H2O2 In Roots Research Articles

Impact of the auxin signaling inhibitor p-chlorophenoxyisobutyric acid on short-term Cd-induced hydrogen peroxide production and growth response in barley root tip

Short-term treatment (30min) of barley roots with a low 10μM Cd concentration induced significant H2O2 production in the elongation and differentiation zone of the root tip 3h after treatment. This elevated H2O2 production was accompanied by root growth inhibition and probably invoked root swelling in the elongation zone of the root tip. By contrast, a high 60μM Cd concentration induced robust H2O2 production in the elongation zone of the root tip already 1h after short-term treatment. This robust H2O2 generation caused extensive cell death 6h after short-term treatment. Similarly to low Cd concentration, exogenously applied H2O2 caused marked root growth inhibition, which at lower H2O2 concentration was accompanied by root swelling. The auxin signaling inhibitor p-chlorophenoxyisobutyric acid effectively inhibited 10μM Cd-induced root growth inhibition, H2O2 production and root swelling, but was ineffective in the alleviation of 60μM Cd-induced root growth inhibition and H2O2 production. Our results demonstrated that Cd-induced mild oxidative stress caused root growth inhibition, likely trough the rapid reorientation of cell growth in which a crucial role was played by IAA signaling in the root tip. Strong oxidative stress induced by high Cd concentration caused extensive cell death in the elongation zone of the root tip, resulting in the cessation of root growth or even in root death.

Relationship Between Polyamines Metabolism and Cell Death in Roots of Malus hupehensis Rehd. Under Cadmium Stress

The free putrescine (Put) content, the hydrogen peroxide (H2O2) content and the polyamine oxidase (PAO) activity in roots of Malus hupehensis Rehd. var. pinyiensis Jiang (PYTC) were significantly increased, and reached its peak at 1, 2 and 6 h, respectively, under cadmium treatment. The free spermine (Spm) and spermidine (Spd) contents were dramatically decreased, and reached the minimum value at 4–6 h, then remained relatively stable. The change in total free polyamines (PAs) content was consistent with that of free Put. The number of root dead cells was gradually increased after treatment for 24 h, and the typical characteristics of programmed cell death (PCD) were displayed at 48 h. Throughout the Cd treatment process, changes in PAs metabolism appeared to be prior to cell death increase, and the H2O2 content was always maintained at a high level. These results indicated that polyamines could initiate cell death by generating H2O2 in roots of Malus hupehensis Rehd. under CdSO4 stress.

Growth and physiological responses of five Malus species to the pH of hydroponic solutions

We characterized the growth and physiological responses of five wild Malus species indigenous to China to acid and alkaline abiotic stresses. At the six-leaf stage, seedlings of Malus sieversii, Malus prunifolia, Malus robusta, Malus mandshurica and Malus hupehensis were transferred to hydroponic systems in which the solution pH was 5.5, 7.0 or 8.5. Fresh and dry weights, plant heights and root lengths were measured over time. After 20 days of treatment, malondialdehyde (MDA), hydrogen peroxide (H2O2), ascorbic acid (AsA), andglutathione (GSH) contents were determined, as well as the rate of superoxide radical (O2.-) generation and antioxidant enzyme activities in the roots. When the pH was raised from 7.0 to 8.5, seedlings of M. sieversiidisplayed significant increases in their growth parameters whereas values for all other traits were significantly decreased. The opposite trend was observed with M. prunifolia and M. hupehensis. When the pH was decreased from 7.0 to 5.5, seedlings of M. sieversii showed a significant decline in their dry weights but a significant improvement in root MDA, H2O2, AsA, and GSH contents, O2.- generation and enzyme activities. In conclusions, these results suggest that growth is not severely inhibited for M. prunifolia, M. mandshurica, andM. hupehensis under acidic conditions (pH 5.5) or alkaline conditions for M. sieversii, M. robusta and M.mandshurica (pH 8.5). In fact, M. sieversii grew better at pH 8.5, whereas growth was significantly inhibited forM. sieversii and M. robusta at pH 5.5 and for M. prunifolia and M. hupehensis at pH 8.5. Key words: Antioxidant enzymes, antioxidants, growth traits, hydroponics pH, Malus species, oxidative stress.

