Dimethylthiourea Research Articles

A multilevel investigation to reveal the regulatory mechanism of lignin accumulation in juice sac granulation of pomelo

Granulation of juice sacs is a physiological disorder, which affects pomelo fruit quality. Here, the transcriptome and ubiquitinome of the granulated juice sacs were analyzed in Guanxi pomelo. We found that lignin accumulation in the granulated juice sacs was regulated at transcription and protein modification levels. In transcriptome data, we found that the genes in lignin biosynthesis pathway and antioxidant enzyme system of the granulated juice sacs were significantly upregulated. However, in ubiquitinome data, we found that ubiquitinated antioxidant enzymes increased in abundance but the enzyme activities decreased after the modification, which gave rise to reactive oxygen species (ROS) contents in granulated juice sacs. This finding suggests that ubiquitination level of the antioxidant enzymes is negatively correlated with the enzyme activities. Increased H2O2 is considered to be a signaling molecule to activate the key gene expressions in lignin biosynthesis pathway, which leads to the lignification in granulated juice sacs of pomelo. This regulatory mechanism in juice sac granulation of pomelo was further confirmed through the verification experiment using tissue culture by adding H2O2 or dimethylthiourea (DMTU). Our findings suggest that scavenging H2O2 and other ROS are important for reducing lignin accumulation, alleviating juice sac granulation and improving pomelo fruit quality.

Involvement of ROS and calcium ions in developing heat resistance and inducing antioxidant system of wheat seedlings under melatonin's effects.

The stress-protective effect of melatonin (N-acetyl-5-methoxytryptamine) on plant cells is mediated by key signaling mediators, in particular calcium ions and reactive oxygen species (ROS). However, the links between changes in calcium and redox homeostasis and the formation of adaptive responses of cultivated cereals (including wheat) to the action of high temperatures have not yet been studied. In the present study, we investigated the possible involvement of ROS and calcium ions as signaling mediators in developing heat resistance in wheat (Triticum aestivum L.) seedlings and activating their antioxidant system. Treatment of 3-day-old etiolated seedlings with melatonin solutions at concentrations 0.01-10µM increased their survival after exposure to 45°C for 10min. The most significant stress-protective effect was exerted by melatonin treatment at 1µM concentration. Under the influence of melatonin, a transient enhancement of superoxide anion radical (O2•-) generation and an increase in hydrogen peroxide content were observed in roots, with a maximum at 1h. Four hours after treatment with melatonin, the activity of catalase and guaiacol peroxidase increased in roots, while the activity of superoxide dismutase did not change significantly. After exposure to 45°C, the activity of catalase and guaiacol peroxidase was higher in the roots of melatonin-treated wheat seedlings, and the indices of ROS generation, content of the lipid peroxidation product malonic dialdehyde, and cell membrane damage were lower than in control seedlings. Melatonin-induced changes in root ROS generation and antioxidant enzyme activities were eliminated by pretreatment with the hydrogen peroxide scavenger dimethylthiourea (DMTU), NADPH oxidase inhibitor imidazole, and calcium antagonists (the extracellular calcium chelator EGTA and phospholipase C inhibitor neomycin). Treatment with DMTU, imidazole, EGTA, and neomycin also abolished the melatonin-induced increase in survival of wheat seedlings after heat stress. The role of calcium ions and ROS, generated with the participation of NADPH oxidase, as signaling mediators in the melatonin-induced antioxidant system and heat stress resistance of wheat seedlings have been demonstrated.

