Characterization of post-flooding recovery-responsive enzymes in soybean root and hypocotyl

Soybean exhibits markedly reduced growth and yields under flooding stress. To determine the functional roles of four soybean proteins in post-flooding recovery, the organ/stress specificity and time-dependency of their enzymatic activities were analyzed. Peroxidase activity decreased in root and hypocotyl exposed to flooding and cold stresses, but increased during the post-stress recovery period. In contrast, its activity increased in both root and hypocotyl under drought stress. Acid phosphatase activity was suppressed in root treated with flooding and cold stresses, and slightly increased during the recovery period; however, the opposite profile was observed in hypocotyl. In response to drought stress, it did not change in root, but was decreased in hypocotyl. Beta-ketoacyl reductase activity did not change in root under flooding conditions, but was decreased in hypocotyl, although the activity increased slightly during the recovery period. In addition, it was decreased in both organs under drought and cold stresses, but again increased during the recovery period. Nucleotidylyl transferase activity was increased in root under flooding and drought stresses, but was decreased in hypocotyl. It was decreased in response to cold stress, but exhibited a slight increase during the recovery period. Furthermore, the treatment with jasmonate and salicylate suppressed the activities of peroxidase and acid phosphatase in root and hypocotyl under flooding stress; however, the activity of acid phosphatase increased during the recovery period. Nucleotidylyl transferase activity in root was also elevated by treatment with jasmonate, but gradually decreased during the recovery period. These results suggest that jasmonate-induced changes in nucleotidylyl transferase activity may facilitate soybean root recovery after flooding stress.     

Characterization of S-adenosylmethionine synthetases in soybean under flooding and drought stresses

Soybean is stress-sensitive crop that exhibits markedly reduced growth under flooding and drought conditions. Three S-adenosylmethionine synthetases (SAMs) proteins were identified as flooding and drought responsive proteins in soybean using a proteomic technique. To better understand the role of these SAMs proteins in soybean under flooding and drought stresses, temporal, organ, and stress specificities were examined at mRNA and enzyme activity levels. The activity of SAMs decreased in response to the flooding, however, it was not significantly changed by NaCl, cold, gibberellic acid, and calcium in soybean roots. The activity of SAMs was induced in roots and hypocotyls under drought. The mRNA expression of the S-adenosylmethionine synthetase (SAMs) family was down-regulated in root tips and roots under the flooding and the drought, and SAMs 1 and SAMs 2 were down-regulated in roots under both stresses. A gene 1-aminocyclopropane-1-carboxylate synthase was up-regulated in root tips, roots, and hypocotyls under drought, however, it was not changed in root tips and roots under the flooding. In addition, 1-aminocyclopropane-1-carboxylate oxidase was induced in root tips under flooding and drought. These results suggest that SAMs was involved in the response to the flooding and drought and it might affect ethylene biosynthesis in soybean.     

中华蚊母树在干旱-水淹交叉胁迫下形态和活性氧代谢的适应机制

为阐明中华蚊母树(Distylium chinense)在消落带干旱-水淹交叉胁迫下的形态和活性氧(ROS)代谢适应机制,通过控制实验模拟了三峡水库消落带的水文节律,研究了干旱-水淹交叉胁迫及恢复过程施加不同外源物质对中华蚊母树形态学和ROS清除的变化。结果表明：(1)前期干旱胁迫增强了中华蚊母树对后期水淹胁迫的适应,主要表现在叶片脱落、大量不定根的形成及茎基部膨大等形态学的变化;(2)干旱或水淹单一胁迫下,中华蚊母树·OH、■等ROS水平明显高于对照,表现出氧化应激反应,其超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、抗坏血酸过氧化物酶(APX)等抗氧化系统酶活性及脯氨酸(Pro)等抗氧化系统小分子含量也均显著高于对照,表现出一定的抗氧化防御作用机制,且在复合胁迫下,SOD、CAT、APX酶活性及Pro含量显著高于单一胁迫;(3)恢复阶段,相关性分析表明,中华蚊母树清除ROS(·OH、■的酶促(SOD、CAT、APX)及非酶促(Pro)系统具有一定的协同性。同时,恢复阶段施加脱落酸(ABA),内源Pro显著高于正常水平;施加Pro, SOD、CAT等抗氧化酶活性显著高于对照;施加可...     

