Methylglyoxal as a novel signal molecule induces the salt tolerance of wheat by regulating the glyoxalase system,the antioxidant system,and osmolytes

Reactive carbonyl species methylglyoxal (MG) has always been regarded as a cytotoxic metabolite, but now is emerging to function as signal molecule in plants. However, whether MG can induce salt tolerance is elusive. In this study, treatment of wheat seeds with NaCl reduced seed germination, plant height, root length, fresh weight, and dry weight, indicating the inhibitive effects of NaCl on seed germination and seedling growth. The inhibitive effects of NaCl were alleviated by applying exogenous MG, but aggravated by the MG scavenger N-acetyl-L-cysteine (NAC), suggesting that MG could induce the salt tolerance of wheat. In addition, MG increased glyoxalase I and glyoxalase II activities and decreased endogenous MG content in wheat seedlings under NaCl stress, whereas coapplication of NAC weakened glyoxalase activity and enhanced the endogenous MG level. Also, MG activated superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase activities; increased glutathione and ascorbic acid levels; and decreased superoxide radical production and H₂O₂ and malondialdehyde contents under NaCl stress, while NAC reversed these physiological parameters. Furthermore, MG also induced the accumulation of proline, glycine betaine, and soluble sugar under NaCl stress, whereas this accumulation was weakened by NAC. This work reported for the first time that MG could induce the salt tolerance of wheat, and the acquisition of this salt tolerance was involved in the activation of the glyoxalase system and antioxidant system, as well as the accumulation of osmolytes.     

Role of rpoS in the regulation of glyoxalase III in Escherichia coli

Methylglyoxal is an endogenous electrophile produced in Escherichia coli by the enzyme methylglyoxal synthase to limit the accumulation of phosphorylated sugars. In enteric bacteria methylglyoxal is detoxified by the glutathione-dependent glyoxalase I/II system, by glyoxalase III, and by aldehyde reductase and alcohol dehydrogenase. Here we demonstrate that glyoxalase III is a stationary-phase enzyme. Its activity reached a maximum at the entry into the stationary phase and remained high for at least 20 h. An rpoS- mutant displayed normal glyoxalase I and II activities but was unable to induce glyoxalase III in stationary phase. It thus appears that glyoxalase III is regulated by rpoS and might be important for survival of non-growing E. coli cultures.     

Signaling molecule methylglyoxal-induced thermotolerance is partly mediated by hydrogen sulfide in maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) seedlings

Methylglyoxal (MG) was traditionally viewed as toxic by-product of glycolysis and photosynthesis in plants, but now is emerging as a signaling molecule, which, similar to hydrogen sulfide (H₂S), participates in regulating seed germination, growth, development, and response to abiotic stress. However, whether exists an mutual effect between MG and H₂S in improving thermotolerance in plants is not found to be reported. In this paper, interplay between MG and H₂S in the formation of thermotolerance in maize seedlings was investigated. The results indicated that MG pretreatment elevated the survival percentage of maize seedlings under high-temperature stress, manifesting that MG could boost the thermotolerance of maize seedlings. Interestingly, MG-induced thermotolerance was reinforced by sodium hydrosulphide (NaHS, H₂S donor), while impaired by dl-propargylglycine (inhibitor of H₂S biosynthesis) and hypotaurine (scavenger of H₂S), respectively. In addition, H₂S could induce the thermotolerance of maize seedlings, which was impaired by aminoguanidine (AG) and N-acetyl-l-cysteine (NAC) (MG scavengers), respectively. Furthermore, MG stimulated the activity of a key enzyme in H₂S biosynthesis, l-cysteine desulfhydrase, which, in turn, triggered the elevation of endogenous H₂S in maize seedlings. In addition, H₂S increased the level of endogenous MG; this increase was crippled by AG and NAC. This paper, for the first time, reported that MG could improve the thermotolerance of maize seedlings, and its acquisition was, at least partly, mediated by H₂S.     

2-Oxoaldehyde metabolism in microorganisms

The properties of methylglyoxal-metabolizing enzymes in prokaryotic and eukaryotic microorganisms were studied systematically and compared with those of mammalian enzymes. The enzymes constitute a glycolytic bypass and convert methylglyoxal into pyruvate via lactate. The first step in this conversion is catalyzed by glyoxalase I, methylglyoxal reductase, or methylglyoxal dehydrogenase. The regulation of the yeast glyoxalase system was analyzed. The system was closely related to the proliferative states of yeast cells, the activity of the system being high in dividing cells and low in nondividing ones. The gene for the glyoxalase I of Pseudomonas putida and the genes responsible for the activity of glyoxalase I and methylglyoxal reductase in Saccharomyces cerevisiae were cloned and their structural and phenotypic characters studied.     