Caspase-like enzymatic activity and the ascorbate-glutathione cycle participate in salt stress tolerance of maize conferred by exogenously applied nitric oxide

Salinity stress causes ionic stress (mainly from high Na+ and Cl- levels) and osmotic stress (as a result of inhibition of water uptake by roots and amplified water loss from plant tissue), resulting in cell death and inhibition of growth and ultimately adversely reducing crop productivity. In this report, changes in root nitric oxide content, shoot and root biomass, root H2O2 content, root lipid peroxidation, root cell death, root caspase-like enzymatic activity, root antioxidant enzymatic activity and root ascorbate and glutathione contents/redox states were investigated in maize (Zea mays L. cv Silverking) after long-term (21 d) salt stress (150 mM NaCl) with or without exogenously applied nitric oxide generated from the nitric oxide donor 2,2′-(Hydroxynitrosohydrazano)bis-ethane. In addition to reduced shoot and root biomass, salt stress increased the nitric oxide and H2O2 contents in the maize roots and resulted in elevated lipid peroxidation, caspase-like activity and cell death in the roots. Altered antioxidant enzymatic activities, along with changes in ascorbate and glutathione contents/redox status were observed in the roots in response to salt stress. The detrimental effects of salt stress in the roots were reversed by exogenously applied nitric oxide. These results demonstrate that exogenously applied nitric oxide confers salt stress tolerance in maize by reducing salt stress-induced oxidative stress and caspase-like activity through a process that limits accumulation of reactive oxygen species via enhanced antioxidant enzymatic activity.

Effects of scion and rootstock genotypes on the anti-oxidant defense systems of grafted cucumber seedlings under NaCl stress

To assess the effects of scion and rootstock genotypes on plant growth and the anti-oxidant defense systems of cucumber, two cucumber cultivars (“Jinchun No. 2”, a relatively salt-sensitive cultivar, and “Jinyu No. 1”, a relatively salt-tolerant cultivar) were grafted onto two rootstocks (“Figleaf Gourd”, a relatively salt-sensitive cultivar, and “Chaojiquanwang”, a relatively salt-tolerant cultivar), respectively. The grafted plants were grown hydroponically and were exposed to 0 and 100 mmol L−1 NaCl stress under greenhouse conditions. The results showed that the salt injury index, leaf Na+ content and root hydrogen peroxide (H2O2) content in the plants grafted onto Figleaf Gourd were higher, whereas the activities of superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) in the leaves and roots were lower than those of the plants grafted onto Chaojiquanwang under NaCl stress, independent of the scion genotypes. This resulted in smaller reductions in root dry weight, shoot dry weight and leaf area in the plants grafted onto Chaojiquanwang. The salt injury index was positively correlated with the leaf Na+ content and H2O2 content in the leaves and roots, but was negatively correlated with the activities of SOD and POD in the roots. The effects of scion on the salt injury index, Na+ content in the leaves, H2O2 content and activities of POD and CAT in the roots were significant, whereas there were no significant effects of scion on the root and shoot dry weights, leaf area and leaf relative water content. Overall, these results suggest the following: (1) the salt-tolerant rootstock was more important in determining the salt tolerance of the grafted cucumber seedlings under NaCl stress compared with the scion genotypes, (2) the higher salt tolerance of the grafted cucumber seedlings could be partially attributed to the lower leaf Na+ content and root H2O2 content and higher activities of SOD, POD and CAT in the roots.