Analysis of flooding-responsive proteins localized in the nucleus of soybean root tips

Flooding stress has negative impact on soybean cultivation as it severely impairs plant growth and development. To examine whether nuclear function is affected in soybean under flooding stress, abundance of nuclear proteins and their mRNA expression were analyzed. Two-day-old soybean seedlings were treated with flooding for 2 days, and nuclear proteins were purified from root tips. Gel-free proteomics analysis identified a total of 39 flooding-responsive proteins, of which abundance of 8 and 31 was increased and decreased, respectively, in soybean root tips. Among these differentially regulated proteins, the mRNA expression levels of five nuclear-localized proteins were further analyzed. The mRNA levels of four proteins, which are splicing factor PWI domain-containing protein, epsilon2-COP, beta-catenin, and clathrin heavy chain decreased under flooding stress, were also down-regulated. In addition, mRNA level of a receptor for activated protein kinase C1(RACK1) was down-regulated, though its protein was accumulated in the soybean nucleus in response to flooding stress. These results suggest that several nuclear-related proteins are decreased at both the protein and mRNA level in the root tips of soybean under flooding stress. Furthermore, RACK1 might have an important role with accumulation in the soybean nucleus under flooding-stress conditions.     

Physiological integration enhanced the tolerance of Cynodon dactylon to flooding

Many flooding‐tolerant species are clonal plants; however, the effects of physiological integration on plant responses to flooding have received limited attention. We hypothesise that flooding can trigger changes in metabolism of carbohydrates and ROS (reactive oxygen species) in clonal plants, and that physiological integration can ameliorate the adverse effects of stress, subsequently restoring the growth of flooded ramets. In the present study, we conducted a factorial experiment combining flooding to apical ramets and stolon severing (preventing physiological integration) between apical and basal ramets of Cynodon dactylon, which is a stoloniferous perennial grass with considerable flooding tolerance. Flooding‐induced responses including decreased root biomass, accumulation of soluble sugar and starch, as well as increased activity of superoxide dismutase (SOD) and ascorbate peroxidase (APX) in apical ramets. Physiological integration relieved growth inhibition, carbohydrate accumulation and induction of antioxidant enzyme activity in stressed ramets, as expected, without any observable cost in unstressed ramets. We speculate that relief of flooding stress in clonal plants may rely on oxidising power and electron acceptors transferred between ramets through physiological integration.     

抗旱性不同的小麦幼苗对水分和NaCl胁迫的反应

分别测定抗旱小麦的８１３９（Ｔｒｉｔｉｃｕｍ ａｅｓｔｉｖｕｍ Ｌ．ｃｖ．８１３９）和干旱敏感品种甘麦８号（Ｔ．ａｅｓｔｉｖｕｍ Ｌ．ｃｖ．Ｇａｎｍａｉ Ｎｏ．８）在２０％ＰＥＧ６０００和１．２％ＮａＣｌ胁迫下的生长、光合作用、蒸腾作用及抗氧化保护系统的变化。结果表明,抗旱小麦８１３９对ＰＥＧ６０００有较强的抗性,但对ＮａＣｌ胁迫的抗性较差。ＮａＣｌ胁迫下,两种小麦根的生长均受到严重抑制,而在ＰＥ     

Ascorbate peroxidase 1 plays a key role in the response of Arabidopsis thaliana to stress combination

Within their natural habitat plants are subjected to a combination of different abiotic stresses, each with the potential to exacerbate the damage caused by the others. One of the most devastating stress combinations for crop productivity, which frequently occurs in the field, is drought and heat stress. In this study we conducted proteomic and metabolic analysis of Arabidopsis thaliana plants subjected to a combination of drought and heat stress. We identified 45 different proteins that specifically accumulated in Arabidopsis in response to the stress combination. These included enzymes involved in reactive oxygen detoxification, malate metabolism, and the Calvin cycle. The accumulation of malic enzyme during the combined stress corresponded with enhanced malic enzyme activity, a decrease in malic acid, and lower amounts of oxaloacetate, suggesting that malate metabolism plays an important role in the response of Arabidopsis to the stress combination. Cytosolic ascorbate peroxidase 1 (APX1) protein and mRNA accumulated during the stress combination. When exposed to heat stress combined with drought, an APX1-deficient mutant (apx1) accumulated more hydrogen peroxide and was significantly more sensitive to the stress combination than wild type. In contrast, mutants deficient in thylakoid or stromal/mitochondrial APXs were not more sensitive to the stress combination than apx1 or wild type. Our findings suggest that cytosolic APX1 plays a key role in the acclimation of plants to a combination of drought and heat stress.     