MiR408 balances plant growth and heat response in rice

MicroRNAs are small noncoding RNAs that regulate gene expression by targeting complementary mRNA molecules. Among them, microRNA408 (miR408) is an ancient microRNA influencing plant growth and aiding in abiotic stress adaptation. We proved that miR408 sliced uclacyanin7 (UCL7) directly by degradome sequencing. However, the mechanisms underlying the balance between growth and stress response of rice (Oryza sativa L.) controlled by miR408 remain unclear. We conducted a study comparing Nipponbare (wild-type, WT) plants with miR408–3p-overexpression (miR408–3p-OX) and UCL7-overexpression (UCL7-OX) plants under normal and heat stress conditions. Through RNA-sequencing, physiological measurements, and phenotype examination, we observed that miR408 overexpression resulted in increased dry weight and grain yield, higher levels of ATP and non-structural carbohydrates (NSC), improved photosynthesis, and reduced H2O2 content, antioxidant enzyme activity, and ATPase activity in seedling plants compared to UCL7-OX plants. Additionally, the application of H2O2 scavengers, such as ascorbic acid (ASA) and dimethylthiourea (DMTU), further enhanced dry weight and photosynthesis by reducing H2O2 levels. Under heat stress conditions, miR408 overexpression sharply increased H2O2 content, antioxidant enzyme activity, ATP levels, and ATPase activity compared to UCL7-OX plants, thereby enhancing heat tolerance in rice seedlings. These findings suggest that miR408 and its target gene UCL7 participate in regulating energy homeostasis to achieve a balance between growth and heat response. This regulation involves modulating H2O2 levels and optimizing the relationship between the target of rapamycin (TOR) and SNF1-related protein kinase 1 (SnRK1) in rice.

H2O2 Significantly Affects Larix kaempferi × Larix olgensis Somatic Embryogenesis.

Larch is widely distributed throughout the world and is an important species for timber supply and the extraction of industrial raw materials. In recent years, the hybrid breeding of Larix kaempferi and Larix olgensis has shown obvious heterosis in quick-growth, stress resistance and wood properties. However, its growth and development cycle is too long to meet general production needs. In order to shorten the breeding cycle, we have for the first time successfully established and optimized a somatic embryogenesis system for Larix kaempferi × Larix olgensis. We found that the highest rate of embryonal-suspensor mass (ESM) induction was observed when late cotyledonary embryos were used as explants. The induced ESMs were subjected to stable proliferation, after which abscisic acid (ABA) and polyethylene glycol (PEG) were added to successfully induce somatic embryos. Treatment with PEG and ABA was of great importance to somatic embryo formation and complemented each other's effect. ABA assisted embryo growth, whereas PEG facilitated the formation of proembryo-like structures. On top of this, we studied in more detail the relationship between redox homeostasis and the efficiency of somatic embryogenesis (frequency of ESM induction). During subculture, we observed the gradual formation of three distinct types of ESM. The Type I ESM is readily able to form somatic embryos. In contrast to type I, the type III ESM suffers from severe browning, contains a higher level of hydrogen peroxide (H2O2) and demonstrates a decreased ability to form somatic embryos. External treatment with H2O2 decreased the somatic embryogenesis efficiency of Type I and type III ESMs, or the higher the exogenous H2O2 content, the lower the resulting somatic embryogenesis efficiency. We found that treatment with the H2O2 scavenger DMTU (dimethylthiourea) could significantly increase the somatic embryogenesis efficiency of the type III ESM, as a result of a decline in endogenous H2O2 content. Overall, these findings have contributed to setting up a successful somatic embryogenesis system for larch production.

Dimethylthiourea as a Quencher in Hydroxyl Radical Protein Footprinting Experiments.

Hydroxyl radical protein footprinting (HRPF) is a mass-spectrometry-based method for studying protein structures, interactions, conformations, and folding. This method is based on the irreversible labeling of solvent-exposed amino acid side chains by hydroxyl radicals. While catalase is commonly used as a quencher after the labeling of a protein by the hydroxyl radicals to efficiently remove the remaining hydrogen peroxide, it has some disadvantages. Catalase quenching adds a relatively high amount of protein to the sample, limiting the sensitivity of the method due to dynamic range issues and causing significant issues when dealing with more complex samples. We evaluated dimethylthiourea (DMTU) as a replacement for catalase in the quenching HRPF reactions. We observed that DMTU is highly effective at quenching HRPF oxidation. DMTU does not cause the background protein issues that catalase does, resulting in an increased number of protein identifications from complex mixtures. We recommend the replacement of catalase quenching with DMTU for all HRPF experiments.

