosis in primary cultured hepatocytes. However, the detailed mechanism, including regulating molecules, is still unclear. In this study, we found that IFN-gamma induced generation of reactive oxygen species (ROS) in primary hepatocytes and that pyrrolidinedithiocarbamate (PDTC), an anti-oxidant reagent, completely suppressed IFN-gamma-induced hepatic apoptosis. PDTC blocked apoptosis downstream from IRF-1 and upstream from caspase activation, suggesting that the generation of ROS occurred between these stages. However, IFN-gamma also induced the generation of ROS in IRF-1-deficient hepatocytes, cells insensitive to IFN-gamma-induced apoptosis. Moreover, a general cyclooxygenase (COX) inhibitor, indomethacin (but not the cyclooxygenase 2-specific inhibitor, NS-398) also inhibited the apoptosis without blocking the generation of ROS. Both PDTC and indomethacin also blocked IFN-gamma-induced release of cytochrome c from mitochondria. These results suggest that ROS are not the only or sufficient mediators of IFN-gamma-induced hepatic apoptosis. In contrast, we also found that IFN-gamma induced endoplasmic reticulum (ER) stress proteins, CHOP/GADD153 and caspase 12, in wild-type primary hepatocytes, but induced only caspase 12 and not CHOP/GADD153 protein in IRF-1-deficient hepatocytes. These results suggest that IFN-gamma induces ER stress in primary hepatocytes. Both the ROS and ER stress induced by IFN-gamma may be complementary mediators that induce apoptosis in primary hepatocytes.     

Seasonal changes in antioxidant defence system of liver and gills of Salmo trutta caspius, Salmo trutta labrax and Salmo trutta macrostigma

Seasonal changes in antioxidant enzyme activities (superoxide dismutase, SOD, EC 1.15.1.1; catalase, CAT, EC 1.11.1.16; glutathione peroxidase, GPx, EC 1.11.1.9; glutathione reductase, GR, EC 1.6.4.2; glucose-6-phosphate dehydrogenase, G6PD, EC 1.1.1.49 and glutathione S -transferase, GST, EC 1.5.1.18) and lipid peroxidation (LPO) levels of livers and gills of female Caspian trout Salmo trutta caspius , Black Sea trout Salmo trutta labrax and mountain trout Salmo trutta macrostigma were investigated. SOD, CAT, GPx, G6PD and GST activities were higher in liver compared to gills of all sub-species; concomitantly, the GR activity was also higher in the livers of S. t. caspius and S. t. labrax , but the reverse was seen in S. t. macrostigma . LPO levels were higher in the gills compared to the liver of all sub-species. There was no general trend in the seasonal changes in the gill antioxidant enzyme (AE) activities or LPO levels. Higher AE activities, however, were found in the liver of each sub-species during autumn, and this coincided with an increase in the gonado-somatic index.