复苏植物旋蒴苣苔早期光诱导蛋白的特征分析

随着全球气候变暖,旱胁迫越来越成为粮食生产的严重威胁。探索复苏植物耐受极端旱胁迫的机理,积累抗旱相关基因资源,对于植物适应逆境的基础研究,和农业可持续发展,均有重要的作用。旋蒴苣苔(B. hygrometrica)广泛分布于我国山区,是复苏植物的代表性物种,而且有高质量的全基因组序列可以分析利用。本研究对旋蒴苣苔(B. hygrometrica)基因组中的早期光诱导蛋白(ELIPs)基因家族进行挖掘和分析。旋蒴苣苔(B. hygrometrica)基因组中早期光诱导蛋白(ELIPs)基因的数目远高于其他植物(拟南芥,水稻,辣椒,番茄)。系统发育分析显示,早期光诱导蛋白(ELIPs)基因序列具有很高的植物特异性。这说明,早期光诱导蛋白(ELIPs)基因对不同植物进化和适应各自的生长环境具有重要的作用。对处于不同程度干旱状态的旋蒴苣苔(B. hygrometrica)叶片的基因表达分析,表明一部分早期光诱导蛋白(ELIPs)基因表现出很强的对旱处理的响应。不同的早期光诱导蛋白(ELIPs)基因表现出三种不同的响应模式。这些结果表明,旋蒴苣苔(B. hygrometrica)早期光诱导蛋白(ELIPs)基因参与植株应对旱胁迫,而且有精细的调控机制控制不同早期光诱导蛋白(ELIPs)基因的表达量。     

复苏植物旋蒴苣苔棉子糖合酶基因的克隆和表达

复苏植物是研究植物耐脱水机制的特殊模式植物和宝贵的耐旱基因资源植物。以复苏植物旋蒴苣苔(Boea hygrometrica) 为材料研究其在脱水和复水过程中棉子糖系列寡糖含量的变化, 并克隆了旋蒴苣苔棉子糖合酶基因BhRFS。荧光定量PCR检测表明, BhRFS受干旱、低温(4°C)、高盐(200 mmol·L^–1NaCl)和ABA(100 μmol·L^–1)诱导表达上调, 而高温(37°C)抑制其表达, H₂O₂(200 μmol·L^–1)处理对其没有影响。研究结果表明, BhRFS可能参与了多种非生物逆境胁迫抗性反应, 并受到ABA依赖的信号通路调控。     

复苏植物旋蒴苣苔C2结构域小蛋白BhC2DP1参与植物对ABA的反应

为揭示植物抗旱的调控机理, 对复苏植物旋蒴苣苔(Boea hygrometrica)的一个编码C2结构域小蛋白的基因BhC2DP1进行研究。Real-time PCR和Pro_BhC2DP1:GUS报告基因检测显示, 该基因只在干旱早期和外源Ca²⁺处理0.5小时时受诱导表达; 分别施加Ca²⁺螯合剂EGTA和逆境激素ABA均抑制该基因表达, 但二者同时处理则显著诱导其表达, 表明ABA对该基因转录水平的调控是Ca²⁺依赖型的。过表达BhC2DP1的拟南芥(Arabidopsis thaliana)对ABA的敏感性增强, EGTA处理可消除其与野生型的差异, 表明Ca²⁺是BhC2DP1蛋白参与ABA反应所必需的。综上所述, ABA和Ca²⁺信号途径的精细调控可能是决定干旱诱导旋蒴苣苔中BhC2DP1基因表达时间、丰度和功能的重要机制。     

