AtsHsp17.6-CⅠ和AtsHsp17.6-CⅡ基因对非生物胁迫的应答研究

小分子热激蛋白是植物受到热胁迫后的主要表达产物之一,与植物细胞耐热有密切关系。该研究发现,拟南芥小分子热激蛋白基因AtsHsp17.6-CⅠ和AtsHsp17.6-CⅡ 除热激之外,重金属离子Ni⁺、Pb²⁺、Cu²⁺、Zn²⁺和Al³⁺均能诱导这2个热激蛋白基因的表达;氧化胁迫和渗透胁迫同样也能诱导它们表达。该研究将由CaMV35S启动子驱动的这2个小分子热激蛋白基因导入拟南芥,RT-PCR分析表明,2个小分子热激蛋白基因在转基因植物中呈现组成型表达。实验结果表明,组成型表达小分子热激蛋白基因AtsHsp17.6-CⅠ的转基因植物表现出对6 μmol·L^-1 Cd²⁺胁迫、0.4% NaCl胁迫的耐受性。研究表明,这2个小分子热激蛋白基因可能参与着多种抗逆途径,推测其能够减轻或抵抗逆境胁迫引起的伤害并对其进行修复。     

不结球白菜热激蛋白基因克隆及表达

从不结球白菜'暑绿'中克隆到一个受热激诱导的小分子量热激蛋白(sHSP)基因,命名为BcHSP(DDBJ登录号为AB367955),该基因核苷酸序列全长722 bp,编码157个氨基酸,与芜菁、芥蓝、拟南芥等有90%以上的相似性.实时定量检测表明,不结球白菜BcHSP转录表达受热激诱导,以叶片中表达量最高,BcHSP在不结球白菜叶片中表达特征说明它可能与植物叶片的耐热性关系更为密切.     

马铃薯HSP17.7基因的克隆及其对高温逆境的响应

热激蛋白和植物对高温的响应密切相关,同时在植物生长发育调控和逆境抵抗等方面具有重要作用。该研究克隆了马铃薯热激蛋白基因StHSP17.7(GenBank登录号为XP_006350804.1),对其全长cDNA序列进行了相关生物信息学分析,并利用qRT-PCR分析StHSP17.7基因在高温胁迫下的差异表达特性。结果显示:(1)StHSP17.7全长755bp,包含465bp开放阅读框,编码154个氨基酸。(2)StHSP17.7分子质量约为17.62kD,等电点7.91,为亲水性蛋白,C端含有保守序列Ⅰ和Ⅱ组成的ACD结构域,属于典型的sHSPs家族成员。(3)系统进化分析发现马铃薯小分子热激蛋白归为12个亚家族,其中StHSP17.7与马铃薯HSP17.6蛋白亲缘关系较近,属于小分子热激蛋白C的Ⅰ类家族成员;基因结构分析显示,在马铃薯48个sHSP基因中,StHSP17.7基因不含内含子,含1个内含子的基因有23个,占47.9%。(4)qRT-PCR分析显示,高温能够快速诱导StHSP17.7基因表达,且基因表达量呈爆发式变化,在24h达到最高值。研究表明,StHSP17.7基因明显参与了马铃薯对高温的响应。     

植物热激蛋白70基因家族及其生物学功能研究进展

植物热激蛋白70(heat shock protein 70,Hsp70)是热激蛋白家族中保守、普遍表达的家族,有两个主要功能区:N端核酸结合区和C端底物结合区。通常Hsp70具有分子伴侣功能,参与新生蛋白的折叠、转运、重折叠变性蛋白、协助降解变性蛋白;Hsp70不仅受高温胁迫诱导,且受其它多种胁迫诱导;同时,Hsp70参与植物正常发育过程。Hsp70基因分布广泛,序列高度保守,在植物基因组中以基因家族形式存在,最近广泛用于遗传多样性和系统发育研究。文章对植物中Hsp70的基因家族、结构、表达调控机制和生物学功能进行了综述。     

基因AtHsp17.6-C2启动子的克隆及其热诱导动力学研究(英文)

