金属硫蛋白和植物螯合肽在植物重金属耐性中的作用

植物螯合肽和金属硫蛋白广泛存在于植物界中,它们对植物耐重金属特别重要,能够与重金属形成复合物,以缓解重金属对植物的危害。本文就这两种金属螯合蛋白的结构、生物合成和基因调控,以及在植物体内缓解重金属毒害的作用方面作了简要介绍。     

铜、镉胁迫下外源NO介导的番茄解毒途径

一氧化氮(NO)作为信号分子,在抵御重金属胁迫中起重要作用,但对不同离子胁迫下的解毒机制尚缺乏研究.本研究采用营养液培养法,研究了铜(Cu)、镉(Cd)单一或复合胁迫下,番茄幼苗对Cu、Cd的吸收转运特性及对外源NO的响应机制.结果表明: 50 μmol·L^-1的Cu²⁺、Cd²⁺均显著抑制番茄植株的生长,其中Cd胁迫对生长的抑制效应远高于Cu胁迫.Cu、Cd单一或复合胁迫均使番茄根系Cu、Cd含量显著升高,但根系对Cu、Cd吸收存在严格选择性.根细胞对必需元素Cu表现出“奢侈吸收”的现象,而对毒性较强的Cd则吸收相对较少,胞内Cd浓度仅为Cu的1/10左右.外源NO处理可不同程度地缓解Cu、Cd胁迫,其中缓解Cd胁迫的效能更强.番茄对被动进入细胞的Cu、Cd具有相似的解毒机制:一方面,Cu、Cd胁迫诱导细胞质中产生谷胱甘肽(GSH)、植物螯合肽(PCs)和金属硫蛋白(MTs),络合过多的Cu、Cd离子,降低其生物毒性;另一方面,过多的Cu、Cd离子或螯合物被转运至液泡区隔化.外源NO通过调控GSH-GSSG(氧化型谷胱甘肽)氧化还原状态及GSH-PCs代谢方向的改变,促进Cu、Cd离子转运至液泡区隔化来缓解胁迫抑制;NO还可诱导植株叶片或根系表达更多的金属硫蛋白、GSH和PCs,而且上述响应普遍存在叠加效应.这可能是NO介导番茄对Cu、Cd胁迫的另一主要解毒途径.     

厚藤cDNA文库的构建和重金属镉耐受相关基因的筛选

厚藤（Ipomoea pes-caprae（L.） Sweet）是一种具有重要生态、观赏及药用价值的沙滩植物,对重金属镉（Cd）具有一定的富集能力,可作为Cd污染滨海地区的修复植物进行引种栽植和利用。本研究通过Gateway技术构建厚藤的cDNA文库,将该文库质粒转化酵母对Cd敏感的突变株ycf1△,采用全长cDNA过表达基因捕获系统（FOX）筛选厚藤重金属Cd胁迫耐受相关基因,并采用酵母互补实验进行基因的功能验证。本研究获得了2个能够恢复ycf1△对Cd敏感表型的重组质粒,经测序分析,该重组质粒包含的cDNA全长序列分别对应厚藤植物螯合肽合成酶基因（phytochelatin synthase）和金属硫蛋白基因（metallothionein）,分别将其命名为IpPCS和IpMT,通过功能分析,初步认定该基因为编码Cd耐受和解毒相关蛋白的候选基因。     

缺硫对镉胁迫下水稻幼苗非蛋白巯基物质含量和谷胱甘肽硫转移酶活性的影响

通过设置缺硫(S)处理,研究了镉(Cd)胁迫下水稻生长情况、幼苗Cd和非蛋白巯基含量以及谷胱甘肽硫转移酶(GST)活性的动态变化.结果表明:Cd胁迫明显抑制了水稻生长,显著诱导了巯基物质[非蛋白巯基(NPT)、谷胱甘肽(GSH)、植物螯合肽(PC)]的合成,GST活性表现出先升后降的趋势.缺S处理下,尽管水稻根部对Cd的吸收和向地上部的转运都有所增加,但Cd胁迫程度并未明显增强,巯基物质含量明显降低,根部GST活性提高.表明巯基物质和GST在水稻抗Cd胁迫过程中互为补充,在一定程度上减轻了Cd的毒性效应.     