Bordeaux mixture accelerates ripening, delays senescence, and promotes metabolite accumulation in jujube fruit

Abstract The old, but eco-friendly pesticide, Bordeaux mixture (Bm), is widely used in agriculture. Here, the effects of Bm on preharvest fruit ripening, and postharvest senescence of fruit and the accumulation of antioxidants were studied in Ziziphus jujuba. A 200× dilution of Bm enhanced preharvest ripening and retarded jujube fruit postharvest senescence. Treatment with Bm increased the reddening index and promoted the accumulation of total phenolics (TP) and the total antioxidant capacity (TAC) in preharvest fruit. However, this Bm-accelerated ripening could be partly reversed using dimethylthiourea (DMTU), a specific scavenger of reactive oxygen species. Compared with preharvest fruit, Bm treatment decreased weight loss and decay and increased firmness in postharvest fruit. Similarly, Bm-delayed senescence was partly reversed using dimethylthiourea. Moreover, the decrease in TP and TAC during storage was partly impeded by Bm. A higher H2O2 content was detected in preharvest fruit than in postharvest fruit. Moreover, this Bm-induced H2O2 accumulation was significantly mitigated using dimethylthiourea. Interestingly, both preharvest and postharvest spraying with Bm significantly enhance copper and calcium accumulation in fruit. Thus, Bm can be widely used before or after harvest to improve fruit quality.

Effect of Reactive Oxygen Scavenger N,N'-Dimethylthiourea (DMTU) on Seed Germination and Radicle Elongation of Maize.

Reactive oxygen species (ROS) are an important part of adaptation to biotic and abiotic stresses and regulate seed germination through positive or negative signaling. Seed adaptation to abiotic stress may be mediated by hydrogen peroxide (H2O2). The effects of the ROS scavenger N,N'-dimethylthiourea (DMTU) on maize seed germination through endogenous H2O2 regulation is unclear. In this study, we investigated the effects of different doses of DMTU on seed endogenous H2O2 and radicle development parameters using two maize varieties (ZD958 and DMY1). The inhibitory effect of DMTU on the germination rate and radicle growth was dose-dependent. The inhibitory effect of DMTU on radicle growth ceased after transferring maize seeds from DMTU to a water medium. Histochemical analyses showed that DMTU eliminated stable H2O2 accumulation in the radicle sheaths and radicles. The activity of antioxidant enzyme and the expression of antioxidant enzyme-related genes (ZmAPX2 and ZmCAT2) were reduced in maize seeds cultured with DMTU compared with normal culture conditions (0 mmol·dm-3 DMTU). We suggest the use of 200 mmol·dm-3 DMTU as an H2O2 scavenger to study the ROS equilibrium mechanisms during the germination of maize seeds, assisting in the future with the efficient development of plant growth regulators to enhance the seed germination performance of test maize varieties under abiotic stress.

The influence of electronic effects on the substitution reactions of N,N′-donor cis-palladium(II) complexes with bidentate (pyridin-2-yl)methyl-aniline ligands

The substitution behaviour of bidentate N,N′-donor (pyridin-2-yl)methyl-aniline chelates with different substituents on the mononuclear Pd(II) complexes was investigated. The complexes were synthesized and characterized using 1H NMR, 13C NMR, FTIR, LC–MS, and elemental analysis. The study explored the kinetics and mechanistic behaviour of N,N′-pyridyl Pd(II) complexes, viz. dichloro-(N-((pyridin-2-yl)methyl)aniline)palladium(II) (PdL1), dichloro-(4-fluoro-N-((pyridin-2-yl)methyl)aniline)-palladium(II) (PdL2), dichloro-(4-methoxy-N-((pyridin-2-yl)methyl)aniline)-palladium(II) (PdL3) and dichloro-(4-ethyl-N-((pyridin-2-yl)methyl)aniline)-palladium(II) (PdL4). The effect of concentration and temperature on substitution behaviour of coordinated chloride ligands by three bio-relevant thiourea nucleophiles, viz. thiourea (TU), N,N′-dimethylthiourea (DMTU) and N,N,N′,N′-tetramethylthiourea (TMTU), of different steric demands was studied in a 0.1 M (LiCl) solution of ultra-pure water under pseudo-first order conditions using standard Stopped-Flow and UV–Visible spectrophotometric techniques. The substitution of the chloride ligands from the Pd(II) metal by thiourea nucleophiles was a two-step reaction, where the chloride trans to the pyridine ligand was substituted first due to the strong trans effect on the pyridine ring compared to the amine group. The rate of substitution of the chloride by thiourea nucleophiles increased with the presence of an electron-withdrawing substituent and decreased when an electron-donating group was attached to the para position of the phenyl moiety. The negative activation entropies and positive activation enthalpy for all the substitution reactions support an associative mode of substitution mechanism.