介绍一个植物表达系统,由热激蛋白基因(AtHsp17.6-C2)的启动子来驱动GUS基因的表达。在22℃生长条件下,稳定遗传的转基因植株中几乎检测不到GUS的活性。但是当温度升至34～37℃时,GUS的活性迅速升高。37℃是该植物表达系统最适诱导温度。转基因植株经37℃热诱导2小时后再返回22℃培育2小时,GUS的活性增加80多倍。多次热诱导实验表明这个表达系统是能够被重复多次热诱导的。实验结果表明这个植物诱导表达系统能够适用于多种目的需要。     

温度,热激蛋白与高粱育性的变化

高粱不育系3A在热激(43～45℃)诱导下结出了种子,由不育系转变为可育系。比较3A和3B线粒体在热激条件的热激蛋白得知,它们的热激蛋白是由核编码的,在细胞质中合成后才运到线粒体中,热激2h,3A出现70、31、24、18和16kDa 5条蛋白带,3B除出现上述5条蛋白带外还多出现96、94kDa 2条,而且70kDa含量比3A大。热激4h,3B的96、94kDa消失,两系趋于一致。此时,3A和3B线粒体总蛋白比热激前大量增加。此后HSPs急剧降低。热激8h,3B线粒体仅有70、31、24和16kDa 4条蛋白带,70kDa特别明显,而3A则全部消失。从而表明,HSPs在3B中是稳定的,在3A中是缺乏或不稳定的,这些差异可能与3B育性稳定性及3A不育性有关。     

柱状田头菇(茶薪菇)金属硫蛋白的分离纯化与特性研究

应用快速灌注色谱系统首次从经Cd2+诱导的柱状田头菇(茶薪菇)Agrocybecylindracea(DC.:Fr:)R.Maoire菌丝体中分离得到一种镉结合蛋白。通过SephadexG-75凝胶过滤层析,原子吸收光谱分析(AAS),巯基含量测定及紫外吸收光谱分析表明这种镉结合蛋白具有金属硫蛋白(metallothionein,MT)的理化性质:即分子量为6kDa、每分子MT含18个半胱氨酸残基并结合7个镉原子、具有镉硫金属簇的特征紫外吸收光谱,初步鉴定为茶薪菇Cd-MT。     

柱状田头菇(茶薪菇)金属硫蛋白的分离纯化与特性研究*

应用快速灌注色谱系统首次从经Cd2+诱导的柱状田头菇(茶薪菇)Agrocybecylindracea(DC.:Fr:)R.Maoire菌丝体中分离得到一种镉结合蛋白。通过SephadexG-75凝胶过滤层析,原子吸收光谱分析(AAS),巯基含量测定及紫外吸收光谱分析表明这种镉结合蛋白具有金属硫蛋白(metallothionein,MT)的理化性质:即分子量为6kDa、每分子MT含18个半胱氨酸残基并结合7个镉原子、具有镉硫金属簇的特征紫外吸收光谱,初步鉴定为茶薪菇Cd-MT。     

盐芥热激同源蛋白70基因(Thhsc70)的分子克隆及鉴定

热激蛋白70(hsp70s)具有分子伴侣的功能,其中在非胁迫条件下表达的hsp70s称为热激同源蛋白70(hsc70).为更好地了解hsc70基因的特性,鉴定了盐芥(Thellungiella halophila(C.A.Mey.)O.E. Schulz)中编码胞质hsc70蛋白的基因Thhsc70.实验结果表明:在非胁迫条件下,Thhsc70基因具有组织特异性表达;Thhsc70基因受热胁迫和冷胁迫的诱导表达,但几乎不受盐诱导和干旱诱导.Thhsc70基因在拟南芥中过量表达后提高了转基因拟南芥的热耐受性和冷耐受性.     