Effects of drought stress on legume symbiotic nitrogen fixation: physiological mechanisms

Drought stress is one of the major factors affecting nitrogen fixation by legume-rhizobium symbiosis. Several mechanisms have been previously reported to be involved in the physiological response of symbiotic nitrogen fixation to drought stress, i.e. carbon shortage and nodule carbon metabolism, oxygen limitation, and feedback regulation by the accumulation of N fixation products. The carbon shortage hypothesis was previously investigated by studying the combined effects of CO2 enrichment and water deficits on nodulation and N2 fixation in soybean. Under drought, in a genotype with drought tolerant N2 fixation, approximately four times the amount of 14C was allocated to nodules compared to a drought sensitive genotype. It was found that an important effect of CO2 enrichment of soybean under drought was an enhancement of photo assimilation, an increased partitioning of carbon to nodules, whose main effect was to sustain nodule growth, which helped sustain N2 rates under soil water deficits. The interaction of nodule permeability to O2 and drought stress with N2 fixation was examined in soybean nodules and led to the overall conclusion that O2 limitation seems to be involved only in the initial stages of water deficit stresses in decreasing nodule activity. The involvement of ureides in the drought response of N2 fixation was initially suspected by an increased ureide concentration in shoots and nodules under drought leading to a negative feedback response between ureides and nodule activity. Direct evidence for inhibition of nitrogenase activity by its products, ureides and amides, supported this hypothesis. The overall conclusion was that all three physiological mechanisms are important in understanding the regulation of N2 fixation and its response of to soil drying.     

Does a terminal drought tolerance QTL contribute to differences in ROS scavenging enzymes and photosynthetic pigments in pearl millet exposed to drought?

The control of reactive oxygen species (ROS) and the stability of photosynthetic pigments under stress conditions are hypothesized to contribute to drought tolerance. Here we studied how ascorbic peroxidase (APX), superoxide dismutase (SOD), catalase (CAT) isozyme activities and chlorophyll a, b (Chl a, b) and carotenoids (Car) contents responded to water stress and whether they related to presence of a terminal drought tolerance QTL in pearl millet. We used PRLT2/89-33 (QTL donor), H77/833-2 (sensitive), and near-isogenic lines (QTL-NILs) introgressed with the QTL in H77/833-2 background. Under water stress there was no significant change in the total APX activity; only the proportional APX5 activity increased, with higher band intensity in tolerant genotypes. There were no significant changes in total activities of CAT and SOD under water stress, with similar band intensities in all genotypes, and a new CAT isozyme was induced in all genotypes. The photosynthetic pigment content decreased under water stress, although not differently in any genotype. Under water stress, the activities of most APX, CAT and SOD isozymes were closely related to the total chlorophyll/carotenoids ratio. Overall, besides APX5, water stress did not lead to major changes in the profile of isoenzymes involved in ROS scavenging. Similarly, the pigment content under stress did not discriminate genotypes according to the presence/absence of the QTL. This absence of discrimination for the ROS scavenging enzymes and for the pigment content under stress suggests that these traits may not play a key role in terminal drought tolerance in pearl millet.     

Drought stress enhanced andrographolides contents in Andrographis paniculata

Andrographis paniculata (AP) is an important medicinal plant in South China, but it is often exposed to drought and flooding stress. Here three drought stresses and two flooding stresses with three treated time were imposed on potted AP to explore the water stress effects. The growth physiology characteristics and major effective ingredients (andrographolide (AG), 14-deoxy-11,12-didehydroandrographolide (DDAG), and total andrographolides (AGs)) accumulations were studied. Results showed that water stress treated time was statistically effective while its interactions with treatment were not, except for a few parameters. Leaf area and stem diameter of water stressed plants were not statistically differentiated from control all the time, but the plant height, root length, dry weight, and root-shoot ratio were significantly affected by severe drought/flooding stress in prolonged treated time. Significant bioactive component changes were only occurred in the longest treated time under severe stresses, especially those of DDAG and AGs. Results also showed that the plant dry weight were relatively well correlated with andrographolides contents when expressed on ‘per plant’ basis. The underlying mechanisms behind these responses were discussed.     