INDUCTION OF RESISTANCE IN PHASEOLUS VULGARIS TO ROOT ROT DISEASE, CAUSED BY RHIZOCTONIA SOLANI, USING JASMONIC ACID

Application of 10 µM of Jasmonic acid (IA) as seed soaking significantly reduced % of root rot incidence in bean plants. Production of reactive oxygen species especially H2O2 was highly increased in both roots and shoots of R. solani infected-plants, as compared with non-infected control. JA significantly decreased the level of H2O2 in roots of 14 and 28 days old plants. Lipid peroxidation decreased in JA-treated plants, as compared with infected control. Activity of antioxidant enzymes (catalase, ascorbate peroxidase and superoxide dismutase) significantly increased in bean tissues in response to both pathogenic fungi (R.solani) and/or elicitor JA. JA-treated plants significantly increased total phenolic compounds and various phenolic acids except gallic acid in bean roots as compared with infected or noninfected control. Coumarin was induced only by JA. Additionally, activities of POX, PPO and PAL were greatly increased in both roots and shoots of bean plants infected by Rhizoctonia solani. JA treated plants showed the highest enzymes activity.

Involvement of root ethylene and oxidative stress-related activities in pre-conditioning of tomato transplants by increased salinity

SummaryIn order to understand the mechanism(s) of pre-conditioning (PC) by increased levels of salinity that enhance transplant establishment in the field, we studied the possible roles of ethylene and oxidative stress in transplants of tomato (Solanum lycopersicum L.) during PC by saline treatments. Prior to transplanting, transplants were fed hydroponically for 6 d with a normal nutrient solution, or solutions containing various concentrations of NaCl, ethephon, or silver thiosulphate (STS), or various combinations of these chemicals. The transplants were then transferred to normal nutrient solution for 10 d to recover. Addition of up to 75 mM NaCl to the nutrient solution decreased root elongation slightly and significantly increased lateral root (LR) formation during PC, but enhanced the subsequent recovery of roots and shoots. All the salinity treatments inhibited root ethylene production. However, the application of 20 mg l–1 ethephon to the roots during PC increased ethylene concentrations, inhibited root elongation, increased LR formation, and stimulated subsequent root growth.An enhancement of root growth during recovery was also observed when 75 mM NaCl plus 20 mg l–1 ethephon was applied to the root system during PC. The ethylene blocker, STS, enhanced root elongation during PC, and acted synergistically with NaCl in enhancing subsequent root growth. As PC treatments, NaCl or ethephon both increased root malonyldialdehyde (MDA) and H2O2 concentrations, and increased root peroxidase and superoxide dismutase (SOD) activities; but, if NaCl or ethephon were combined with STS, the extent of these increases was diminished. Application of ≤ 1 mM H2O2 at PC increased subsequent root and shoot growth in seedlings, but higher H2O2 concentrations were inhibitory. These results suggest that subsequent root growth upon recovery from transplantation benefits from PC applications of NaCl or ethephon and this might be due the enhancement of LR initiation during PC by these treatments, possibly through the signalling role of H2O2. STS alleviates oxidative stress in the root by inhibiting the action of ethylene and decreasing ethylene sensitivity, thereby promoting root recovery. These morphological and biochemical changes may explain the beneficial effect of PC on transplant development under field conditions.

Methylene blue increases the tolerance of tomato plants to abiotic stresses

SummaryEnvironmental stresses such as high temperature and high salinity are known to cause oxidative stress in plants by enhancing the generation of reactive oxygen species (ROS), which have a strong impact on plant development. Mitochondria constitute one of the major sources of ROS in roots. Recently, it has been shown in mammalian systems that methylene blue (MB) at very low concentrations can attenuate mitochondrial ageing by scavenging ROS. In the present study, we tested whether MB could be used to protect tomato (Lycopersicon esculentum L.) seedlings against the harmful effects of high salinity and high root temperature. Tomato seedlings were grown hydroponically and were subjected to two abiotic stresses: high salinity (150 mM NaCl) in the nutrient medium, or high root temperature (35ºC) for 14 d. These stress treatments were applied with or without the addition of 10–8 M MB. The results showed that, under normal conditions, MB enhanced root and shoot growth, increased root respiration, decreased root H2O2 and malonyldialdehyde (MDA) concentrations, and enhanced root superoxide dismutase (SOD) activity. Both high salinity and high root temperature stress impaired root and shoot growth, and caused an enhancement of root oxidative stress. MB had a significant protective effect against both abiotic stresses and restored the levels of oxidative stress-related components (i.e., H2O2 and MDA) in root and leaf tissues almost to normal levels. It is hypothesised that MB could confer its protective effect not only through interactions with root mitochondria, but also via additional cellular targets in the root.