The OsTIL1 lipocalin protects cell membranes from reactive oxygen species damage and maintains the 18:3-containing glycerolipid biosynthesis under cold stress in rice.

Lipocalins constitute a conserved protein family that binds to and transports a variety of lipids while fatty acid desaturases (FADs) are required for maintaining the cell membrane fluidity under cold stress. Nevertheless, it remains unclear whether plant lipocalins promote FADs for the cell membrane integrity under cold stress. Here, we identified the role of OsTIL1 lipocalin in FADs-mediated glycerolipid remodeling under cold stress. Overexpression and CRISPR/Cas9 mediated gene edition experiments demonstrated that OsTIL1 positively regulated cold stress tolerance by protecting the cell membrane integrity from reactive oxygen species damage and enhancing the activities of peroxidase and ascorbate peroxidase, which was confirmed by combined cold stress with a membrane rigidifier dimethyl sulfoxide or a H2 O2 scavenger dimethyl thiourea. OsTIL1 overexpression induced higher 18:3 content, and higher 18:3/18:2 and (18:2 + 18:3)/18:1 ratios than the wild type under cold stress whereas the gene edition mutant showed the opposite. Furthermore, the lipidomic analysis showed that OsTIL1 overexpression led to higher contents of 18:3-mediated glycerolipids, including galactolipids (monoglactosyldiacylglycerol and digalactosyldiacylglycerol) and phospholipids (phosphatidyl glycerol, phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine and phosphatidyl inositol) under cold stress. RNA-seq and enzyme linked immunosorbent assay analyses indicated that OsTIL1 overexpression enhanced the transcription and enzyme abundance of four ω-3 FADs (OsFAD3-1/3-2, 7, and 8) under cold stress. These results reveal an important role of OsTIL1 in maintaining the cell membrane integrity from oxidative damage under cold stress, providing a good candidate gene for improving cold tolerance in rice.

Stimulating Role of Calcium and Cyclic GMP in Mediating the Effect of Magnetopriming for Alleviation of Salt Stress in Soybean Seedlings

This current study examined the role of calcium (Ca) and Cyclic GMP (cGMP) in mitigating the adverse effect of salt stress through magnetopriming of soybean cultivar JS-335 seeds with a static magnetic field (SMF, 200 mT for 1 h). The salt stress (50 mMNaCl) extensively reduced the early seedling growth (64%), vigour Index-I (71%), vigour Index-II (39%), total amylase (59%), protease (63%), and nitrate reductase (NR, 19%) activities in un-primed soybean seedlings. However, magnetopriming and Ca treatment enhanced all of these measured parameters along with remarkable increase in reactive oxygen species (ROS) and nitric oxide (NO) content. The exogenous application of Ca2+, cGMP and ROS regulators such as nifedipine (Ca2+ channel blocker), EGTA, ethylene glycol-β-amino ethyl ether tetra acetic acid (Ca2+chelators), genistein (cGMP blocker), and dimethyl thiourea (DMTU, H2O2 inhibitor) negatively affects the SMF-induced seedling length, seedling vigour, ROS, NO, and enzyme activities such as protease, total amylase, and NR in soybean seedlings. Results presented by using specific various biochemical inhibitors of Ca, cGMP, or ROS signalling in vivo indicated that Ca and cGMP are also involved with ROS and NO in the signal transduction of magnetic field enthused soybean seed germination and seedling growth under salt stress.