马铃薯甲虫热激蛋白基因Ld-HSP60的克隆、序列分析及温度胁迫下的表达

【目的】马铃薯甲虫Leptinotarsa decemlineata是一种世界性检疫害虫,对温度胁迫具有极强的适应性,为进一步明确其对温度胁迫适应性的分子机制,研究了热激蛋白HSP60在马铃薯甲虫温度胁迫应答过程中的作用。【方法】采用RT-PCR及RACE技术克隆马铃薯甲虫热激蛋白HSP60基因的cDNA全长序列;利用生物信息学软件分析该基因及其编码蛋白质的序列特性;运用实时荧光定量PCR技术分析该基因在温度胁迫下的表达模式。【结果】克隆得到马铃薯甲虫热激蛋白HSP60基因,命名为Ld-HSP60(Gen Bank登录号:KC556801),其cDNA全长2 234 bp,开放阅读框(ORF)长1 731 bp,编码576个氨基酸,相对分子量约为61.27 kD,理论等电点为5.51,5'端非翻译区(UTR)长101 bp,3'UTR长402 bp。氨基酸序列中含有HSP60家族典型的特征序列。实时荧光定量PCR结果表明,低温胁迫(-10和0℃)下未检测到马铃薯甲虫雌雄成虫中Ld-HSP60的诱导表达;高温胁迫(38和44℃)诱导马铃薯甲虫雄成虫Ld-HSP60上调表达,随着胁迫温度的升高LdHSP60表达量呈现先升高后降低的趋势,38℃高温胁迫下表达量最高,胁迫时间越长Ld-HSP60表达量也越高。【结论】相比其他热激蛋白,HSP60对温度敏感性较低,推测HSP60可能在马铃薯甲虫雄成虫抵御高温胁迫中发挥作用。     

拟南芥受Cd2+诱导表达基因的筛选及其在Cd2+胁迫下的功能

镉是一种毒性很大的重金属。土壤溶液中即使存在极低浓度Cd2+也能对植物造成伤害。早在植物做出结构和代谢的调整以适应逆境之前,由于Cd2+的刺激,植物的基因表达已经发生了变化。这里我们采用一种新的基于引物退火控制技术的差异显示方法来筛选受镉离子诱导表达的基因。获得的19条差异条带代表着18个基因。经过RT-PCR方法验证,其中6个基因确实是受Cd2+诱导表达,包括LEA(胚胎发育晚期丰富蛋白), AtGSTF2(谷胱甘肽-S-转移酶2), AtGSTF6(谷胱甘肽-S-转移酶6), HSP70(热激蛋白70), sHSP17.6B-CI(17.6 kDa 类型 I小分子热激蛋白)和sHSP17.6-CII(17.6 kDa类型II 小分子热激蛋白)。 这些结果有助于研究植物对镉离子胁迫的解毒机制。其中的三个热激蛋白基因的启动子也能考虑用于植物修复。     

一个水稻多逆境响应基因OsMsr3克隆与表达分析

为深入探讨水稻对逆境的反应机理并寻找新的植物耐逆基因,采用Affymetrix水稻表达芯片(含51279个转录本)分析了培矮64S全基因组在不同逆境(高温、干旱、低温)胁迫下、不同生育时期叶片和穗中的表达谱,从中筛选出一个受多种逆境诱导表达的基因OsMsr3(Oryza sativa L.multiple stresses responsive gene3,GenBank登陆号为 FJ383169).定量实时PCR分析结果进一步证实了此基因在逆境条件下的诱导表达模式.用 RT-PCR方法扩增获得了包含其完整开放阅读框的cDNA克隆,序列分析表明,其ORF大小为480 bp,编码一个具有160个氨基酸残基的低分子量热激蛋白,推测分子量约为18.0 kD;pI约为6.8.对其编码的蛋白质进行分析,发现其羧基端存在一个HSP20的蛋白保守结构域,与其他植物中的低分子量热激蛋白的相似性介于33.7%～97.5%.对其可能的启动子序列分析,发现6类与逆境反应有关的顺式作用元件.推测该基因在逆境反应中起着重要的作用,进一步的研究正在进行中.     