Proline and betaine provide protection to antioxidant and methylglyoxal detoxification systems during cold stress in <Emphasis Type="Italic">Camellia sinensis</Emphasis> (L.) O. Kuntze

The aim of this study was to monitor the influence of proline and betaine exposure on antioxidant and methylglyoxal (MG) detoxification system during cold stress in Camellia sinensis (L.) O. Kuntze. Cold stress enhanced MG and lipid peroxidation levels in tea bud (youngest topmost leaf). This increase was resisted upon the exposure of tea bud to proline and betaine. Exposure of tea bud with proline and betaine also help in maintaining thiol/disulfide ratio during cold stress. Proline exposure enhanced glutathione-S-transferase and glutathione reductase (GR) activity, while betaine exposure increased only GR activity during cold stress. Furthermore, effect of proline/betaine was studied on glyoxalase pathway enzymes that are involved in MG detoxification and comprise of two enzymes glyoxalase I and glyoxalase II. Both proline and betaine showed protective effect on glyoxalase I and activating effect on glyoxalase II during cold stress in tea bud. This investigation, therefore, suggest that proline and betaine might provide protection to cold stress in tea by regulating MG and lipid peroxidation formation as well as by activating or protecting some of antioxidant and glyoxalase pathway enzymes.     

外源甲基乙二醛对干旱胁迫下板栗幼苗的影响

甲基乙二醛(MG)是一种在植物中具有多种功能的新型信号分子.为探究MG对板栗幼苗干旱胁迫的影响,以两年生'黄棚'板栗幼苗为试材,通过聚乙二醇(PEG)模拟干旱胁迫并进行MG及其清除剂N-乙酰半胱氨酸(NAC)处理,分析板栗幼苗叶片超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)、抗坏血酸过氧化物酶(...     

Paclobutrazol mitigates salt stress in <Emphasis Type="Italic">indica</Emphasis> rice seedlings by enhancing glutathione metabolism and glyoxalase system

Stress-induced methylglyoxal (MG) functions as a toxic molecule, inhibiting plant physiological processes such as photosynthesis and antioxidant defense systems. In the present study, an attempt was made to investigate the MG detoxification through glutathione metabolism in indica rice [Oryza sativa L. ssp. indica cv. Pathumthani 1] under salt stress by exogenous foliar application of paclobutrazol (PBZ). Fourteen-day-old rice seedlings were pretreated with 15 mg L^?1 PBZ foliar spray. After 7 days, rice seedlings were subsequently exposed to 0 (control) or 150 mM NaCl (salt stress) for 12 days. Prolonged salt stress enhanced the production of MG molecules and the oxidation of proteins, leading to decreased activity of glyoxalase enzymes, glyoxalase I (Gly I) and glyoxalase II (Gly II). Consequently, the decreased glyoxalase activities were also associated with a decline in reduced glutathione (GSH) content and glutathione reductase (GR) activity. PBZ pretreatment of rice seedlings under salt stress significantly lowered MG production and protein oxidation, and increased the activities of both Gly I and Gly II. PBZ also increased GSH content and GR activity along with the up-regulation of glyoxalase enzymes, under salt stress. In summary, salinity induced a high level of MG and the associated oxidative damage, while PBZ application reduced the MG toxicity by up-regulating glyoxalase and glutathione defense system in rice seedlings.     

热激诱导的玉米幼苗耐热性及其与脯氨酸的关系

研究了热激对玉米幼苗耐热性的效应及其与脯氨酸的关系。结果表明,培养2.5 d的玉米幼苗经过42℃热激4 h并于26.5℃下恢复4 h后,提高了玉米幼苗在48℃下的存活率,并且热激及其后的恢复过程中都表现出脯氨酸的积累。不同浓度的外源脯氨酸预处理也可提高玉米幼苗内源脯氨酸的水平和抗氧化酶抗坏血酸过氧化物酶(APX)、过氧化氢酶(CAT)、超氧化物歧化酶(SOD)、谷胱甘肽还原酶(GR)、过氧化物酶(GPX)的活性,从而提高玉米幼苗在高温胁迫下的存活率。这些结果暗示热激过程中脯氨酸的积累所诱发的抗氧化酶活性的增强可能是热激诱导的玉米幼苗耐热性形成的生理基础之一。     