Hydrogen Peroxide and GA3 Levels Regulate the High Night Temperature Response in Pistils of Wheat (Triticum aestivum L.).

High night temperature (HNT) impairs crop productivity through the reproductive failure of gametes (pollen and pistil). Though female gametophyte (pistil) is an equal partner in the seed-set, the knowledge of the antioxidant system(s) and hormonal control of HNT tolerance or susceptibility of pistils is limited and lacking. The objectives of this study were to determine the antioxidant mechanism for homeostatic control of free radicals, and the involvement of abscisic acid (ABA) and gibberellic acid (GA3) in HNT stress protection in the wheat pistils of contrasting wheat genotypes. We hypothesized that HNT tolerance is attributed to the homeostatic control of reactive oxygen species (ROS) and hormonal readjustment in pistils of the tolerant genotype. The ears of two contrasting wheat genotypes-HD 2329 (susceptible) and Raj 3765 (tolerant) were subjected to two HNTs (+5 °C and +8 °C) over ambient, in the absence and presence of dimethylthiourea (DMTU), a chemical trap of hydrogen peroxide (H2O2). Results showed that HNTs significantly increased ROS in pistils of susceptible genotype HD 2329 to a relatively greater extent compared to tolerant genotype Raj 3765. The response was similar in the presence or absence of DMTU, but the H2O2 values were lower in the presence of DMTU. The ROS levels were balanced by increased activity of peroxidase under HNT to a greater extent in the tolerant genotype. Cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) activity was inversely related to H2O2 production within a critical range in Raj 3765, indicating its modulation by H2O2 levels as no change was observed at the transcriptional level. The hormonal status showed increased ABA and decreased GA3 contents with increasing temperature. Our study elucidates the role of H2O2 and GA3 in stress tolerance of pistils of tolerant genotype where GAPC acts as a ROS sensor due to H2O2-mediated decrease in its activity.

Selective Enrichment of Nitrososphaera viennensis-Like Ammonia-Oxidizing Archaea over Ammonia-Oxidizing Bacteria from Drinking Water Biofilms.

Ammonia-oxidizing archaea (AOA) can oxidize ammonia to nitrite for energy gain. They have been detected in chloraminated drinking water distribution systems (DWDS) along with the more common ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB). To date, no members of the AOA have been isolated or enriched from drinking water environments. To begin the investigation of the role of AOA in chloraminated DWDS, we developed a selective approach using biofilm samples from a full-scale operational network as inoculum. A Nitrososphaera viennensis-like AOA taxon was enriched from a mixed community that also included Nitrosomonas-like AOB while gradually scaling up the culture volume. Dimethylthiourea (DMTU) and pyruvate at 100 μM were added to promote the growth of AOA while inhibiting AOB. This resulted in the eventual washout of AOB, while NOB were absent after 2 or 3 rounds of amendment with 24 μM sodium azide. The relative abundance of AOA in the enrichment increased from 0.2% to 39.5% after adding DMTU and pyruvate, and further to 51.6% after filtration through a 0.45-μm pore size membrane, within a period of approximately 6 months. IMPORTANCE Chloramination has been known to increase the risk of nitrification episodes in DWDS due to the presence of ammonia-oxidizing microorganisms. Among them, AOB are more frequently detected than AOA. All publicly available cultures of AOA have been isolated from soil, marine or surface water environments, meaning they are allochthonous to DWDS. Hence, monochloramine exposure studies involving these strains may not accurately reflect their role in DWDS. The described method allows for the rapid enrichment of autochthonous AOA from drinking water nitrifying communities. The high relative abundance of AOA in the resulting enrichment culture reduces any confounding effects of co-existing heterotrophic bacteria when investigating the response of AOA to varied levels of monochloramine in drinking water.