橡胶树胶乳高表达热激蛋白HbHSP90.4基因抗逆功能分析

为了分析热激蛋白90（HSP90）基因在橡胶树（Hevea brasiliensis）逆境胁迫和激素转导中的作用,利用PCR技术从橡胶树品种热研73397胶乳中克隆得到HbHSP90.4基因全长cDNA序列,该基因含有1个2 451 bp开放阅读框（ORF）,编码816个氨基酸。生物信息学分析结果表明,HbHSP90.4含有HSP90 superfamily和HATPase superfamily结构域,属于HSP90家族成员。系统进化分析发现该蛋白与木薯MeHSP90具有较近的亲缘关系。亚细胞定位预测显示HbHSP90.4基因定位在内质网。qRT-PCR结果表明HbHSP90.4基因主要在橡胶树胶乳中表达。干旱、冷胁迫、橡胶树白粉菌侵染、H₂O₂和MeJA处理均可促进胶乳HbHSP90.4基因上调表达,而其在ETH、SA和ABA处理中均呈现显著下调表达。构建植物表达载体HbHSP90.4-mScarlet,为进一步的转基因植物的做成准备了材料。本研究为阐明胶乳HbHSP90.4基因响应橡胶树逆境胁迫过程和植物激素信号传导途径分子调控机制奠定坚实基础。     

热激转录因子在植物抗非生物胁迫中的功能研究进展

植物在遭受环境胁迫时会产生一系列应激反应,而热激转录因子可通过介导热激蛋白或其他热诱导基因的转录和表达,来参与调控植物抵抗逆境胁迫过程和其他生命活动。主要介绍了植物热激转录因子的基本蛋白结构域,阐述了3类热激转录因子在抗极端温度（高温、低温）胁迫、干旱胁迫、高盐胁迫、活性氧胁迫中的功能与作用机制,并探讨和展望了植物热激转录因子在植物育种和提高植物抗逆性的研究中的发展与应用前景,以期为深入研究热激转录因子在调控植物抵抗逆境胁迫中的生物学功能与机制提供理论参考。     

中国南瓜CmHSP90基因的克隆及其在逆境胁迫下的表达分析

热激蛋白90(heat shock protein 90, HSP90)广泛介导胁迫信号的传递,在植物逆境应答反应中起重要作用。为探究HSP90基因的功能,该研究采用RT-PCR方法从中国南瓜(Cucurbitamoschata)幼苗中分离到2个HSP90基因的开放阅读框(ORF)全长,分别命名为CmHSP90-5和CmHSP90-6。二者的序列长度分别为2 357和2 472 bp,编码793和824个氨基酸。蛋白序列分析显示, 2个CmHSP90蛋白均属于亲水性蛋白,含信号肽,无跨膜区,结构上都含有典型的ATPase保守结构域,亚细胞分别定位于叶绿体和线粒体。同源比对和进化分析发现, 2个CmHSP90蛋白与苦瓜(Momordica charantia)、甜瓜(Cucumis melo)和黄瓜(Cucumis sativus)HSP90蛋白的相似性最高且遗传距离最近。qRT-PCR分析显示, 2个CmHSP90基因对高温、低温、ABA、高盐、H2O2、干旱和水杨酸(SA)等多种胁迫均具有明显的应答反应,但响应的速度和强度并不相同,其中对高温和水杨酸处理均表现出短时间强烈响应,推测CmHSP90基因可能在中国南瓜热胁迫和水杨酸信号途径中发挥调节作用。该研究结果为深入了解HSP90基因功能以及阐明中国南瓜的抗逆机制提供理论依据。     

植物热激蛋白70

植物热激蛋白70(HsP70)由多基因家族编码.除热胁迫外,其它环境因素如低温、干旱等也能诱导HSP70基因的大量表达.HSP70主要参与新生肽的成熟与分拣、变性蛋白的复性或降解等细胞活动.该文介绍HSP70的结构、功能和调控的研究现状.     

The oxidative stress response in Enterococcus faecalis: relationship between H2O2 tolerance and H2O2 stress proteins

The hydrogen peroxide (H₂O₂) stress response in Enterococcus faecalis ATCC19433 was investigated. A 2·4 mmol l⁻¹ H₂O₂ pretreatment conferred protection against a lethal concentration (45 mmol l⁻¹) of this agent. The relatively high concentrations of H₂O₂ used for adaptation and challenge treatments in Ent. faecalis emphasised the strong resistance towards oxidative stress in this species. Various stresses (NaCl, heat, ethanol, acidity and alkalinity) induced weak or strong H₂O₂ cross-protection. This paper describes the involvement of protein synthesis in the active response to lethal dose of H₂O₂, in addition to the impressive enhancement of synthesis of five H₂O₂ stress proteins. Combined results suggest that these proteins might play an important role in the H₂O₂ tolerance response.     