酸性土壤上植物应对铝胁迫的过程与机制

铝胁迫是酸性土壤上影响作物产量最重要的因素之一.目前,全球土壤酸化程度进一步加剧了铝胁迫.植物可通过将铝离子与有机酸螯合储藏于液泡和从根系中排出铝毒.排出铝毒主要通过苹果酸转运蛋白ALMT和柠檬酸转运蛋白MATE的跨膜运输来实现.编码ABC转运蛋白和锌指转录因子的基因与植物抗铝胁迫有关.这些抗铝毒基因的鉴别使得通过转基因和分子标记辅助育种等生物技术来提高农作物的抗铝毒能力成为可能.最后提出了植物抗铝胁迫研究中需要解决的关键问题及今后的研究方向.     

过表达谷胱甘肽合成酶基因增强烟草对镉的耐受性

随着工业的发展,土壤污染问题愈发严重,利用基因工程修复土壤技术备受青睐,因此,开发重金属应答中的限速酶基因,将为植物修复重金属污染的土壤提供可应用的基因资源.通过RT-PCR及末端克隆方法获得枸杞谷胱甘肽合成酶 (Lycium chinense,Glutathione synthetase,LcGS>) 基因,采用半定量RT-PCR分析了枸杞LcGS在不同时间镉(Cd)胁迫下表达量的变化,LcGS表达量随着胁迫时间的延长而增强,胁迫9h、12h和24h 后LcGS表达量维持在较高水平.同时构建了植物双元表达载体pCAMBIA2300-LcGS,通过农杆菌介导的方法将LcGS基因转入烟草,PCR证明了LcGS基因成功整合到烟草基因组中,在Cd处理条件下,转基因植株谷胱甘肽 (glutathione,GSH)、植物螯合肽(phytochelatins,PCs)和叶绿素含量比对照组明显高,即转基因植株对重金属的耐逆性比对照组更强,因此,过表达GS植物将是植物修复重金属污染的一个有效策略.     

拟南芥MT-Ⅱ过量表达提高抗旱性

富含巯基的植物Ⅱ型金属硫蛋白(MT)对植物抵抗重金属胁迫具有重要作用,其中一个可能机制是金属硫蛋白可能猝灭重金属引起的氧化胁迫.利用转MT-Ⅱ基因和野生型拟南芥(Arabidopsis thaliana)植株来对比研究MT在胁迫过程中通过清除氧自由基,特别是H2O2而对植物抗旱性的影响.研究表明,转基因型拟南芥能有效维持体内氧化-还原势,减少MDA的产生,从而缓解干旱胁迫引起的伤害,提高抗旱性.     

植物金属硫蛋白及其重金属解毒机制研究进展

金属硫蛋白是一类分子量较小、富含Cys的金属结合蛋白,广泛分布于生物界。近年来从植物中克隆到许多编码金属硫蛋白的基因,并在研究基因表达模式、组织表达特异性以及基因结构,如启动子、内含子在染色体上的定位等方面取得了一定进展,但对其功能的研究还处于起步阶段。很多实验表明,植物金属硫蛋白可以通过其大量的Cys残基螯合重金属并清除活性氧,使植物避免氧化损伤。文章介绍了植物金属硫蛋白的分类、特征、基因结构及其在植物重金属解毒中的作用。      

重金属镉的转运蛋白研究进展

重金属镉(Cadmium,Cd)是一种工业和环境污染物,长期接触可对人体及其他动植物造成毒害。镉作为机体的非必需金属,需要借助必需金属的转运蛋白进入细胞。文章总结了机体中参与镉运输的关键转运蛋白,包括金属硫蛋白、谷胱甘肽、植物螯合素等富含半胱氨酸的蛋白质以及必需金属离子的转运蛋白等,以期为后续镉中毒防治靶点的研究提供一些参考。     