Substitution reactions of cis-platinum(II) complexes containing bidentate N,N-donor pyridinecarboxamide ligands with different substituents

Substitution reactions of [2-(pyridinecarboxamide)dichloride Pt(II)] [PtCl2 ], [N-phenyl-(2-pyridinecarboxamide)dichloride Pt(II)] [PhPtCl2 ], [N-(4-methylphenyl)-2-pyridinecarboxamide)dichloride Pt(II)] [CH3PhPtCl2 ], [N-(4-methoxyphenyl)-(2-pyridine-carboxamide)dichloride Pt(II)] [CH3OPhPtCl2 ] and [N-(4-fluorophenyl)-(2-pyridinecarboxamide)dichloride Pt(II)] [FPhPtCl2 ], with nucleophiles; thiourea (TU), N,N′-dimethylthiourea (DMTU) and N,N,N′,N′-tetramethylthiourea (TMTU) were studied under pseudo first-order conditions. The rates of substitution were investigated as a function of nucleophile concentration and temperature using stopped-flow and UV-visible absorption spectrophotometers. Substitutions of the two coordinated chloride ligands of the Pt(II) complexes occur consecutively, with the first substitution occurring opposite the coordinated pyridyl. The observed pseudo first-order rate constants regressed linearly with concentration of the incoming nucleophiles according to the equation k obs = k 2[Nu]. The highest substitution rates were measured for PtCl2, which is attributed to the unsubstituted non-leaving carboxamide ligand. The Pt(II) center of this complex is the most electrophilic as a result of the strong withdrawal of electron density through π-resonance by the carboxamide group. The introduction of a 4′-substituted phenyl group on the amido N of the carboxamide reduces the reactivity of the complexes due to proportional electronic effects from the ancillary substituents on the phenyl ring as well as its conformational disposition with respect to the plane of the complex. The order of reactivity of studied nucleophiles is TU > DMTU > TMTU. The substitution is associatively activated as supported by the negative entropy of activation values for the reactions.

Hydrogen peroxide regulates the biosynthesis of phenolic compounds and antioxidant quality enhancement in lettuce under low nitrogen condition.

Reduced nitrogen availability is an efficient strategy for increasing the accumulation of phenolic compounds in vegetables, but related mechanisms remain unknown. Here, the production of hydrogen peroxide (H2O2) and its potential roles in regulating phenolic biosynthesis and enhancing the antioxidant quality of lettuce under low nitrogen (LN) conditions were investigated. The LN treatment caused a rapid production of H2O2, which effectively increased lettuce quality by enhancing the levels of phenolic compounds and other nutrients such as ascorbic acid, glutathione, soluble sugar, and soluble protein. The increased phenolic content was related to the higher expression levels of phenolic biosynthesis genes, including PAL, CHS, DFR, F35H, and UFGT, and higher photosynthetic capacity after H2O2 addition under LN conditions. However, these positive effects were suppressed by dimethylthiourea (DMTU), a scavenger of H2O2. These results suggest that H2O2 as an important signal molecular mediates the LN-caused phenolic accumulation and antioxidant quality enhancement in lettuce.

Hydrogen peroxide regulated salicylic acid– and jasmonic acid–dependent systemic defenses in tomato seedlings

To investigate the regulation of hydrogen peroxide (H2O2) in SA- and JA-dependent systemic defense pathways in tomato seedlings damaged by cotton bollworm. The lower two leaves of four-leaf tomato plant seedlings were treated with cotton bollworms. The upper leaves were then removed at various time points and assessed for systemic changes in H2O2 levels, enzyme activity and defense-related genes expression. Wild-type seedlings, def-5 mutant seedlings (deficient in JA accumulation) and transgenic nahG seedlings(deficient in SA content) were constructed at four-leaf stage. Wild-type plants were further treated with dimethylthiourea (DMTU) and 2,5-dihydroxycinnamic acid methyl ester (DHC). Bollworm feeding in the lower leaves induced rapid systemic accumulation of H2O2 in the upper leaves. H2O2 accumulation further increased activities for SA- and JA-dependent enzymes including proteinase inhibitors (PIs), pathogenesis-related gene 1 (PR-1), cathepsin D inhibitor (CDI), β-1,3-glucanse (BGL) and polyphenol oxidase (PPO). Furthermore, H2O2 accumulation enhanced mRNA expression of BGL-2, PR-1, PI, CDI and PPO genes. Suppression of H2O2 accumulation using H2O2 scavengers substantially diminished these effects. mRNA expression of PR-1 was not induced in transgenic nahG plants. SA- and JA-dependent signaling pathways are involved in the tomato systemic defense responses to herbivores, and that H2O2 generation is required for both systemic pathways.