Characterization of Lactobacillus collinoides response to heat, acid and ethanol treatments

Tolerance to stress and cross-protection in Lactobacillus collinoides were examined after exposure to ethanol, acid or heat shock. Ethanol and heat-adapted cells demonstrate induced homologous␣tolerance and cross-resistance to acid stress. No cross-protection of acid-adapted cells against ethanol and heat stresses was observed. Heat was the only pretreatment leading to cross-protection against the two other stresses. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins; the synthesis of some of these polypeptides being induced by more than one condition. The greatest overlap was observed between ethanol and heat treatments. Ten proteins were found to be common to these stresses. Received: 7 November 1998 / Received revision: 10 February 1999 / Accepted: 12 February 1999     

盐碱胁迫对平欧杂种榛枝条电阻抗图谱参数及离子含量的影响

为探讨平欧杂种榛枝条对盐碱胁迫的适应性,测定‘辽榛3号’当年生枝条在中性盐(NaCl)、碱性盐(Na₂CO₃)及混合盐3种盐碱胁迫下各项电阻抗图谱(EIS)参数的变化及主要矿质元素含量,分析各电阻抗参数及矿质离子含量的相关性.结果表明： 随着NaCl浓度的增加,电阻抗参数高频电阻率(r)、低频电阻率(r₁)、胞内电阻率(r_i)和胞外电阻率(r_e)均表现出先降低后增加再降低的变化趋势;Na₂CO₃胁迫下,各参数则呈现持续上升的变化规律;混合盐碱胁迫下,r₁、r_e随盐浓度增加持续降低,Na⁺含量在3种类型盐碱胁迫下均随浓度的增加而上升,且NaCl胁迫>混合盐碱胁迫> Na₂CO₃胁迫;Na₂CO₃胁迫导致了Zn、B、Ca 3种元素含量的降低.5种阳离子总量与r₁、r_e、弛豫时间(τ)、弛豫时间分布系数(Ψ)呈显著负相关.平欧杂种榛枝条能耐200 mmol·L^-1碱性盐胁迫,而对中性盐的忍耐能力介于100～150 mmol·L^-1.     

Hexavalent chromium,a lung carcinogen,confers resistance to thermal stress and interferes with heat shock protein expression in human bronchial epithelial cells

Exposure to hexavalent chromium [Cr(VI)], a lung carcinogen, triggers several types of cellular stresses, namely oxidative, genotoxic and proteotoxic stresses. Given the evolutionary character of carcinogenesis, it is tempting to speculate that cells that survive the stresses produced by this carcinogen become more resistant to subsequent stresses, namely those encountered during neoplastic transformation. To test this hypothesis, we determined whether pre-incubation with Cr(VI) increased the resistance of human bronchial epithelial cells (BEAS-2B cells) to the antiproliferative action of acute thermal shock, used here as a model for stress. In line with the proposed hypothesis, it was observed that, at mildly cytotoxic concentrations, Cr(VI) attenuated the antiproliferative effects of both cold and heat shock. Mechanistically, Cr(VI) interfered with the expression of two components of the stress response pathway: heat shock proteins Hsp72 and Hsp90α. Specifically, Cr(VI) significantly depleted the mRNA levels of the former and the protein levels of the latter. Significantly, these two proteins are members of heat shock protein (Hsp) families (Hsp70 and Hsp90, respectively) that have been implicated in carcinogenesis. Thus, our results confirm and extend previous studies showing the capacity of Cr(VI) to interfere with the expression of stress response components.     