Comparative proteomics analysis of differentially expressed proteins in soybean cell wall during flooding stress

Flooding is a major problem for soybean crop as it reduces the growth and grain yield. To investigate the function of the soybean cell wall in the response to flooding stress, cell wall proteins were analyzed. Cell wall proteins from roots and hypocotyls of soybeans, which were germinated for 2 days and subjected to 2 days of flooding, were purified, separated by two-dimensional polyacrylamide gel electrophoresis and stained with Coomassie brilliant blue. Sixteen out of 204 cell wall proteins showed responses to flooding stress. Of these, two lipoxygenases, four germin-like protein precursors, three stem 28/31 kDa glycoprotein precursors, and one superoxide dismutase [Cu–Zn] were downregulated. A copper amine oxidase was found to have shifted from the basic to acidic zone following flooding stress. Based on these results, it was confirmed by the lignin staining that the lignification was suppressed in the root of soybean under the flooding stress. These results suggest that the roots and hypocotyls of soybean caused the suppression of lignification through decrease of these proteins by downregulation of reactive oxygen species and jasmonate biosynthesis under flooding stress.     

Proteomic and Biochemical Analyses of the Cotyledon and Root of Flooding-Stressed Soybean Plants

Background

Flooding significantly reduces the growth and grain yield of soybean plants. Proteomic and biochemical techniques were used to determine whether the function of cotyledon and root is altered in soybean under flooding stress.

Results

Two-day-old soybean plants were flooded for 2 days, after which the proteins from root and cotyledon were extracted for proteomic analysis. In response to flooding stress, the abundance of 73 and 28 proteins was significantly altered in the root and cotyledon, respectively. The accumulation of only one protein, 70 kDa heat shock protein (HSP70) (Glyma17g08020.1), increased in both organs following flooding. The ratio of protein abundance of HSP70 and biophoton emission in the cotyledon was higher than those detected in the root under flooding stress. Computed tomography and elemental analyses revealed that flooding stress decreases the number of calcium oxalate crystal the cotyledon, indicating calcium ion was elevated in the cotyledon under flooding stress.

Conclusion

These results suggest that calcium might play one role through HSP70 in the cotyledon under flooding stress.     

Effects of Drought Stress on Key Enzymes of Carbon Metabolism,Photosynthetic Characteristics and Agronomic Traits of Soybean at the Flowering Stage under Different Soil Substrates

Soybean is an important legume food crop, and its seeds are rich in nutrients, providing humans and animals with edible oil and protein feed. However, soybean is sensitive to water requirements, and drought is an important factor limiting soybean yield and quality. This study used Heinong 84 (drought resistant variety) and Hefeng 46 (intermediate variety) as tested varieties planted in chernozem, albic, and black soils. The effects of drought stress on the activities of key enzymes in carbon metabolism and photosynthetic characteristics of soybean were studied during the flowering stage, most sensitive to water. (1) The activities of SS-1, 6PGDH, and G6PDH enzymes in soybean leaves first increased and then decreased under drought stress. The enzyme activity was the highest under moderate drought stress and weakest in the blank group. (2) Drought stress increased Phi2, PhiNO, and Fm in soybean leaves and reached the highest value under severe drought; with the increase in drought stress, PhiNPQ and Fv/Fm of soybean leaves gradually decreased, reaching the lowest under severe drought. (3) With the increase in drought stress, F₀ and Fs of soybean leaves showed a single peak curve, and the maximum was at moderate drought. (4) Correlation analysis showed that F₀ was greatly affected by varieties and soil types; Fs, F0, and Fm soil varieties had a great influence, and chlorophyll fluorescence parameters were affected differently under drought stress with different drought degrees. (5) Drought stress changed the agronomic traits and yield of soybean. With the increase of drought degree, plant height, node number of main stem, effective pod number, 100-seed weight and total yield decreased continuously. (6) Drought stress affected the dry matter accumulation of soybean. With the increase of drought degree, the dry matter accumulation gradually decreased. Among them, the leaf was most seriously affected by drought, and SD decreased by about 55% compared with CK. Under the condition of black soil, the dry matter accumulation of soybean was least affected by drought.     

