Ascorbate peroxidase. A prominent membrane protein in oilseed glyoxysomes.

The glyoxysomes of growing oilseed seedlings produce H2O2, a reactive oxygen species, during the beta-oxidation of lipids stored in the cotyledons. An expression library of dark-grown cotton (Gossypium hirsutm L.) cotyledons was screened with antibodies that recognized a 31-kD glyoxysomal membrane polypeptide. A full-length cDNA clone (1258 bp) was isolated that encodes a 32-kD subunit of ascorbate peroxidase (APX) with a single, putative membrane-spanning region near the C-terminal end of the polypeptide. Internal amino acid sequence analysis of the cotton 31-kD polypeptide verified that this clone encoded this protein. This enzyme, designated gmAPX, was immunocytochemically and enzymatically localized to the glyoxysomal membrane in cotton cotyledons. The activity of monodehydroascorbate reductase, a protein that reduces monodehydroascorbate to ascorbate with NADH, also was detected in these membranes. The co-localization of gmAPX and monodehydroascorbate reductase within the glyoxysomal membrane likely reflects an essential pathway for scavenging reactive oxygen species and also provides a mechanism to regenerate NAD+ for the continued operation of the glyoxylate cycle and beta-oxidation of fatty acids. Immunological cross-reactivity of 30- to 32-kD proteins in glyoxysomal membranes of cucumber, sunflower, castor bean, and cotton indicate that gmAPX is common among oilseed species.     

Development of Endoplasmic Reticulum and Glyoxysomal Membrane Redox Activities during Castor Bean Germination

Redox activities, NADH:ferricyanide reductase, NAD(P)H:cytochrome reductases, and NADH:ascorbate free-radical reductase, are present in endoplasmic reticulum (ER) and glyoxysomal membranes from the endosperm of germinating castor bean (Ricinus comminus L. var Hale). The development of these functions was followed in glyoxysomes and ER isolated on sucrose gradients from castor bean endosperm daily from 0 through 6 days of germination. On a per seed basis, glyoxysomal and ER protein, glyoxysomal and ER membrane redox enzyme activities, and glyoxylate cycle activities peaked at day 4 as did the ER membrane content of cytochrome P-450. NADH:ferricyanide reductase was present in glyoxysomes and ER isolated from dry seed. This activity increased only about twofold in glyoxysomes and threefold in ER during germination relative to the amount of protein in the respective fractions. The other reductases, NADH:cytochrome reductase and NADH:ascorbate free-radical reductase, increased about 10-fold in the ER relative to protein up to 4 to 5 days, then declined. NADPH:cytochrome reductase reached maximum activity relative to protein at day 2 in both organelles. The increases in redox activities during germination indicate that the membranes of the ER and glyoxysome are being enriched with redox proteins during their development. The development of redox functions in glyoxysomes was found to be coordinated with development of the glyoxylate cycle.     

Ascorbate free-radical reduction by glyoxysomal membranes

Glyoxysomal membranes from germinating castor bean (Ricinus communis L. cv Hale) endosperm contain an NADH dehydrogenase. This enzyme can utilize extraorganellar ascorbate free-radical as a substrate and can oxidize NADH at a rate which can support intraglyoxysomal demand for NAD⁺. NADH:ascorbate free-radical reductase was found to be membrane-associated, and the activity remained in the membrane fraction after lysis of glyoxysomes by osmotic shock, followed by pelleting of the membranes. In whole glyoxysomes, NADH:ascorbate free-radical reductase, like NADH:ferricyanide reductase and unlike NADH:cytochrome c reductase, was insensitive to trypsin and was not inactivated by Triton X-100 detergent. These results suggest that ascorbate free-radical is reduced by the same component which reduces ferricyanide in the glyoxysomal membrane redox system. NADH:ascorbate free-radical reductase comigrated with NADH:ferricyanide and cytochrome c reductases when glyoxy-somal membranes were solubilized with detergent and subjected to rate-zonal centrifugation. The results suggest that ascorbate free-radical, when reduced to ascorbate by membrane redox system, could serve as a link between glyoxysomal metabolism and other cellular activities.     