Nitric oxide alleviates Fe deficiency-induced stress in Solanum nigrum

The possible involvement of nitric oxide in response of Solanum nigrum seedlings to Fe deficiency was investigated. Iron deficiency resulted in decreased shoot height and chlorophyll content and increased proliferation of root hairs and H2O2, K+ and Ca2+ content. NO donor S-nitrosoglutathione (GSNO) was effective in preventing Fe deficiency-induced increase in content of H2O2 and the ion uptake. The protective effects of GSNO were reversed by cPTIO, an NO scavenger, and tungstate, a nitrate reductase (NR) inhibitor.

Changes in fatty acids composition, hydrogen peroxide generation and lipid peroxidation of salt-stressed corn (Zea mays L.) roots

Comparative study about the salt-induced oxidative stress and lipid composition has been realised in primary root tissues for two varieties of maize (Zea mays L.) in order to evaluate their responses to salt stress. The root growth, root water content (WC), hydrogen peroxide (H2O2) generation, lipid peroxidation, membrane stability index and the changes in the profile of fatty acids composition were investigated. Salinity impacts in term of root growth, water content, H2O2 generation, lipid peroxidation and membrane destabilisation were more pronounced in primary roots of Aristo than in those of Arper indicating more sensitivity of the first variety. It was confirmed by gas chromatography that the composition of fatty acids in roots of both varieties was constituted mainly by 16:0 and 18:0 as major saturated fatty acids and 18:1ω9, 18:2ω6 and 18:3ω3 as major unsaturated fatty acids. Total lipid extracts from the roots of both varieties showed that the lipid saturation level increased under salt stress, notwithstanding the increased proportion of polyunsaturated fatty acids. The changes in lipid saturation being predominantly due to decreases in oleic acid (18:1ω9) and increases in palmitic acid (16:0). However, Arper root extracts contained a lower proportion of saturated lipids than Aristo. The enhanced proportion of highly polyunsaturated fatty acids especially linolenic and eicosapentaenoic acids was considered to be the characteristic of the relatively salt tolerance in Arper roots.

Influences of phosphorus starvation on OsACR2.1 expression and arsenic metabolism in rice seedlings

The effects of phosphorus (P) status on arsenate reductase gene (OsACR2.1) expression, arsenate reductase activity, hydrogen peroxide (H2O2) content, and arsenic (As) species in rice seedlings which were exposed to arsenate after −P or +P pretreatments were investigated in a series of hydroponic experiments. OsACR2.1 expression increased significantly with decreasing internal P concentrations; more than 2-fold and 10-fold increases were found after P starvation for 30 h and 14 days, respectively. OsACR2.1 expression exhibited a significant positive correlation with internal root H2O2 accumulation, which increased upon P starvation or exposure to H2O2 without P starvation. Characterization of internal and effluxed As species showed the predominant form of As was arsenate in P-starved rice root, which contrasted with the +P pretreated plants. Additionally, more As was effluxed from P-starved rice roots than from non-starved roots. In summary, an interesting relationship was observed between P-starvation induced H2O2 and OsACR2.1 gene expression. However, the up-regulation of OsACR2.1 did not increase arsenate reduction in P-starved rice seedlings when exposed to arsenate.