Drought tolerance in cereals in terms of water retention,photosynthesis and antioxidant enzyme activities

Experiments were carried out on three bread wheat varieties, one barley and one durum wheat variety grown in pots in the phytotron and subjected to water withdrawal for 7 days during grain-filling. Leaf water loss, net assimilation rate and transpiration showed marked differences, allowing the genotypes to be ranked. Although the most resistant variety had the highest activity for ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR) and glutathione-S-transferase (GST), which did not rise further in response to drought and the most susceptible variety had the lowest values, which increased to the greatest extent under drought, the level of sensitivity could not be predicted for all the genotypes from the enzyme activity values alone. The largest increases were recorded for the APX, CAT and GR activities. In most genotypes the GR activity was correlated with that of GST, CAT and APX. Changes in the enzyme activities were observed after a decline in transpiration and photosynthesis. The range of soil moisture values over which the antioxidant enzyme activity levels remained relatively unchanged was a better indication of tolerance to drought than either basic or stress-induced activity levels.     

四季桂抗氧化防御系统对干旱、高温及协同胁迫的响应

以天香台阁四季桂(Osmanthus fragrans cv. ‘Tian Xiang TaiGe’)为材料, 研究干旱(轻度、中度和重度)、高温(40°C)及干旱高温协同胁迫对四季桂叶片抗氧化防御系统的影响。结果显示, 干旱胁迫下, 四季桂活性氧(ROS)逐渐积累, 膜脂过氧化程度加深; 轻度和中度干旱胁迫下, 抗氧化酶活性显著升高; 重度干旱胁迫下, 抗坏血酸(AsA)及其还原力(AsA/DHA)显著降低, 谷胱甘肽(GSH)及其还原力(GSH/GSSG)以及抗坏血酸-谷胱甘肽(AsA-GSH)循环相关酶活性呈先上升后下降的趋势, 在中度干旱胁迫时达到峰值。高温胁迫显著增强ROS积累、抗氧化酶活性、抗氧化剂含量及AsA-GSH循环效率。干旱高温协同胁迫下, 四季桂所受伤害大于单一胁迫, ROS在抗氧化酶的作用下增幅减缓; 随着胁迫强度的加剧, AsA-GSH循环效率呈先增加后下降的趋势, 重度协同胁迫时显著降低, 无法维持氧化还原平衡。四季桂在干旱高温胁迫下能快速启动体内抗氧化防御系统, 清除体内过量的ROS, 增加机体还原力, 以减缓胁迫带来的伤害。     

干旱胁迫下转基因甘薯块根膨大期水分利用效率和生理代谢特征

以转Cu/Zn-SOD和APX基因及其非转基因甘薯进行盆栽试验,在甘薯块根膨大期进行正常供水(田间最大持水量的80%)、中度缺水(田间最大持水量的60%)和重度缺水(田间最大持水量的40%)3种水分处理,分别测定转基因植株和对照植株在薯块膨大期的第20天和第70天的抗氧化酶系统、可溶性糖含量、光合系统之间的差异,以及在不同水分胁迫处理下产量和水分利用效率之间的差异。以此研究外源基因的超表达是否可以提高甘薯的产量及水分利用效率。结果显示:(1)转基因甘薯(TS)的SOD、APX活性以及可溶性糖含量均高于非转基因对照株(NT),但POD活性低于NT;TS和NT植株的APX活性、可溶性糖含量、净光合速率以及蒸腾速率均随干旱胁迫加重呈递减趋势。(2)干旱胁迫70d时,TS和NT植株光合参数均较胁迫20d时降低,且TS和NT间的净光合速率没有明显差异。(3)TS和NT两株系的块根产量在中度胁迫下最高而在重度胁迫下最低,而TS具有较高的块根产量且在重度胁迫下产量降低幅度较小。(4)TS的气孔导度和蒸腾速率显著低于NT,且TS的水分利用效率较NT更高。研究表明,Cu/Zn-SOD和APX基因可以显著增加干旱胁迫下甘薯块根膨大期的SOD、APX活性和可溶性糖含量,提高其水分利用效率,从而减轻干旱胁迫对产量的影响。