Ex vivo imaging and analysis of ROS generation correlated with microglial activation in rat model with acute neuroinflammation induced by intrastriatal injection of LPS

Neuroinflammation and oxidative stress are hallmarks of neurodegenerative diseases. Microglia, the major important regulators of neuroinflammation, are activated in response to excessive generation of reactive oxygen species (ROS) from damaged cells and resulting in elevated and sustained damages. However, the relationship between microglia and ROS-regulatory system in the early stages of neuroinflammation prior to the appearance of neuronal damages have not been elucidated in detail. In this study, we analyzed the time-dependent changes in ROS generation during acute neuroinflammation in rats that were given an intrastriatal injection of lipopolysaccharide (LPS). We evaluated the effects of minocycline, an anti-inflammatory antibiotic, and N,N′-dimethylthiourea (DMTU), a radical scavenger, to understand the correlation between activated microglia and ROS generation. Ex vivo fluorescence imaging using dihydroethidium (DHE) clearly demonstrated an increased ROS level in the infused side of striatum in the rats treated with LPS. The level of ROS was changed in time-dependent manner, and the highest level of ROS was observed on day 3 after the infusion of LPS. Immunohistochemical studies revealed that time-dependent changes in ROS generation were well correlated to the presence of activated microglia. The inhibition of microglial activation by minocycline remarkably reduced ROS levels in the LPS-injected striatum, which indicated that the increased ROS generation caused by LPS was induced by activated microglia. DMTU decreased ROS generation and resulted in remarkable inhibitory effect on microglial activation. This study demonstrated that ROS generation during acute neuroinflammation induced by LPS was considerably associated with microglial activation, in an intact rat brain. The results provides a basis for understanding the interaction of ROS-regulatory system and activated microglia during neuroinflammation underlying neurodegenerative diseases.

Abscisic acid and reactive oxygen species were involved in slightly acidic electrolyzed water-promoted seed germination in watermelon

Seed germination is a vital process in seedling stage. The delayed seed germination and irregular emergence frequently occur in watermelon. It is critical to exploit technologies for promoting seed germination. This study aims to explore the effects of slightly acidic electrolyzed water (SAEW) on the seed germination of watermelon. The results showed that SAEW significantly improved the germination indexes and caused a mass of changes to nutrient substances and enzymatic activities in watermelon seeds. In addition, SAEW treatment decreased abscisic acid (ABA) contents, while increased gibberellin (GA) contents of watermelon seeds by regulating the gene expressions of metabolic related enzymes. Thereinto, SAEW remarkably upregulated the transcriptional level of CYP707A1 and CYP707A2 involved in ABA catabolism, which inhibited the accumulation of ABA. Moreover, SAEW could inhibit reactive oxygen species (ROS) accumulation in seeds and improve the seed germination which was identical to the effect of DMTU (dimethyl thiourea, ROS inhibitor) on the seed germination. In addition, the expressions of RbohC and RbohD involved in ROS accumulation were notably depressed by SAEW. This study demonstrated that exogenous SAEW could inhibit the accumulation of ABA content and decrease the accumulation of ROS, which finally improves the germination ability of watermelon seeds.

Coronatine Modulated the Generation of Reactive Oxygen Species for Regulating the Water Loss Rate in the Detaching Maize Seedlings