Steady-state kinetic mechanism of recombinant avocado ACC oxidase: initial velocity and inhibitor studies

The gaseous plant hormone ethylene modulates a wide range of biological processes, including fruit ripening. It is synthesized by the ascorbate-dependent oxidation of 1-aminocyclopropyl-1-carboxylate (ACC), a reaction catalyzed by ACC oxidase. Recombinant avocado (Persea americana) ACC oxidase was expressed in Escherichia coli and purified in milligram quantities, resulting in high levels of ACC oxidase protein and enzyme activity. An optimized assay for the purified enzyme was developed that takes into account the inherent complexities of the assay system. Fe(II) and ascorbic acid form a binary complex that is not the true substrate for the reaction and enhances the degree of ascorbic acid substrate inhibition. The K(d) value for Fe(II) (40 nM, free species) and the K(m)'s for ascorbic acid (2.1 mM), ACC (62 microM), and O(2) (4 microM) were determined. Fe(II) and ACC exhibit substrate inhibition, and a second metal binding site is suggested. Initial velocity measurements and inhibitor studies were used to resolve the kinetic mechanism through the final substrate binding step. Fe(II) binding is followed by either ascorbate or ACC binding, with ascorbate being preferred. This is followed by the ordered addition of molecular oxygen and the last substrate, leading to the formation of the catalytically competent complex. Both Fe(II) and O(2) are in thermodynamic equilibrium with their enzyme forms. The binding of a second molecule of ascorbic acid or ACC leads to significant substrate inhibition. ACC and ascorbate analogues were used to confirm the kinetic mechanism and to identify important determinants of substrate binding.     

Effect of abscisic acid on active oxygen species, antioxidative defence system and oxidative damage in leaves of maize seedlings.

Leaves of maize (Zea mays L.) seedlings were supplied with different concentrations of abscisic acid (ABA). Its effects on the levels of superoxide radical (O(2)(-)), hydrogen peroxide (H(2)O(2)) and the content of catalytic Fe, the activities of several antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR), the contents of several non-enzymatic antioxidants such as ascorbate (ASC), reduced glutathione (GSH), alpha-tocopherol (alpha-TOC) and carotenoid (CAR), and the degrees of the oxidative damage to the membrane lipids and proteins were examined. Treatment with 10 and 100 microM ABA significantly increased the levels of O(2)(-) and H(2)O(2), followed by an increase in activities of SOD, CAT, APX and GR, and the contents of ASC, GSH, alpha-TOC and CAR in a dose- and time-dependent pattern in leaves of maize seedlings. An oxidative damage expressed as lipid peroxidation, protein oxidation, and plasma membrane leakage did not occur except for a slight increase with 100 microM ABA treatment for 24 h. Treatment with 1,000 microM ABA led to a more abundant generation of O(2)(-) and H(2)O(2) and a significant increase in the content of catalytic Fe, which is critical for H(2)O(2)-dependent hydroxyl radical production. The activities of these antioxidative enzymes and the contents of alpha-TOC and CAR were still maintained at a higher level, but no longer further enhanced when compared with the treatment of 100 microM ABA. The contents of ASC and GSH had no changes in leaves treated with 1,000 microM ABA. These results indicate that treatment with low concentrations of ABA (10 to 100 microM) induced an antioxidative defence response against oxidative damage, but a high concentration of ABA (1,000 microM) induced an excessive generation of AOS and led to an oxidative damage in plant cells.     