超积累植物伴矿景天镉耐受基因SpMT2的分离及功能鉴定

超积累植物由于其对重金属具有地上部超积累以及超耐受等特性,不仅是研究植物离子转运及毒性耐受的理想模式,而且在植物修复的发展和应用中具有不可替代的作用。伴矿景天是近年在我国境内发现的一种景天科镉(Cd)/锌(Zn)超积累植物。为鉴定其富集和耐受Cd的关键基因,笔者构建了其酵母表达cDNA文库,利用酵母的遗传互补系统筛选到一个极大提高了酵母抗Cd能力的基因SpMT2。SpMT2属于富含半胱氨酸(Cys)的金属硫蛋白(Metallothionein)家族。亚细胞定位表明SpMT2表达于酵母细胞质中,并特异地提高酵母对Cd的抗性。进一步研究发现SpMT2的表达显著降低了酵母液泡中Cd含量,但酵母吸收的总Cd含量无显著变化。推测抗性增加是由于SpMT2在酵母细胞质中通过螯合Cd从而降低Cd对酵母的毒害。qRT-PCR分析表明SpMT2在伴矿景天的根和地上部都高丰度表达,且不受Cd诱导变化。鉴于SpMT2也定位于植物细胞质中,结合上述结果,推测SpMT2可能在伴矿景天细胞质中螯合Cd,在降低Cd毒害的同时可能还保持Cd在细胞质中的流动性,从而在Cd长途转运过程中也发挥重要作用。     

Cadmium toxicity in crop plants and its alleviation by arbuscular mycorrhizal (AM) fungi: An overview

Cadmium (Cd), a toxic metal released into agricultural settings induces numerous changes in plant growth and physiology. The main known mechanisms of Cd toxicity include its affinity for sulfhydryl groups in proteins and its ability to replace some essential metals in active sites of enzymes, thus causing inhibition of enzyme activities and protein denaturation. This article reviews detrimental effects of Cd toxicity on the functional biology of plants and summarizes the mechanisms that are activated by plants to prevent the absorption or to detoxify Cd ions such as synthesis of antioxidants, osmolytes, phytochelatins, metallothioneins, etc. Arbuscular mycorrhizal (AM) fungi are reported to be present on the roots of plants growing in metal-contaminated soils and play an important role in metal tolerance. Through mycorrhizal symbiosis, heavy metals are immobilized in the rhizosphere through precipitation in the soil matrix, adsorption onto the root surface or accumulation within roots, and compartmentalized in aboveground parts of the plant. This article unfolds the potential role of AM fungi in enhancing Cd tolerance of plants.     

植物对重金属镉的耐受机制

镉离子（Cd^2＋）具有强植物毒性,抑制植物生长,甚至使植物死亡。由于长期的环境选择和适应进化,植物发展出耐受机制,可减轻或避免Cd^2＋的毒害。硫转运蛋白、硫还原相关酶类以及半胱氨酸、谷胱甘肽和植物螯合肽合成基因的表达受Cd^2＋调控。同时这些基因的过表达也能提高植物对Cd^2＋的耐性。植物抗氧化系统对Cd^2＋胁迫诱发的活性氧的清除作用,具转运Cd^2＋活性的质膜转运蛋白促进Cd^2＋经共质体途径向木质部运输、装载,而后随蒸腾流向地上部迁移,具转运Cd^2＋活性的液泡膜转运蛋白促进Cd^2＋进入液泡的隔离作用,都在植物对Cd^2＋的耐性中起作用。     

Effects of nitrogen on the activity of antioxidant enzymes and gene expression in leaves of Populus plants subjected to cadmium stress

The research aimed to verify the important physiological effect of nitrogen (N) on plants exposed to cadmium (Cd). The poplar plants were grown in a Hoagland nutrient solution and treated with extra N, Cd, and N + Cd. After treatment, plant growth and chlorophyll content were recorded. The oxidative stress, the activity of antioxidant enzymes, and the expression of related genes were also examined. The results indicated the plants treated with sole Cd presented obvious toxicity symptoms, i.e. growth inhibition, reactive oxygen species accumulation, and chlorophyll content decrement. However, when N was added to the plants under Cd stress, plant growth was enhanced, chlorophyll synthesis was promoted, and the oxidative stress was alleviated. Further, the expression of antioxidant enzymes genes was upregulated by N. The results indicated that N partially reversed the toxic effect of Cd on poplar plants, which can provide new methodology to enhance the phytoremediation technology for heavy metal pollution soil.     

