Expression and signal regulation of the alternative oxidase genes under abiotic stresses

Plants in their natural environment frequently face various abiotic stresses, such as drought, high salinity, and chilling. Plant mitochondria contain an alternative oxidase （AOX）, which is encoded by a small family of nuclear genes. AOX genes have been shown to be highly responsive to abiotic stresses. Using transgenic plants with varying levels of AOX expression, it has been confirmed that AOX genes are im- portant for abiotic stress tolerance. Although the roles of AOX under abiotic stresses have been extensively studied and there are several excellent reviews on this topic, the differential expression patterns of the AOX gene family members and the signal regulation of AOX gene（s） under abiotic stresses have not been extensively summarized. Here, we review and discuss the current progress of these two important issues.     

菜用大豆CIPK基因对逆境胁迫及激素的响应特征

CIPK(calcineurin B-like-interacting protein kinase)是一类丝氨酸/苏氨酸蛋白激酶,在植物响应逆境胁迫和激素信号转导中发挥重要作用。本研究利用大豆基因组数据库,在全基因组水平鉴定获得52个CIPK蛋白激酶。蛋白比对分析发现所有Gm CIPK含有高度保守特征性的N端激酶区、连接区和C端调控区。系统进化树分析发现大豆Gm CIPK与拟南芥、水稻CIPK分类一致,分为4个亚家族,且每个亚家族含有3个不同物种的成员,表明Gm CIPK基因的分化早于物种的分化。启动子分析表明,多数Gm CIPK基因的启动子区含有逆境和激素应答元件。组织表达分析发现,Gm CIPK基因呈现多样化的组织表达特性。进一步选取组织表达量相对较高的14个Gm CIPK进行荧光定量PCR分析,结果表明这些菜用大豆CIPK基因在不同程度上均受高温、干旱、高盐胁迫以及ABA、ACC、SA、Me JA激素的诱导表达。采用蛋白同源比对和蛋白互作在线数据库对拟南芥及大豆同源CIPK蛋白激酶与其他蛋白的互作关系进行了预测分析,发现17对同源CIPK与其他蛋白(激酶、磷酸酶、转录因子等)存在互作。本研究为菜用大豆CIPK基因的功能研究与利用奠定了基础。     

Zinc-finger protein GmZF351 improves both salt and drought stress tolerance in soybean

Abiotic stress is one of the most important factors reducing soybean yield. It is essential to identify regulatory factors contributing to stress responses.A previous study found that the tandem CCCH zincfinger protein Gm ZF351 is an oil level regulator. In this study, we discovered that the Gm ZF351 gene is induced by stress and that the overexpression of Gm ZF351 confers stress tolerance to transgenic soybean. Gm ZF351 directly regulates the expression of Gm CIPK9 and Gm SnRK, leading to stoma...     

大豆出苗期和苗期对盐胁迫的响应及耐盐指标评价

比较了4个大豆品种出苗期和苗期的耐盐性,测定150 mmol/L NaCl胁迫下的株高、下胚轴长、侧根数、地上干/鲜重、根干/鲜重、MDA含量、SOD活性、游离Pro含量,并将幼苗移栽到田间生长至成熟。结果表明:出苗期和苗期盐胁迫下4个品种的株高都显著降低、地上干/鲜重和根干/鲜重降低;出苗期胁迫侧根数减少,下胚轴长降低;而苗期胁迫侧根数增加,下胚轴长升高。未胁迫条件下,出苗期和苗期耐盐性强的品种22021-1的MDA含量和SOD活性高于耐盐性弱的品种22293-1。胁迫后,22021-1的MDA含量降低、SOD活性升高,其MDA含量分别比对照低51.03%和21.45%,SOD活性比对照高5.85%和45.77%;22293-1的MDA含量出苗期比对照高58.97%,苗期基本无变化,SOD活性出苗期和苗期升高都不显著。MDA和SOD可以作为大豆耐盐性筛选指标。早期的短时胁迫对不同耐盐性大豆品种的经济产量影响不同。     