Kinetics of the Anti-oxidant Response to Salinity in the Halophyte Cakile maritima

The effects of NaCl stress on the activity of anti-oxidant enzymes （superoxide dismutase, catalase （CAT）, peroxidase （POD）, ascorbate peroxidase （APX）, monodehydroascorbate reductase, dehydroascorbate reductase （DHAR）, and glutathione reductase （GR））, anti-oxidant molecules （ascorbate and glutathione）, and parameters of oxidative stress （malondialdehyde （MDA）, electrolyte leakage, and H2O2 concentrations） were investigated in Cakile maritima, a halophyte frequent along the Tunisian seashore. Seedlings were grown in the presence of salt （100, 200, and 400 mmol/L NaCl）. Plants were harvested periodically over 20 days. Growth was maximal in the presence of 0-100 mmol/L NaCl. At 400 mmol/L NaCl, growth decreased significantly. The salt tolerance of C. maritima, at moderate salinities, was associated with the lowest values of the parameters indicative of oxidative stress, namely the highest activities of POD, CAT, APX, DHAR, and GR and high tissue content of ascorbate and glutathione. However, prolonged exposure to high salinity resulted in a decrease in anti-oxidant activities and high MDA content, electrolyte leakage, and H2O2 concentrations. These results suggest that anti-oxidant systems participate in the tolerance of C. maritima to moderate salinities.     

外源一氧化氮对氯化钠处理下秋茄幼苗抗氧化系统的调节效应

研究了外源一氧化氮(NO)供体硝普钠(SNP)对NaCl处理下红树植物秋茄(Kan-deliacandel)幼苗叶片中抗氧化酶活性、抗氧化物质及脯氨酸含量的影响。结果表明:NaCl处理下,秋茄幼苗叶片中超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)等4种活性氧清除酶的活性均受到明显抑制(P<0.05),SNP可以不同程度地恢复SOD、POD、CAT的活性,但对APX活性影响不大;SNP提高谷胱甘肽(GSH)及类胡萝卜素(Car)的含量,促进脯氨酸含量的上升,显著降低叶片中过氧化氢(H2O2)和丙二醛(MDA)的累积。表明外源NO可以缓解NaCl处理诱导的秋茄幼苗叶片氧化损伤,降低膜脂过氧化水平,有利于秋茄适应盐生环境。     

Ascorbic acid regulation of norepinephrine biosynthesis in isolated chromaffin granules from bovine adrenal medulla

The effect of ascorbic acid on the conversion of dopamine to norepinephrine was investigated in isolated chromaffin granules from bovine adrenal medulla. Ascorbic acid was shown to double the rate of [3H]norepinephrine formation from [3H]dopamine, despite no demonstrable accumulation of ascorbic acid into chromaffin granules. The enhancement of norepinephrine biosynthesis by ascorbic acid was dependent on the external concentrations of dopamine and ascorbate. The apparent Km of the dopamine beta-hydroxylation system for external dopamine was approximately 20 microM in the presence or absence of ascorbic acid. However, the apparent maximum velocity of norepinephrine formation was nearly doubled in the presence of ascorbic acid. By contrast, the apparent Km and Vmax of dopamine uptake into chromaffin granules were not affected by ascorbic acid. Norepinephrine formation was increased by ascorbic acid when the concentration of ascorbate was 200 microM or higher; a concentration of 2 mM appeared to induce the maximal effect under the experimental conditions used here. The effect of ascorbic acid on conversion of dopamine to norepinephrine required Mg-ATP-dependent dopamine uptake into chromaffin granules. In contrast to ascorbic acid, other reducing agents such as NADH, glutathione, and homocysteine were unable to enhance norepinephrine biosynthesis. These data suggest that ascorbic acid provides reducing equivalents for hydroxylation of dopamine despite the lack of ascorbate accumulation into chromaffin granules. These findings imply the functional existence of an electron carrier system in the chromaffin granule which transfers electrons from external ascorbic acid for subsequent intragranular norepinephrine biosynthesis.     