钙-钙调素信号系统参与热激信号转导的研究

根据作者实验室的研究工作结合国内外的研究动态讨论热激信号转导的Ca2 -CaM途径。作者实验室的工作表明,钙一钙调素(Ca^2 -CaM)信号系统参与植物热激信号转导。激光共聚焦扫描显微镜的观察结果表明,37℃热激可引起小麦胞内自由Ca。’浓度迅速提高。在Ca^2 存在条件下,热激也引起小麦CaM基因CaM1-2表达及CaM蛋白含量增加。Ca^2 可促进小麦热激基因hsp26和mp70表达和热激蛋白合成,而Ca^2 螯合剂EGTA、Ca^2 通道阻断剂异搏定和LaCl3、CaM抑制剂W7、TFP和CPZ明显降低热激基因hsp26和mp70表达和热激蛋白合成。EGTA、异搏定、TFP或CPZ也阻止小麦耐热性的获得。小麦CaM基因与热激基因的表达动力学研究表明CaM位于热激信号转导的上游,而Ca^2 是启动热激反应的胞内关键因子。凝胶阻滞分析的结果表明,Ca^2 -CaM在热激信号转导中的作用是通过激活热激转录因子的DNA结合活性来实现的。根据大量实验证据,作者提出在植物细胞内存在一条新的热激信号转导途径——钙一钙调素途径。     

Altered gene expression in plants with constitutive expression of a mitochondrial small heat shock protein suggests the involvement of retrograde regulation in the heat stress response

Heat stress can negatively affect crop productivity. One way in which plants attempt to alleviate the effects of heat stress is to induce the expression of genes encoding heat shock proteins (HSPs), including small HSPs (sHSPs). We produced transgenic lines of Arabidopsis thaliana expressing a transgene encoding a maize mitochondrial sHSP, ZmHSP22. The transgene, under the control of the cauliflower mosaic virus 35S promoter, is constitutively highly expressed in these lines. As demonstrated by confocal immunofluorescence microscopy and analyses of isolated mitochondria, ZmHSP22 is directed to the mitochondria of Arabidopsis and is processed into the mature form. These transgenic lines demonstrated altered expression of nuclear genes encoding the endogenous mitochondrial sHSP, AtHSP23.6, chloroplast localized AtHSP25.3, class I cytosolic AtHSP17.4, cytosolic AtHSP70-1 and chloroplast localized AtHSP70-6, but not cytosolic AtHSP70-15, following exposure to heat stress. This suggests that the expression of HSPs can be affected by heat-induced mitochondrial retrograde regulation. Three-week-old plants from the transgenic Arabidopsis lines expressing ZmHSP22 have increased thermotolerance, as measured by the maintenance of higher leaf mass following successive days with short periods of heat stress.     

Examination of cadmium-induced expression of the small heat shock protein gene, hsp30, in Xenopus laevis A6 kidney epithelial cells

Cadmium is a highly toxic environmental pollutant that has been classified as a human carcinogen. Toxicological responses to cadmium exposure include respiratory diseases, neurological disorders and kidney damage. In the present study, we have characterized the effect of cadmium on the accumulation of the small heat shock protein (HSP), HSP30, in Xenopus laevis A6 kidney epithelial cells. Incubation of A6 cells with cadmium chloride induced the accumulation of HSP30 protein and hsp30 mRNA. While HSP70 protein and hsp70 mRNA accumulation were also induced, the relative levels of actin remained relatively unaffected. Elevated levels of HSP30 were detected in cells undergoing prolonged exposure of cells to cadmium chloride or in cells recovering from cadmium chloride treatment. Immunocytochemical analysis of cadmium chloride-treated A6 cells revealed HSP30 accumulation primarily in the cytoplasm in a punctate pattern supplemented with larger HSP30 staining structures. Also, HSP30 co-localized with the F-actin cytoskeleton at higher cadmium chloride concentrations. The combination of mild heat shock temperatures plus cadmium chloride concentrations employed in this study resulted in a synergistic accumulation of HSP30 protein and hsp30 mRNA. Finally, in contrast to heat shock, prior exposure of Xenopus A6 cells to cadmium chloride treatment, sufficient to induce the accumulation of HSPs, did not protect the cells against a subsequent thermal challenge.     