基因芯片研究植物逆境基因表达新进展

逆境胁迫影响植物的正常生长, 导致作物减产, 甚至绝收。提高作物的抗逆性一直是作物遗传育种学家追求的目标, 大量研究也正试图揭示这一复杂的生物学机制。传统的从生理生化水平到单一基因的研究都难以揭示植物复杂的抗逆机制, 而基因芯片(Gene chip)的应用使得这一目标成为了可能, 基因芯片从整个转录水平入手, 能够揭示大量基因的表达和调控情况, 同时结合蛋白质组学和代谢组学的研究方法, 将基因定位于代谢途径的某个位置, 寻找逆境胁迫响应的关键基因, 完善植物逆境胁迫响应的分子网络, 为今后利用生物技术手段提高作物抗逆境胁迫能力提供依据。文章主要对近年来基因芯片在植物逆境胁迫基因表达研究中的进展进行了综述。     

Differential protection of photosynthetic capacity in trehalose-and lea protein-producing transgenic plants under abiotic stresses

We previously demonstrated that both trehalose and LEA protein protect plants from damage by drought, salt, and heat. Here, we compared their effectiveness in preserving photosynthetic capacity under those abiotic stresses. Upon dehydration, the Pmax (maximal photosynthetic rate) of O₂ evolution decreased similarly in both nontransformants andotsA plants. Contrastingly, Pmax was maintained at a considerably higher level inCaLEA6 plants. However, no significant differences in Chl fluorescence parameters were observed between transformants and nontransformants. Under salinity stress,CaLEA6 plants were also better thanotsA plants in terms of their values for Pmax, photochemical efficiency of PSII(Fv/Fm), and photochemical quenching (qP). After heat bothotsA andCaLEA6 plants maintained a higher Pmax as well as more favorable Chl fluorescence parameters, although the latter transformant performed slightly better overall. Therefore, despite the comparable effectiveness of trehalose and LEA protein in enhancing tolerance against those abiotic stresses, they confer differential protection in maintaining photosynthetic capacity. Compared with trehalose, the CaLEA6 protein appears to be a more universal and effective agent under those stresses.     

大豆GolS基因家族鉴定及盐旱胁迫下的表达分析

肌醇半乳糖苷合成酶（galactinol synthase,GolS）是棉子糖家族寡糖（raffinose family oligosaccharides,RFOs）生物合成途径中的关键酶,在植物对非生物胁迫的反应中发挥重要作用。然而,关于大豆（Glycine max）GolS基因家族成员的分子结构特征还未见研究报道。本研究在全基因组水平上鉴定了6个大豆GolS基因家族成员,并对其理化性质、染色体定位、进化关系、基因结构、保守基序、二级结构、三级结构、组织特异性表达模式以及盐和干旱胁迫下的表达量进行了分析。结果表明：6个大豆GolS基因不均匀地分布在4条染色体上,6个大豆GolS蛋白的等电点为5.45-6.08,分子量变化范围为37 567.07-38 817.59 Da,氨基酸数量为324-339 aa;亚细胞定位预测结果发现4个蛋白定位在叶绿体上,2个蛋白定位在细胞质。系统进化树分析表明,大豆GolS基因家族成员在进化树中呈现出两两紧邻的现象,在进化上较为保守。6个基因成员含有的外显子数目为3或4。二级结构和三级结构预测表明,该家族所有成员蛋白质的空间结构主要由α螺旋和无规则卷曲结构组成,有较少的β转角结构和延伸链结构。组织特异性表达分析表明,6个GmGolS家族成员在种子、根、根毛、花、茎、豆荚、根瘤和叶中均有不同程度表达。基于qRT-PCR的表达分析显示,盐旱处理后所有GmGolS基因成员表现出不同程度的上调表达,表明这些基因可能与植物的耐盐抗旱响应有关。本研究结果为后续开展大豆GolS基因的功能解析奠定了基础。     

Expression of soybean plant hemoglobin gene family under abiotic stresses

Many abiotic stresses induce the generation of nitric oxide (NO) in plant tissues, where it functions as a signal molecule in stress responses. Plants modulate NO by oxidizing it to NO₃⁻ with plant hemoglobin (GLB), because excess NO is toxic to cells. At least eight genes encoding GLB have been identified in soybean, in three clades: GLB1, GLB2, and GLB3. However, it is still unclear which GLB genes are responsible for NO regulation under abiotic stress in soybean. We exposed soybean roots to flooding, salt, and two NO donors—sodium pentacyanonitrosylferrate (III) dihydrate (SNP) and S-nitroso-N-acetyl-d,l-penicillamine (SNAP)—and analyzed expression of GLB genes. GmGLB1, one of two GLB1 genes of soybean, significantly responded to both SNP and SNAP, and its induction was almost completely repressed by a NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide. GmGLB1 responded to flooding but not to salt, suggesting that it is responsible for NO regulation under NO-inducing abiotic stresses such as flooding. GmGLB3, one of two GLB3 genes of soybean, did not respond to NO donors at all but did respond to flooding, at a lower level than GmGLB1. These results suggest that flooding induces not only NO but also unknown factor(s) that induce GmGLB3 gene in soybean.     