Role of Antioxidant Systems in Wheat Genotypes Tolerance to Water Stress

The role of plant antioxidant systems in stress tolerance was studied in leaves of three contrasting wheat genotypes. Drought imposed at two different stages after anthesis resulted in an increase in H2O2 accumulation and lipid peroxidation and decrease in ascorbic acid content. Antioxidant enzymes like superoxide dismutase, ascorbate peroxidase and catalase significantly increased under water stress. Drought tolerant genotype C 306 which had highest ascorbate peroxidase and catalase activity and ascorbic acid content also showed lowest H2O2 accumulation and lipid peroxidation (malondialdehyde content) under water stress in comparison to susceptible genotype HD 2329 which showed lowest antioxidant enzyme activity and ascorbic acid content and highest H2O2 content and lipid peroxidation. HD 2285 which is tolerant to high temperature during grain filling period showed intermediate behaviour. Superoxide dismutase activity, however, did not show significant differences among the genotypes under irrigated as well as water stress condition. It seems that H2O2 scavenging systems as represented by ascorbate peroxidase and catalase are more important in imparting tolerance against drought induced oxidative stress than superoxide dismutase alone. This revised version was published online in July 2006 with corrections to the Cover Date.     

Ascorbate activates soluble guanylate cyclase via H2O2-metabolism by catalase

Conditions necessary for the activation by ascorbic acid of soluble guanylate cyclase purified from bovine lung have been examined. Ascorbic acid (0.1-10 mM) did not directly activate the enzyme, nonetheless, pronounced activation by ascorbate (3-10 mM) was observed in incubation mixtures containing 1 microM bovine liver catalase. Superoxide dismutase (SOD) and mannitol did not affect the catalase-dependent activation of guanylate cyclase elicited by ascorbate, suggesting that superoxide anion and hydroxyl radical were not mediating the activation of the enzyme. However, SOD enhanced the relatively low level activation of the enzyme elicited by catalase in the absence of added ascorbate. Pronounced inhibition (both with and without added ascorbate) was observed of catalase-dependent activation of guanylate cyclase by either ethanol (100 mM) or a fungal catalase preparation. Neither ethanol nor fungal catalase inhibited activation of guanylate cyclase by S-nitrosyl-N-acetyl-penicillamine (SNAP), a source of the nitric oxide free radical. These observations indicate that autoxidation of ascorbic acid or thiols present with the guanylate cyclase preparation leads to generation of H2O2, and its metabolism by bovine liver catalase mediates the concomitant activation of guanylate cyclase. The mechanism of activation appears to be associated with the presence of Compound I of catalase and to be inhibited by superoxide anion.     

Determination of ascorbic acid with immobilized green zucchini ascorbate oxidase

Ascorbate oxidase from zucchini squash was immobilized onto CH-Sepharose via carbodiimide. The properties of the immobilized enzyme were found to be similar to those of the free ascorbate oxidase. The immobilized enzyme was utilized in a flow-through system equipped with a polarographic detector which monitors the oxygen depletion due to the reaction ascorbic acid + 1/2 O2----dehydroascorbic acid + H2O. This method, the response of which is linear between 3 X 10(-7) and 5 X 10(-4) M ascorbate, was utilized to measure the ascorbic acid in biological samples such as human plasma and fruit juices at a rate of about 60 determinations every hour with a standard deviation lower than 5%.     

臭氧浓度升高对油松抗氧化系统活性的影响

以生长在开顶箱内的油松为试材,对高浓度臭氧(80 nmol·mol^-1)条件下油松(Pinus tabulaeformis)针叶中超氧阴离子自由基(O₂^·)产生速率、过氧化氢(H₂O₂)含量、超氧化物歧化酶(SOD)、抗坏血酸过氧化物酶、脱氢抗坏血酸还原酶、单脱氢抗坏血酸还原酶、谷胱甘肽还原酶活性与抗坏血酸(ASA)含量进行测定.结果表明:高浓度臭氧使O₂^·产生速率提高,H₂O₂ 和MDA含量增加.ASA含量与SOD、抗坏血酸过氧化物酶、脱氢抗坏血酸还原酶、单脱氢抗坏血酸还原酶、谷胱甘肽还原酶活性在高浓度臭氧熏蒸的前期升高,随后下降并低于对照.说明生长季前期,油松抗氧化系统对高浓度臭氧存在适应性反应,但不能抵抗长期臭氧胁迫带来的氧化伤害.     