Manganese-induced salt stress tolerance in rice seedlings: regulation of ion homeostasis,antioxidant defense and glyoxalase systems

Hydroponically grown 12-day-old rice (Oryza sativa L. cv. BRRI dhan47) seedlings were exposed to 150 mM NaCl alone and combined with 0.5 mM MnSO₄. Salt stress resulted in disruption of ion homeostasis by Na⁺ influx and K⁺ efflux. Higher accumulation of Na⁺ and water imbalance under salinity caused osmotic stress, chlorosis, and growth inhibition. Salt-induced ionic toxicity and osmotic stress consequently resulted in oxidative stress by disrupting the antioxidant defense and glyoxalase systems through overproduction of reactive oxygen species (ROS) and methylglyoxal (MG), respectively. The salt-induced damage increased with the increasing duration of stress. However, exogenous application of manganese (Mn) helped the plants to partially recover from the inhibited growth and chlorosis by improving ionic and osmotic homeostasis through decreasing Na⁺ influx and increasing water status, respectively. Exogenous application of Mn increased ROS detoxification by increasing the content of the phenolic compounds, flavonoids, and ascorbate (AsA), and increasing the activities of monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), superoxide dismutase (SOD), and catalase (CAT) in the salt-treated seedlings. Supplemental Mn also reinforced MG detoxification by increasing the activities of glyoxalase I (Gly I) and glyoxalase II (Gly II) in the salt-affected seedlings. Thus, exogenous application of Mn conferred salt-stress tolerance through the coordinated action of ion homeostasis and the antioxidant defense and glyoxalase systems in the salt-affected seedlings.     

Chlorate Reducing Activity of Spinach Nitrate Reductase

The activities of 2-oxoaldehyde-metabolizing enzymes (glyoxalase I, glyoxalase II, methyl- glyoxal reductase, methylglyoxal dehydrogenase and lactaldehyde dehydrogenase) were found to be widely distributed among microorganisms. One of the enzymes, methylglyoxal reductase, which catalyzes the reductive conversion of methylglyoxal into lactaldehyde, was purified from Escherichia coli cells. The enzyme was judged to be homogeneous on polyacrylamide gel electrophoresis and was a monomer with a molecular weight of 43000. The enzyme was most active at pH 6.5 and 45°C. The enzyme utilized both NADPH and NADH for the reduction of 2- oxoaldehydes (glyoxal, methylglyoxal, phenylglyoxal and 4,5-dioxovalerate) and some aldehydes (glycolaldehyde, D,l-glyceraldehyde, propionaldehyde and acetaldehyde). The Km values of the enzyme for methylglyoxal, NADPH and NADH were 4.0 mm, 1.7 fiM and 2.8 /¿m, respectively. The product of methylglyoxal reduction was identified as lactaldehyde. The enzyme from E. coli cells was different from the yeast and goat liver enzymes in both molecular structure and substrate specificity.     

Salicylic acid dependent signaling promotes basal thermotolerance but is not essential for acquired thermotolerance in Arabidopsis thaliana

Salicylic acid (SA) is reported to protect plants from heat shock (HS), but insufficient is known about its role in thermotolerance or how this relates to SA signaling in pathogen resistance. We tested thermotolerance and expression of pathogenesis-related (PR) and HS proteins (HSPs) in Arabidopsis thaliana genotypes with modified SA signaling: plants with the SA hydroxylase NahG transgene, the nonexpresser of PR proteins (npr1) mutant, and the constitutive expressers of PR proteins (cpr1 and cpr5) mutants. At all growth stages from seeds to 3-week-old plants, we found evidence for SA-dependent signaling in basal thermotolerance (i.e. tolerance of HS without prior heat acclimation). Endogenous SA correlated with basal thermotolerance, with the SA-deficient NahG and SA-accumulating cpr5 genotypes having lowest and highest thermotolerance, respectively. SA promoted thermotolerance during the HS itself and subsequent recovery. Recovery from HS apparently involved an NPR1-dependent pathway but thermotolerance during HS did not. SA reduced electrolyte leakage, indicating that it induced membrane thermoprotection. PR-1 and Hsp17.6 were induced by SA or HS, indicating common factors in pathogen and HS responses. SA-induced Hsp17.6 expression had a different dose-response to PR-1 expression. HS-induced Hsp17.6 protein appeared more slowly in NahG. However, SA only partially induced HSPs. Hsp17.6 induction by HS was more substantial than by SA, and we found no SA effect on Hsp101 expression. All genotypes, including NahG and npr1, were capable of expression of HSPs and acquisition of HS tolerance by prior heat acclimation. Although SA promotes basal thermotolerance, it is not essential for acquired thermotolerance.     