Physiological and oxidative stress responses of four potato clones to aluminum in nutrient solution

Aluminum toxicity is a serious problem in Brazilian soils and selecting potato clones is an important strategy to produce this crop on these kinds of soils. Potato clones, Macaca, SMIC148-A, Dakota Rose, and Solanum microdontum, were grown in a nutrient solution (pH 4.0) with 0, 50, 100, 150 and 200 mg Al L-1. After 7 d, Al concentration in both root system and shoot of all clones increased linearly with increasing Al levels. Based on relative root growth, S. microdontum and SMIC148-A were considered Al-tolerant clones, whereas Macaca and Dakota Rose were considered Al-sensitive. Shoot growth in Macaca linearly decreased with increasing Al levels. Root H2O2 concentration in both Al-sensitive clones increased with increasing Al supply, whereas in Al-tolerant clones it either decreased (SMIC148-A) or demonstrated no alteration (S. microdontum). Shoot H2O2 concentration increased linearly in Macaca, whereas for Dakota Rose it showed a quadratic relationship with Al levels. On the other hand, shoot H2O2 concentration in the Al-tolerant clones either demonstrated no alteration (S. microdontum) or presented lower levels (SMIC148-A). Root catalase (CAT) activity in both Al-sensitive clones increased with increasing Al levels, whereas in Al-tolerant clones it either demonstrated no alteration (SMIC148-A) or presented lower levels (S. microdontum). Shoot CAT activity in the S. microdontum increased curvilinearly with increasing Al levels. In all potato clones, chlorophyll concentration showed a curvilinear response to Al supply, where in Al-sensitive clones it decreased upon addition of Al exceeding 100 mg L-1, but in SMIC148-A it increased at levels between approximately 100 and 150 mg L-1, and decreased in S. microdontum regardless of the Al level. Carotenoid concentrations in the Al-sensitive clones were linearly decreased with increasing Al levels. Aluminum supply caused root lipid peroxidation only in the Al-sensitive clones, whereas in the shoot it increased linearly in the Al-sensitive clones and in S. microdontum it only increased at around 50 mg L-1. Most of root protein oxidation was only observed in the Al-sensitive clones. However, shoot protein oxidation was increased with increasing Al levels for all potato clones. These results indicate that oxidative stress caused by Al in potato may harm several components of the cell, mainly in Al-sensitive clones.

Expression of ASCORBATE PEROXIDASE 8 in roots of rice (Oryza sativa L.) seedlings in response to NaCl

Reactive oxygen species are thought to play an important role in NaCl stress. Therefore, the expression patterns of the gene family encoding the H(2)O(2)-scavenging enzyme ascorbate peroxidase (APx; EC1.11.1.11) were analysed in roots of etiolated rice (Oryza sativa L.) seedlings in response to NaCl stress. Applying semi-quantitative RT-PCR, the mRNA levels were quantified for two cytosolic (OsAPx1 and OsAPx2), two peroxisomal (OsAPx3 and OsAPx4), and four chloroplastic (OsAPx5, OsAPx6, OsAPx7, and OsAPx8) isoforms identified in the rice genome. NaCl at 150 mM and 200 mM increased the expression of OsAPx8 and the activities of APx, but had no effect on the expression of OsAPx1, OsAPx2, OsAPx3, OsAPx4, OsAPx5, OsAPx6, and OsAPx7 in rice roots. However, NaCl at 300 mM up-regulated OsAPx8 expression, increased APx activity, and down-regulated OsAPx7 expression, but had no effect on the expression of OsAPx1, OsAPx2, OsAPx3, OsAPx4, OsAPx5, and OsAPx6. The accumulation of abscisic acid (ABA) in response to NaCl was observed in rice roots. Exogenously applied ABA also specifically enhanced the expression of OsAPx8 in rice roots. The accumulation of ABA in rice roots in response to NaCl was inhibited by fluridone (Flu), an inhibitor of carotenoid biosynthesis. Flu treatment also suppressed NaCl-enhanced OsAPx8 expression and APx activity. The effect of Flu on the expression of OsAPx8 and increase in APx activity was reversed by the application of ABA. It appears that NaCl-enhanced expression of OsAPx8 in rice roots is mediated through an accumulation of ABA. Evidence is provided to show that Na(+) but not Cl(-) is required for enhancing OsAPx8 expression, APx activity, and ABA accumulation in rice roots treated with NaCl. H(2)O(2) treatment resulted in an enhancement of OsAPx8 induction but no accumulation of ABA. Diphenylene iodonium treatment, which is known to inhibit NaCl-induced accumulation of H(2)O(2) in rice roots, did not suppress OsAPx8 induction and ABA accumulation by NaCl. It appears that H(2)O(2) is not involved in the regulation of NaCl-induced OsAPx8 expression in rice roots.