环境胁迫和抗坏血酸的氧化还原状态

为了解环境胁迫对植物体中抗坏血酸含量及氧化还原状态的影响,以不同强度的冰冻和干旱两种胁迫为例,研究了它们对沈阳几种针叶树离体叶抗坏血酸、脱氢抗坏血酸含量以及抗坏血酸-谷胱甘肽循环中4种酶活性的影响.结果表明,两种胁迫达到一定强度后,都能使还原态抗坏血酸含量下降而使脱氢抗坏血酸含量上升.冰冻使抗坏血酸过氧化酶和单脱氢抗坏血酸还原酶活性下降.轻度失水使这两种酶活性上升,失水加重后转而趋于下降.脱氢抗坏血酸还原酶和谷胱甘肽还原酶活性对两种胁迫反应均不如前两种酶敏感.结合以前的研究结果,认为这一H2O2清除系统在导致驯化(acclimation)的轻度胁迫作用下可以得到加强,而当胁迫强度过大时则其清除能力下降并使组织受到伤害.文中还报告了沈阳几种针叶树抗寒性和针叶中抗坏血酸含量及上述4种酶活性之间的相关关系.     

环境胁迫和抗坏血酸的氧化还原状态

为了解环境胁迫对植物体中抗坏血酸含量及氧化还原状态的影响，以不同强度的冰冻和干旱两种胁迫为例，研究了它们对沈阳几种针叶树离体叶抗坏血酸、脱氢抗坏血酸含量以及抗坏血酸-谷胱甘肽循环中4种酶活性的影响。结果表明，两种胁迫达到一定强度后，都能使还原态抗坏血酸含量下降而使脱氢抗坏血酸含量上升。冰冻使抗坏血酸过氧化酶和单脱氢抗坏血酸还原酶活性下降。轻度失水使这两种酶活性上升，失水加重后转而趋于下降。脱氢抗坏血酸还原酶和谷胱甘肽还原酶活性对两种胁迫反应均不如前两种酶敏感。结合以前的研究结果，认为这一H2O2清除系统在导致驯化(acclimation)的轻度胁迫作用下可以得到加强，而当胁迫强度过大时则其清除能力下降并使组织受到伤害。文中还报告了沈阳几种针叶树抗寒性和针叶中抗坏血酸含量及上述4种酶活性之间的相关关系。     

Silicon-mediated alleviation of Mn toxicity in Cucumis sativus in relation to activities of superoxide dismutase and ascorbate peroxidase

The effects of exogenous silicon (Si) on plant growth, activities of superoxide dismutase (SOD), guaiacol peroxidase (GPX), ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), glutathione reductase (GR) and catalase, and concentrations of ascorbate and glutathione were investigated in cucumber (Cucumis sativus L.) plants treated with excess manganese (Mn) (600 microM). Compared with the treatment of normal Mn (10 microM), excess Mn significantly increased H2O2 concentration and lipid peroxidation indicated by accumulation of thiobarbituric acid reactive substances. The leaves showed apparent symptoms of Mn toxicity and the plant growth was significantly inhibited by excess Mn. The addition of Si significantly decreased lipid peroxidation caused by excess Mn, inhibited the appearance of Mn toxicity symptoms, and improved plant growth. This alleviation of Mn toxicity by Si was related to a significant increase in the activities of SOD, APX, DHAR and GR and the concentrations of ascorbate and glutathione.     

Recycling of the ascorbate free radical by human erythrocyte membranes.