Zinc Dyshomeostasis in Cardiomyocytes after Acute Hypoxia/Reoxygenation

This study aimed to assess the protective roles of polysaccharides from Agaricus blazei Murill (ABP) against cadmium (Cd)-induced damage in chicken livers. A total of 80 Hy-Line laying chickens (7 days old) were randomly divided into four groups (n = 20). Group I (control) was fed with a basic diet and 0.2 ml saline per day, group II (Cd-treated group) was fed with a basic diet containing 140 mg/kg cadmium chloride (CdCl₂) and 0.2 ml saline per day, group III (Cd + ABP-treated group) was fed with a basic diet containing 140 mg/kg CdCl₂ and 0.2-ml ABP solution (30 mg/ml) per day via oral gavage, and group IV (ABP-treated group) was fed with 0.2-ml ABP solution (30 mg/ml) per day via oral gavage. The contents of Cd and malondialdehyde (MDA), the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), the messenger RNA (mRNA) levels of inflammatory cytokines and heat shock proteins (HSPs), the protein levels of HSPs, and the histopathological changes of livers were evaluated on days 20, 40, and 60. The results showed that Cd exposure resulted in Cd accumulating in livers and inhibiting the activities of antioxidant enzymes (SOD and GSH-PX). Cd exposure caused histopathological damage and increased the MDA content, the mRNA levels of inflammatory cytokines (TNF-α, IL-1β, and IL-6) and HSPs (HSP27, HSP40, HSP60, HSP70, and HSP90) and the protein levels of HSPs (HSP60, HSP70, and HSP90). ABP supplementation during dietary exposure to Cd reduced the histopathological damage and decreased the contents of Cd and MDA and the expression of inflammatory cytokines and HSPs and improved the activities of antioxidant enzymes. The results indicated that ABP could partly ameliorate the toxic effects of Cd on chicken livers.     

Proteomic study of β-aminobutyric acid-mediated cadmium stress alleviation in soybean

The present study highlights the protective role of β-aminobutyric acid (BABA) in alleviating cadmium (Cd) stress in soybean. Proteomic analyses revealed that out of 66 differentially abundant protein spots in response to Cd challenge, 17 were common in the leaves of BABA-primed and non-primed plants. Oxygen-evolving enhancer protein 1 and ribulose bisphosphate carboxylase small chain 1 were detected in increase abundance in both groups of leaves. Among the 15 commonly decreased protein spots, the relative intensity levels of heat shock cognate 70-kDa protein, carbonic anhydrase, methionine synthase, and glycine dehydrogenase were partially restored after BABA treatment. Moreover, BABA priming significantly enhanced the abundance of the defense-related protein peroxiredoxin and glycolytic enzymes in response to Cd exposure. Additionally, the impact of Cd on the physiological state of BABA-primed and non-primed plants was analyzed using a biophoton technique. The finding of comparatively low biophoton emission in BABA-primed leaves under Cd stress indicates that these plants experienced less oxidative damage than that of non-primed plants. Proteomic study coupled with biophoton analysis reveals that BABA pretreatment helps the plants to combat Cd stress by modulating plants' defence mechanism as well as activating cellular detoxification system to protect the cells from Cd induced oxidative stress damages.     

Expression analysis of heat shock genes in the skin, spleen and blood of common carp (Cyprinus carpio) after cadmium exposure and hypothermia

Heat shock proteins are chaperones that play a pivotal role in controling multiple regulatory pathways such as stress defense, hormone signaling, cell cycle control, cell proliferation and differentiation, and apoptosis. In this study, the expression patterns of four well-known heat shock genes (hsp70, hsc70-1, hsc70-2 and hsp90α) were characterized in the skin, spleen and blood cells of the common carp, under unstressed conditions and after Cd2+ treatment or hypothermia. The examined genes were expressed in a tissue-specific manner: hsc70-2 was expressed constitutively, and was at best only slightly inducible; hsp90α exhibited a high basic expression in all three tissues, whereas hsc70-1 did so only in the blood cells, the expression of hsp70 proved to be below the level of detection in unstressed fish. Cold shock induced the expression of hsp genes in the spleen (hsp90α) and blood cells (hsp70, hsc70-1 and hsp90α), while Cd2+ treatment has no effect on the expression pattern. The highest inducibilities were detected in the skin: for hsp70 an induction of at least 20-fold after cadmium exposure, for hsc70-1 of at least 30-fold and for hsp90α of 3-fold after hypothermia.