Exogenous nitric oxide donor and arginine provide protection against short-term drought stress in wheat seedlings

Nitric oxide (NO) is an important plant signaling molecule that has a vital role in abiotic stress tolerance. In the present study, we assessed drought-induced (15 and 30% PEG, polyethylene glycol) damage in wheat (Triticum aestivum L. cv. Prodip) seedlings and mitigation by the synergistic effect of exogenous Arg (0.5 mM l-Arginine) and an NO donor (0.5 mM sodium nitroprusside, SNP). Drought stress sharply decreased the leaf relative water content (RWC) but markedly increased the proline (Pro) content in wheat seedlings. Drought stress caused overproduction of reactive oxygen species (ROS) and methylglyoxal (MG) due to the inefficiency of antioxidant enzymes, the glyoxalase system, and the ascorbate-glutathione pool. However, supplementation with the NO donor and Arg enhanced the antioxidant defense system (both non-enzymatic and enzymatic components) in drought-stressed seedlings. Application of the NO donor and Arg also enhanced the glyoxalase system and reduced the MG content by increasing the activities of the glyoxalase system enzymes (Gly I and Gly II), which restored the leaf RWC and further increased the Pro content under drought stress conditions. Exogenous NO donor and Arg application enhanced the endogenous NO content, which positively regulated the antioxidant system and reduced ROS production. Thus, the present study reveals the crucial roles of Arg and NO in enhancing drought stress tolerance in wheat seedlings by upgrading their water status and reducing oxidative stress and MG toxicity.     

Hydrogen sulfide donor sodium hydrosulfide-improved heat tolerance in maize and involvement of proline

Hydrogen sulfide (H₂S) has long been considered as a phytotoxin, but nowadays as a cell signal molecule involved in growth, development, and the acquisition of stress tolerance in higher plants. In the present study, hydrogen sulfide donor, sodium hydrosulfide (NaHS), pretreatment markedly improved germination percentage of seeds and survival percentage of seedlings of maize under heat stress, and alleviated an increase in electrolyte leakage of roots, a decrease in tissue vitality and an accumulation of malondialdehyde (MDA) in coleoptiles of maize seedlings. In addition, pretreatment of NaHS could improve the activity of Δ¹-pyrroline-5-carboxylate synthetase (P5CS) and lower proline dehydrogenase (ProDH) activity, which in turn induced accumulation of endogenous proline in maize seedlings. Also, application of proline could enhance endogenous proline content, followed by mitigated accumulation of MDA and increased survival percentage of maize seedlings under heat stress. These results suggest that sodium hydrosulfide pretreatment could improve heat tolerance of maize and the acquisition of this heat tolerance may be involved in proline.     

Protective Effect of Betaine on Respiratory Enzymes

Effects of betaine and NaC1 in various concentrations on the activities of enzymes in tricarboxylic acid cycle (isocitric dehydrogenase, malic dehydrogenase, succinic dehydrogenase and fumarase), terminal oxidation (cytochrome oxidase) and photorespiratory pathway (glycolate oxidase and hydroxypyruvate reductase) have been studied. Betaine, in contrast to electolyte NaC1 was non-inhibitory to these enzymes up to 500 mmol/L. Partial protection against NaC1 inhibition to the activities of these enzymes were afforded by betaine. These results were consistent with the postulated role of betaine in cytoplasmic osmoregulation. These results showed that betaine was a superior compatible solute.     