Reduction of the ascorbate free radical (AFR) at the plasma membrane provides an efficient mechanism to preserve the vitamin in a location where it can recycle alpha-tocopherol and thus prevent lipid peroxidation. Erythrocyte ghost membranes have been shown to oxidize NADH in the presence of the AFR. We report that this activity derives from an AFR reductase because it spares ascorbate from oxidation by ascorbate oxidase, and because ghost membranes decrease steady-state concentrations of the AFR in a protein- and NADH-dependent manner. The AFR reductase has a high apparent affinity for both NADH and the AFR (< 2 microM). When measured in open ghosts, the reductase is comprised of an inner membrane activity (both substrate sites on the cytosolic membrane face) and a trans-membrane activity that mediates extracellular AFR reduction using intracellular NADH. However, the trans-membrane activity constitutes only about 12% of the total measured in ghosts. Ghost AFR reductase activity can also be differentiated from NADH-dependent ferricyanide reductase(s) by its sensitivity to the detergent Triton X-100 and insensitivity to enzymatic digestion with cathepsin D. This NADH-dependent AFR reductase could serve to recycle ascorbic acid at a crucial site on the inner face of the plasma membrane.     

Inhibition of human leukocyte 3-hydroxy-3-methylglutaryl coenzyme A reductase activity by ascorbic acid. An effect mediated by the free radical monodehydroascorbate

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity in microsomes isolated from cultured lymphoid (IM-9) cells or freshly isolated human leukocytes was markedly decreased by either ascorbic acid or its oxidized derivative, dehydroascorbate. Inhibition of IM-9 leukocyte HMG-CoA reductase activity was log linear between 0.01 and 10 mM ascorbic acid (25 and 81% inhibition, respectively) and 0.1 and 10 mM dehydroascorbate (5 and 75% inhibition, respectively). Inhibition was noncompetitive with respect to HMG-CoA (Km = 10.2 microM (RS); ascorbic acid, Ki = 6.4 mM; dehydroascorbate, Ki = 15 mM) and competitive with respect to NADPH (Km = 16.3 microM; acetic acid, Ki = 6.3 mM; dehydroascorbate, Ki = 3.1 mM). Ascorbic acid and dehydroascorbate are interconverted through the free radical intermediate monodehydroascorbate. Reducing agents are required to convert dehydroascorbate to monodehydroascorbate, but prevent formation of the free radical from ascorbate. In microsomes from IM-9 cells, the reducing agent, dithiothreitol, abolished HMG-CoA reductase inhibition by ascorbate but enhanced inhibition by dehydroascorbate. In addition, the concentration of monodehydroascorbate present in ascorbate solutions was directly proportional to the degree of HMG-CoA reductase inhibition by 1.0 mM ascorbate. Fifty per cent inhibition of enzyme activity occurred at a monodehydroascorbate concentration of 14 microM. These data indicate that monodehydroascorbate mediates inhibition of HMG-CoA reductase by both ascorbate and dehydroascorbate. This effect does not appear to be due to free radical-induced membrane lipid modification, however, since both ascorbate and dehydroascorbate inhibited the protease-solubilized, partially purified human liver enzyme. Since inhibition of HMG-CoA reductase occurs at physiological concentrations of ascorbic acid in the human leukocyte (0.2-1.72 mM), this vitamin may be important in the regulation of endogenous cholesterol synthesis in man.     