Role of mucilage during achene germination and sprout growth of the endangered Tibetan medicinal herb Mirabilis himalaica (Nyctaginaceae) exposed to abiotic stresses

Aim Mirabilis himalaica (Nyctaginaceae) is an endangered medicinal plant mainly distributed in the plateau region of northern Tibet, China. The outer surface of M. himalaica achenes is covered by a pectinaceous mucilaginous layer upon hydration. However, the role of the achene mucilage is poorly understood. In this study, we investigated the effects of mucilage on achene germination and sprout growth under abiotic stress to explain how M. himalaica survive the alpine environment.Methods We investigated the effect of mucilage on achenes germination by contrast the capacity of water absorption, dehydration and respiration of intact achene and the achene with mucilage removal. We performed abiotic stresses experiments including drought stress, salt stress, cold stress and high temperature stress, and quantified the effects of mucilage removal on achene germination rate, root and shoot lengths of seedlings.Important findings Mucilage is extremely hydrophilic, and the mass of intact achenes can be 9-fold greater than that of demucilaged achenes. The removal of the mucilaginous layer did not significantly change final germination percentages under ideal conditions, but intact achenes (i.e. with mucilage) took longer to germinate. The mucilage significantly decreased seed respiration rates by acting as a physical barrier that prevented oxygen diffusion. Germination rates, shoot and root growth of intact achenes were higher than those of demucilaged ones during exposures to cold, heat, osmotic and salt stresses. Achene mucilage presumably plays an ecologically important role in the life cycle of M. himalaica by aiding the critical achene germination and early seedling growth in the stressful habitats of the plateau region of northern Tibet.     

Role of abscisic acid in regulation of wheat seedling growth under salinity stress

Growth of wheat seedlings (Triticum aestivum L. cv. Mehran-89), in hydroponic culture, was affected by abscisic acid (ABA). Using salinity stress and exogenous ABA application (10-6 M) to enhance endogenous ABA level, the growth of roots was more suppressed than the growth of shoots. On the other hand, norflurazon, which inhibits ABA biosynthesis, reduced only the growth of shoots. This revised version was published online in July 2006 with corrections to the Cover Date.     

The involvement of hydrogen peroxide in plant responses to stresses

The role of reactive oxygen species, especially H₂O₂, in plant response to stresses has been the focus of much attention. Hydrogen peroxide has been postulated to play multiple functions in plant defence against pathogens. (1) H₂O₂ may possess direct microbicidal activity at the sites of pathogen invasion. (2) It is used for cell-wall reinforcing processes: lignification and oxidative cross-linking of hydroxyproline-rich proteins and other cell-wall polymers. (3) It was found to be necessary for phytoalexin synthesis. (4) H₂O₂ may trigger programmed plant cell death during the hypersensitive response that restricts the spread of infection. (5) H₂O₂ has been suggested to act as a signal in the induction of systemic acquired resistance and (6) it induces defence genes. Recently H₂O₂ has been proposed to be involved in the signal transduction pathways leading to acclimation and protection from abiotic stresses. The present review discusses new insights into the function of H₂O₂ in plant responses to biotic and abiotic stresses.     

盐胁迫对大豆根系木质部压力和Na+吸收的影响

取栽培大豆的水培幼苗为材料,用木质部压力探针和原子吸收分光光度计测定了盐胁迫条件下其根木质部压力和伤流液中Na~+含量的变化,以分析大豆抗盐吸水的机制.结果表明:在25～150 mmol/L NaCl的浓度范围内,随着盐胁迫强度的增加,大豆根木质部负压力的绝对值逐渐增大,但相对负压力和根的径向反射系数则逐渐减小;木质部伤流液中Na~+含量逐渐增加,但Na~+的相对含量则逐渐降低.同时,虽然根系吸水所需的木质部负压力(压力势)及根木质部伤流液的渗透势随着盐胁迫强度的增加都有所下降,但两者共同作用使木质部水势下降的幅度远远小于根外溶液水势(渗透势)下降的幅度,即随着根外溶液盐浓度的升高,根木质部溶液的总水势逐渐高出根外溶液的水势.上述结果说明,在盐胁迫下大豆可以利用相对小的木质部负压力逆水势梯度吸水,且通过避免对Na~+的过量吸收来适应盐胁迫环境.     