黄瓜幼苗光合作用对高温胁迫的响应与适应

以‘津优35号’黄瓜幼苗为试材,研究高温(HT: 42 ℃/32 ℃)和亚高温(SHT: 35 ℃/25 ℃)胁迫对黄瓜幼苗光合作用及生长量的影响.结果表明: 高温、亚高温明显抑制幼苗生长.随着胁迫时间的延长,黄瓜幼苗叶片的光合速率(P_n)逐渐降低,胞间CO₂浓度(C_i)趋于升高,气孔导度(g_s)、蒸腾速率(T_r)、光呼吸速率(P_r)和暗呼吸速率(D_r)先上升后下降,高温、亚高温引起P_n降低的主要原因是非气孔限制.高温、亚高温可使黄瓜幼苗叶片的暗下光系统Ⅱ最大光化学效率(F_v/F_m)、光下实际光化学效率(Φ_PSII)、光化学猝灭系数(q_P)和电子传递效率(ETR)显著降低,初始荧光(F_o)和非化学猝灭系数(NPQ)逐渐升高.随着胁迫时间的延长,HT处理的RuBP羧化酶(RuBPCase)和Rubisco活化酶(RCA)活性及其mRNA表达量逐渐降低,而SHT处理的胁迫初期变化不大,3 d后趋于降低;HT和SHT处理的景天庚酮糖-1,7-二磷酸酶(SBPase)和果糖-1,6-二磷酸醛缩酶(FBA)活性与mRNA表达均呈先升高后降低趋势.可见,适宜光强下短时亚高温处理黄瓜幼苗不会产生明显光抑制,高温胁迫会对其PSⅡ反应中心造成严重损伤;光合酶受高温胁迫诱导,但其诱导效应与温度升高幅度和高温持续时间有关.     

Heat and salt stress in the food pathogen Bacillus cereus

AIMS: The effects of stresses imposed on bacterial contaminants during food processing and treatment of packaging material were evaluated on the food pathogen Bacillus cereus. METHODS AND RESULTS: Conditions were established which allowed the cells to adapt to heat, ethanol and hydrogen peroxide stresses, but not to osmotic shock. Cross protection between stresses indicated a clear hierarchy of resistance with salt protecting against hydrogen peroxide, which protected against ethanol, which protected against heat shock. The cultures were shown to be most sensitive to heat, ethanol and oxidative stress at mid-exponential phase and to become resistant at stationary phase. Adaptive levels of stressor were found to induce synthesis of general stress and stress-specific proteins and differential accumulation of proteins was demonstrated between heat- or salt-stressed and unstressed cells. CONCLUSIONS: Sequencing revealed that a number of glycolytic enzymes were regulated by heat and osmotic shocks and that the chaperone GroEL was induced by heat shock. SIGNIFICANCE AND IMPACT OF THE STUDY: The implications of the physiological data in designing storage and processing conditions for food are discussed. The identification of stress-regulated proteins reveals a clear role for glycolysis in adaptation to heat shock and osmotic stress.     

C. elegans STI-1, the Homolog of Sti1/Hop, Is Involved in Aging and Stress Response

Environmental and physiological stresses such as heat shock, oxidative stress, heavy metals, and pathogenic conditions induce cellular stress response. This response is often mediated by heat shock proteins that function as molecular chaperones. A stress-inducible cochaperone, Sti1/Hop (Hsp organizer protein), functions as an adaptor protein that simultaneously binds with Hsp70 and Hsp90 to transfer client proteins from Hsp70 to Hsp90. However, the biological role of STI-1 in vivo is poorly understood in metazoans. Here, we report the characterization of the Caenorhabditis elegans homolog of Sti1/Hop, which is approximately 56% identical with human STI-1. C. elegans STI-1 (CeSTI-1) is expressed in the pharynx, intestine, nervous system, and muscle from larvae to adults. Analysis of proteins immunoprecipitated with anti-STI-1 antibody by mass spectrometry revealed that CeSTI-1 can bind with both Hsp70 and Hsp90 homologs like its mammalian counterpart. sti-1 expression is elevated by heat stress, and an sti-1(jh125) null mutant shows decreased fertility under heat stress conditions. These mutants also show abnormally high lethality in extreme heat and may be functioning with DAF-16 in thermotolerance. In addition, sti-1(jh125) mutants have a shortened life span. Our results confirm that CeSTI-1 is a cochaperone protein that may maintain homeostatic functions during episodes of stress and can regulate longevity in nematodes.