Rapid inactivation of chloroplastic ascorbate peroxidase is responsible for oxidative modification to Rubisco in tomato (Lycopersicon esculentum) under cadmium stress

To investigate the sensitive site of antioxidant systems in chloroplast under cadmium stress and its consequence on reactive oxygen species production and action, the sub-organellar localization of chloroplast superoxide dismutases (SOD,EC 1.15.1.1) and ascorbic peroxidase (APX, EC 1.11.1.11) isoenzymes and changes of enzymes activities under cadmium stress were investigated in tomato seedlings. Two APX isoforms, one thylakoid-bound and one stromal, were detected. Cd at 50 μM induced a moderate increase of SOD activities but a rapid inactivation of both APX isoenzymes. APX inactivation was mainly related to the decrease of ascorbate concentration, as supported by in vitro treatment of exogenous ascorbate and APX kinetic properties under Cd stress. H2O2 accumulation in chloroplast, as a consequence of APX inactivation,was associated with a 60% loss of Rubisco (EC 4.1.1.39) activity, which could be partially accounted for by a 10% loss of Rubisco content. Protein oxidation assay found that the Rubisco large subunit was the most prominent carbonylated protein; the level of carbonylated Rubisco large subunit increased fivefold after Cd exposure. Thiol groups in the Rubisco large subunit were oxidized, as indicated by non-reducing electrophoresis. Treating crude extract with H2O2 resulted in a similar pattern of protein oxidation and thiols oxidation with that observed in Cd-treated plants. Our study indicates that APXs in the chloroplast is a highly sensitive site of antioxidant systems under Cd stress, and the inactivation of APX could be mainly responsible for oxidative modification to Rubisco and subsequent decrease in its activity.     

The plasma membrane NADH oxidase of soybean has vitamin K(1) hydroquinone oxidase activity

Isolated plasma membrane vesicles and the plasma membrane NADH oxidase partially purified from soybean plasma membrane vesicles exhibited a cyanide-insensitive vitamin K(1) hydroquinone oxidase activity with isolated plasma membrane vesicles. Reduced vitamin K(1) (phylloquinol) was oxidized at a rate of about 10 nmol/min/mg protein as determined by reduced vitamin K(1) reduction or oxygen consumption. The K(m) for reduced K(1) was 350 microM. With the partially purified enzyme, reduced vitamin K(1) was oxidized at a rate of about 600 nmol/min/mg protein and the K(m) was 400 microM. When assayed in the presence of 1 mM KCN, activities of both plasma membrane vesicles and of the purified protein were stimulated (0.1 microM) or inhibited (0.1 mM) by the synthetic auxin growth factor 2, 4-dichlorophenoxyacetic acid. The findings suggest the potential participation of the plasma membrane NADH oxidase as a terminal oxidase of plasma membrane electron transport from cytosolic NAD(P)H via reduced vitamin K(1) to acceptors (molecular oxygen or protein disulfides) at the cell surface.     

Up-regulation of the leaf mitochondrial and peroxisomal antioxidative systems in response to salt-induced oxidative stress in the wild salt-tolerant tomato species Lycopersicon pennellii

The response of the antioxidative systems of leaf cell mitochondria and peroxisomes of the cultivated tomato Lycopersicon esculentum (Lem) and its wild salt-tolerant related species Lycopersicon pennellii (Lpa) to NaCl 100 mM stress was investigated. Salt-dependent oxidative stress was evident in Lem mitochondria as indicated by their raised levels of lipid peroxidation and H2O2 content whereas their reduced ascorbate and reduced glutathione contents decreased. Concomitantly, SOD activity decreased whereas APX and GPX activities remained at control level. In contrast, the mitochondria of salt-treated Lpa did not exhibit salt-induced oxidative stress. In their case salinity induced an increase in the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR) and glutathione-dependent peroxidase (GPX). Lpa peroxisomes exhibited increased SOD, APX, MDHAR and catalase activity and their lipid peroxidation and H2O2 levels were not affected by the salt treatment. The activities of all these enzymes remained at control level in peroxisomes of salt-treated Lem plants. The salt-induced increase in the antioxidant enzyme activities in the Lpa plants conferred cross-tolerance towards enhanced mitochondrial and peroxisomal reactive oxygen species production imposed by salicylhydroxamic acid (SHAM) and 3-amino-1,2,4-triazole (3-AT), respectively.