Small RNAs from MITE-derived stem-loop precursors regulate abscisic acid signaling and abiotic stress responses in rice

Small silencing RNAs (sRNAs) are non-coding RNA regulators that negatively regulate gene expression by guiding mRNA degradation, translation repression or chromatin modification. Plant sRNAs play crucial roles in various developmental processes, hormone signaling, immune responses and adaptation to a variety of abiotic stresses. miR441 and miR446 were previously annotated as microRNAs (miRNAs) because their precursors can form typical stem-loop structures, but are not considered as real miRNAs because of inconsistency with some ancillary criteria of the recent guidelines for annotation of plant miRNAs. We tentatively rename them small interfering (si)R441 and siR446, respectively, in this study. It has recently been shown that the precursors of siR441 and siR446 might originate from the miniature inverted-repeat transposable element (MITE) Stowaway1. In this report, we show that, in contrast with Dicer-like (DCL)3- and RNA-dependent RNA polymerase (RDR)2-dependent MITE-derived ra-siRNAs, siR441 and siR446 are processed by OsDCL3a but independent of OsRDR2, indicating that siR441 and siR446 are generated from single-stranded stem-loop precursors. We also show that abscisic acid (ABA) and abiotic stresses downregulate the expression of siR441 and siR446 but, surprisingly, increase the accumulation of their precursors in rice plants, implying that processing of siRNA precursors is inhibited. We provide evidence to show that this defective processing is due to increased precursor accumulation per se, possibly by intermolecular self-pairing of the processing intermediate sequences, thus hindering their normal processing. Functional examinations indicate that siR441 and siR446 are positive regulators of rice ABA signaling and tolerance to abiotic stress, possibly by regulating MAIF1 expression.     

Integration of abscisic acid signalling into plant responses

The phytohormone abscisic acid (ABA) plays a major role as an endogenous messenger in the regulation of plant's water status. ABA is generated as a signal during a plant's life cycle to control seed germination and further developmental processes and in response to abiotic stress imposed by salt, cold, drought, and wounding. The action of ABA can target specifically guard cells for induction of stomatal closure but may also signal systemically for adjustment towards severe water shortage. At the molecular level, the responses are primarily mediated by regulation of ion channels and by changes in gene expression. In the last years, the molecular complexity of ABA signal transduction surfaced more and more. Many proteins and a plethora of "secondary" messengers that regulate or modulate ABA-responses have been identified by analysis of mutants including gene knock-out plants and by applying RNA interference technology together with protein interaction analysis. The complexity possibly reflects intensive cross-talk with other signal pathways and the role of ABA to be part of and to integrate several responses. Despite the missing unifying concept, it is becoming clear that ABA action enforces a sophisticated regulation at all levels.     

大豆异黄酮浸种对盐胁迫大豆幼苗的生理效应

大豆异黄酮( Soybean isoflavones)是在大豆生长过程中形成并在成熟种子和叶片中积累较多的一类具有生物活性的次生代谢物,通常可作为人们日常生活中的一类营养保健品.研究了外源大豆苷或染料木苷溶液(0.01 mg/L)浸种处理对盐胁迫栽培大豆(N23674品种)和滩涂野大豆(BB52种群)及其经逐代耐盐性筛选的杂交后代(4076株系,F5)幼苗叶片伤害率、光合作用、Na+含量和Na+/K+值、活性氧清除酶活性及内源大豆异黄酮含量等生理指标的影响.结果表明:盐胁迫下,两种外源大豆异黄酮浸种处理均可显著抑制叶片相对电解质渗透率和硫代巴比妥酸反应物(TBARS)含量的上升及净光合速率(Pn)的下降,降低Na+含量和Na+/K+值,增强超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)活性,提高内源大豆异黄酮含量,从而表现对盐害的缓解效应,其中对耐盐性较弱的栽培大豆N23674品种效应更明显.这为大豆异黄酮在大豆耐盐育种、化学调控和盐碱地种植利用等提供了理论依据.     