甜菜碱对呼吸酶的保护效应

以菠菜（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）叶片为材料,研究了不同浓度的甜菜碱和ＮａＣｌ对三羧酸循环、末端氧化和光呼吸的组成酶的活性的影响。与电解质ＮａＣｌ不同,高浓度的甜菜碱对这些酶的活性是非抑制性的,并对ＮａＣｌ的抑制作用有一定保护效应。甜菜碱是很好的有机渗透调节剂。这与甜菜碱在细胞质中起渗透调节作用,以及是无机渗透调节剂的配伍溶质的假设是一致的。     

Genetic manipulation of Japonica rice using the <Emphasis Type="Italic">OsBADH1</Emphasis> gene from Indica rice to improve salinity tolerance

Betaine aldehyde dehydrogenase (BADH) is a major oxidative enzyme that converts betaine aldehyde to glycine betaine (GB), an osmoprotectant compound in plants. Japonica rice (salt-sensitive) was genetically engineered to enhance salt tolerance by introducing the OsBADH1 gene from Indica rice (salt-tolerant), which is a GB accumulator. We produced transgenic rice plants overexpressing the modified OsBADH1 gene under the control of the maize ubiquitin promoter. The transgenic rice showed increased OsBADH1 gene expression and OsBADH1 enzyme production, resulting in the accumulation of GB. It also exhibited enhanced salt tolerance in immature and mature transgenic rice seedlings. The adverse effect of salt stress on seed germination, the growth of immature and mature seedlings, water status, and photosynthetic pigments was alleviated in transgenic seedlings.     

Modulation of nitrosative stress by S-nitrosoglutathione reductase is critical for thermotolerance and plant growth in Arabidopsis

Nitric oxide (NO) is a key signaling molecule in plants. This analysis of Arabidopsis thaliana HOT5 (sensitive to hot temperatures), which is required for thermotolerance, uncovers a role of NO in thermotolerance and plant development. HOT5 encodes S-nitrosoglutathione reductase (GSNOR), which metabolizes the NO adduct S-nitrosoglutathione. Two hot5 missense alleles and two T-DNA insertion, protein null alleles were characterized. The missense alleles cannot acclimate to heat as dark-grown seedlings but grow normally and can heat-acclimate in the light. The null alleles cannot heat-acclimate as light-grown plants and have other phenotypes, including failure to grow on nutrient plates, increased reproductive shoots, and reduced fertility. The fertility defect of hot5 is due to both reduced stamen elongation and male and female fertilization defects. The hot5 null alleles show increased nitrate and nitroso species levels, and the heat sensitivity of both missense and null alleles is associated with increased NO species. Heat sensitivity is enhanced in wild-type and mutant plants by NO donors, and the heat sensitivity of hot5 mutants can be rescued by an NO scavenger. An NO-overproducing mutant is also defective in thermotolerance. Together, our results expand the importance of GSNOR-regulated NO homeostasis to abiotic stress and plant development.     

Enhancing salt tolerance in a crop plant by overexpression of glyoxalase II

Earlier we have shown the role of glyoxalase overexpression in conferring salinity tolerance in transgenic tobacco. We now demonstrate the feasibility of same in a crop like rice through overproduction of glyoxalase II. The rice glyoxalase II was cloned in pCAMBIA1304 and transformed into rice (Oryza sativa cv PB1) via Agrobacterium. The transgenic plants showed higher constitutive activity of glyoxalase II that increased further upon salt stress, reflecting the upregulation of endogenous glyoxalase II. The transgenic rice showed higher tolerance to toxic concentrations of methylglyoxal (MG) and NaCl. Compared with non-transgenics, transgenic plants at the T1 generation exhibited sustained growth and more favorable ion balance under salt stress conditions. Sneh L. Singla-Pareek and Sudesh Kumar Yadav have contributed equally to this work.     

Engineering of enhanced glycine betaine synthesis improves drought tolerance in maize

Glycine betaine plays an important role in some plants, including maize, in conditions of abiotic stress, but different maize varieties vary in their capacity to accumulate glycine betaine. An elite maize inbred line DH4866 was transformed with the betA gene from Escherichia coli encoding choline dehydrogenase (EC 1.1.99.1), a key enzyme in the biosynthesis of glycine betaine from choline. The transgenic maize plants accumulated higher levels of glycine betaine and were more tolerant to drought stress than wild-type plants (non-transgenic) at germination and the young seedling stage. Most importantly, the grain yield of transgenic plants was significantly higher than that of wild-type plants after drought treatment. The enhanced glycine betaine accumulation in transgenic maize provides greater protection of the integrity of the cell membrane and greater activity of enzymes compared with wild-type plants in conditions of drought stress.