Expression of ascorbate peroxidase and glutathione reductase in roots of rice seedlings in response to NaCl and H 2O 2

The accumulation of H2O2 by NaCl was observed in the roots of rice seedlings. Treatment with NaCl caused an increase in the activities of ascorbate peroxidase (APX) and glutathione reductase (GR) and the expression of OsAPX and OsGR in rice roots. Exogenously applied H2O2 also enhanced the activities of APX and GR and the expression of OsAPX and OsGR in rice roots. The accumulation of H2O2 in rice roots in response to NaCl was inhibited by the NADPH oxidase inhibitors, diphenyleneiodonium chloride (DPI) and imidazole (IMD). However, DPI, IMD, and dimethylthiourea, a H2O2 trap, did not reduce NaCl-enhanced activities of APX and GR and expression of OsAPX and OsGR. It appears that H2O2 is not involved in the regulation of NaCl-induced APX and GR activities and OsAPX and OsGR expression in rice roots.

Luminol-dependent chemiluminescence analysis of chitooligosaccharide-induced rapid production of hydrogen peroxide by intact wheat seedlings.

The feasibility of a luminol-dependent chemiluminescence analysis of hydrogen peroxide production by intact wheat seedlings using a KhL-003 chemiluminometer was determined. It was shown that the minimal H2O2 concentration that can be detected in a 0.5-ml sample with this instrument is 0.125 microM. Analysis of biological activity of a mixture of chitooligosaccharides with molecular masses from 5 to 10 kD and acetylation degree of 65% demonstrated that, at a concentration of 1 microg/ml, they induce rapid overproduction of H2O2 in roots of 3-day-old wheat seedlings.

Hydrogen peroxide production by roots and its stimulation by exogenous NADH

H2O2 production by roots of young seedlings was monitored using a non‐destructive in vivo assay at pH 5.0. A particularly high rate of H2O2 production was measured in the roots of soybean (Glycine max L. cv. Labrador) seedlings which were used for further investigation of the physiological and enzymological properties of apoplastic H2O2 production. In the soybean root H2O2 production can be stimulated 10‐fold by exogenous NADH or NADPH. This response displays typical features of a peroxidase‐catalyzed oxidase reaction using NAD(P)H as electron donor for the reduction of O2 to H2O2. Comparative measurements showed that the NADH‐induced H2O2 production of the roots resembles the H2O2‐forming activity of horseradish peroxidase with respect to NADH and O2 concentration requirements and sensitivity to inhibition by KCN, NaN3, superoxide dismutase and catalase. NADH‐induced H2O2 production can be observed with similar intensity in all regions of the root, in agreement with the distribution of apoplastic peroxidase activity. In contrast, the activity responsible for the basal H2O2 production in the absence of exogenous NADH was mainly confined to a short subapical zone of the root and differs from the NADH‐induced reaction by insensitivity to inhibition by superoxide dismutase and a strikingly lower requirement for O2. It is concluded that the basal H2O2 production of the root is mediated by an enzyme different from peroxidase, possibly a plasma membrane O2−‐producing oxidase.