Molecular titration and ultrasensitivity in regulatory networks

Protein sequestration occurs when an active protein is sequestered by a repressor into an inactive complex. Using mathematical and computational modeling, we show how this regulatory mechanism (called “molecular titration”) can generate ultrasensitive or “all-or-none” responses that are equivalent to highly cooperative processes. The ultrasensitive nature of the input-output response is mainly determined by two parameters: the dimer dissociation constant and the repressor concentration. Because in vivo concentrations are tunable through a variety of mechanisms, molecular titration represents a flexible mechanism for generating ultrasensitivity. Using physiological parameters, we report how details of in vivo protein degradation affect the strength of the ultrasensitivity at steady state. Given that developmental systems often transduce signals into cell-fate decisions on timescales incompatible with steady state, we further examine whether molecular titration can produce ultrasensitive responses within physiologically relevant time intervals. Using Drosophila somatic sex determination as a developmental paradigm, we demonstrate that molecular titration can generate ultrasensitivity on timescales compatible with most cell-fate decisions. Gene duplication followed by loss-of-function mutations can create dominant negatives that titrate and compete with the original protein. Dominant negatives are abundant in gene regulatory circuits, and our results suggest that molecular titration might be generating an ultrasensitive response in these networks.     

Germination variation in Arabidopsis thaliana accessions under moderate osmotic and salt stresses

Background

Water and salt stresses are two important environmental factors that limit the germination of seeds in most ecological environments. Most studies conducted so far to address the genetic basis of the above phenomenon have used stress conditions that are much more extreme than those found in natural environments. Furthermore, although an excess of ions and water restrictions have similar osmotic effects on germination, the common and divergent signalling components mediating the effects of both factors remain unknown.

Methods

The germination of seeds was compared under solutions of NaCl (50 mm) and polyethylene glycol (PEG, −0·6 MPa), that establish mild stress conditions, in 28 Arabidopsis thaliana accessions. Because Bayreuth (Bay) and Shadara (Sha) accessions showed contrasting sensitivity responses to both stresses, a quantitative trait locus (QTL) analysis was carried out using Bay × Sha recombinant inbred lines (RILs) to identify loci involved in the control of germination under mild salt and osmotic stresses.

Key Results

Two loci associated with the salt sensitivity response, named SSR1 and SSR2 QTLs, and four loci for the osmotic sensitivity response, named OSR1–OSR4 QTLs, were mapped. The effects of the SSR1 QTL on toxic salt sensitivity, and the osmotic contribution of OSR1, were confirmed by heterogeneous inbred families (HIFs). Whilst the SSR1 QTL had a significant effect under a wide range of NaCl concentrations, the OSR1 QTL was confirmed only under moderate drought stress. Interestingly the OSR1 QTL also showed pleiotropic effects on biomass accumulation in response to water deficit.

Conclusions

The regulation of germination under moderate salt and osmotic stresses involves the action of independent major loci, revealing the existence of loci specifically associated with the toxic component of salt and not just its osmotic effect. Furthermore, this work demonstrates that novel loci control germination under osmotic stress conditions simulating more realistic ecological environments as found by populations of seeds in nature.     

Effect of abscisic acid application on root isoflavonoid concentration and nodulation of wild-type and nodulation-mutant soybean plants

Isoflavonoids (daidzein, genistein, and coumestrol) are involved in induction of nod genes in Bradyrhizobium japonicum and may be involved in nodule development as well. Abscisic acid (ABA) may also impact nodulation since ABA is reportedly involved in isoflavonoid synthesis. The current study was conducted to evaluate whether ABA plays a role in differential nodulation of a hypernodulated soybean (Glycine max L. Merr.) mutant and the Williams parent. Exogenous ABA application resulted in a decrease in nodule number and weight in both lines. Isoflavonoid concentrations were also markedly decreased in response to ABA application in both inoculated and noninoculated soybean roots. The inoculation treatment itself resulted in a marked increase in isoflavonoid concentrations of NOD1-3, regardless of ABA levels, while only slight increases occurred in Williams. The nodule numbers of both soybean lines across several ABA concentration treatments were highly correlated with the concentration of all three isoflavonoids. However, differences in internal levels of ABA between lines were not detected when grown in the absence of external ABA additions. It is concluded that differential nodule expression between the wild type and the hypernodulated mutant is not likely due to differential ABA synthesis.