Coronatine (COR), a structural and functional mimic of jasmonates, is involved in a wide array of effects on plant development and defense response. The present study aims to investigate the role of COR, in counteracting drought stress by modulating reactive oxygen species (ROS) homeostasis, water balance, and antioxidant regulation in detached maize plants. Our results showed that COR can markedly decrease the water loss rate, but the antioxidants diphenyleneiodonium chloride (DPI) and dimethylthiourea (DMTU) eliminate the effect of water loss induced by COR. Using the dye 2′,7′-dichlorofluorescein diacetate (H2DCF-DA) loaded in the maize epidermis guard cells, it is observed that COR could increase ROS production, and then antioxidants DPI and DMTU decreased ROS production induced by COR. In addition, the expression of ZmRBOHs genes, which were associated with ROS generation was increased by COR in levels and ZmRBOHC was highly expressed in the epidermis guard cells. Moreover, COR-treated plants increased H2O2 and O2−· accumulation, antioxidant enzyme activities in control plants, while COR relieved the ROS accumulation and antioxidant enzyme activities under PEG treatment. These results indicated that COR could improve maize performance under drought stress by modulating ROS homeostasis to maintain water loss rate and antioxidant enzyme activities.

Chloroplast thylakoid ascorbate peroxidase PtotAPX plays a key role in chloroplast development by decreasing hydrogen peroxide in Populus tomentosa.

Chloroplast development is a complex process that is critical to the growth and development of plants. However, the detailed mechanism of chloroplast development in woody plants remains unclear. In this study, we showed that chloroplasts with elaborate thylakoids could develop from proplastids in the cells of calli derived from leaf tissues of Populus tomentosa upon exposure to light. Chloroplast development was confirmed at the molecular and cellular levels. Transcriptome analysis revealed that genes related to photoreceptors and photosynthesis were significantly up-regulated during chloroplast development in a time-dependent manner. In light-induced chloroplast development, a key process was the removal of hydrogen peroxide, in which thylakoid-localized PtotAPX played a major role; light-induced chloroplast development was enhanced in PtotAPX-overexpressing transgenic P. tomentosa callus with lower levels of hydrogen peroxide, but was suppressed in PtotAPX antisense transgenic callus with higher levels of hydrogen peroxide. Moreover, the suppression of light-induced chloroplast development in PtotAPX antisense transgenic callus was relieved by the exogenous reactive oxygen species scavenging agent N,N'-dimethylthiourea (DMTU). Based on these results, we propose that PtotAPX-mediated removal of reactive oxygen species plays a key role in chloroplast development from proplastids upon exposure to light in P. tomentosa.

ROS-Dependent Induction of Antioxidant System and Heat Resistance of Wheat Seedlings by Hemin

Effect of 24-h-long treatment with hemin (a donor of carbon monoxide, CO, produced with the participation of heme oxygenase) on heat resistance and the processes of ROS formation and neutralization was investigated in wheat (Triticum aestivum L.) seedlings. It was found that treatment of seedlings with hemin in the concentration range from 0.5 to 10 µM induced an elevation of their resistance to damaging heating (45°C, 10 min). The greatest effect was produced by 5 µM hemin. Hemin treatment was associated with a transient elevation in the roots of extracellular peroxidase activity and a rise in the content of hydrogen peroxide. Elevation of H2O2 content was not observed in the presence of its scavenger dimethyl thiourea (DMTU) and upon treatment of seedlings’ roots with CO absorber hemoglobin and inhibitor of extracellular peroxidase sodium azide; however, this pattern was not followed in the presence of NADPH oxidase inhibitor imidazole. Since decomposition of hemin is associated with the production of redox active Fe2+ ions, the specificity of its action as a CO donor was tested by comparing the effects of hemin and 5 µM FeSO4. At the examined concentration, ferrous sulfate did not affect the generation of hydrogen peroxide in the roots and activity of extracellular peroxidase. In 24 h after the beginning of hemin treatment, activities of superoxide dismutase, catalase, and intracellular peroxidase in the roots rose. Elevation in the activity of antioxidant enzymes induced by hemin was not observed in the presence of DMTU, and treatment of seedling roots with FeSO4 did not modify their activity. Exposure to the CO donor after damaging heating also reduced outflux from the seedling roots of compounds absorbed in the ultraviolet spectral region. Positive influence of hemin on heat resistance of the seedlings was eliminated by CO scavenger hemoglobin and hydrogen peroxide absorber DMTU. Treatment of seedlings with FeSO4 did not alter their survival after heating. Hydrogen peroxide is assumed to participate as a signal mediator in realization of the influence of CO donor on the antioxidant system of wheat seedlings and their heat resistance.