硅缓解植物镉毒害的生理生态机制

镉是对生物毒性最强的污染物之一。过量的镉能够抑制植物的生长和光合作用,干扰矿质代谢并诱发氧化胁迫。硅作为一种有益元素,主要以Si(OH)_4的形态通过主动或被动方式被植物体吸收并转运到地上部分。硅对植物镉毒害具有缓解作用,但其缓解机制在不同物种、品种或生态型之间存在显著差异,并表现出一定的硅/镉浓度依赖性。总体上可概括为避性机制和耐性机制。避性机制包括:(1)在器官水平,减少植物根系对镉的吸收及其向地上部的转运;(2)在细胞水平,增强细胞壁对镉的吸附能力,减少共质体中镉的含量。耐性机制包括:(1)诱导细胞产生小分子螯合剂,增强对镉的螯合作用,减少细胞中游离态镉的含量;(2)增强抗氧化机制,减轻氧化胁迫;(3)改善光合作用和无机营养,促进植物生长。从植物对硅的吸收和转运、镉对植物的毒害作用以及硅对缓解植物镉毒害的生理生态机制3个方面进行了综述,并基于目前的研究现状和薄弱之处,对今后的研究重点进行了展望。     

Zinc resistance impairs sensitivity to oxidative stress in HeLa cells: protection through metallothioneins expression.

To analyze the effects of high concentrations of zinc ions on oxidative stress protection, we developed an original model of zinc-resistant HeLa cells (HZR), by using a 200 microM zinc sulfate-supplemented medium. Resistant cells specifically accumulate high zinc levels in intracellular vesicles. These resistant cells also exhibit high expression of metallothioneins (MT), mainly located in the cytoplasm. Exposure of HZR to Zn-depleted medium for 3 or 7 d decreases the intracellular zinc content, but only slightly reduces MT levels of resistant cells. No changes of the intracellular redox status were detected, but zinc resistance enhanced H2O2-mediated cytotoxicity. Conversely, zinc-depleted resistant cells were protected against H2O2-induced cell death. Basal- and oxidant-induced DNA damage was increased in zinc resistant cells. Moreover, measurement of DNA damage on zinc-depleted resistant cells suggests that cytoplasmic metal-free MT ensures an efficient protection against oxidative DNA damage, while Zn-MT does not. This newly developed Zn-resistant HeLa model demonstrates that high intracellular concentrations of zinc enhance oxidative DNA damage and subsequent cell death. Effective protection against oxidative damage is provided by metallothionein under nonsaturating zinc conditions. Thus, induction of MT by zinc may mediate the main cellular protective effect of zinc against oxidative injury.     

植物对重金属耐性的分子生态机理

植物适应重金属元素胁迫的机制包括阻止和控制重金属的吸收、体内螯合解毒、体内区室化分隔以及代谢平衡等。近年来,随着分子生物学技术在生态学研究中的深入应用,控制这些过程的分子生态机理逐渐被揭示出来。菌根、根系分泌物以及细胞膜是控制重金属进入植物根系细胞的主要生理单元。外生菌根能显著提高寄主植物的重金属耐性,根系分泌物通过改变根际pH、改变金属物质的氧化还原状态和形成络合物等机理减少植物对重金属的吸收。目前,控制菌根和根系分泌物重金属抗性的分子生态机理还不清楚。但细胞膜跨膜转运器已得到深入研究,相关金属离子转运器被鉴定和分离,一些控制基因如铁锌控制运转相关蛋白(ZIP)类、自然抵抗相关巨噬细胞蛋白(Nramp)类、P_1B-type ATPase类基因已被发现和克隆。金属硫蛋白(MTs)、植物螯合素(PCs)、有机酸及氨基酸等是植物体内主要的螯合物质,它们通过螯合作用固定金属离子,降低其生物毒性或改变其移动性。与MTs合成相关的MT-like基因已经被克隆,PCs合成必需的植物螯合素合酶(PCS), 即γ-Glu-Cys二肽转肽酶(γ-ECS) 的编码基因已经被克隆,控制麦根酸合成的氨基酸尼克烟酰胺(NA)在重金属耐性中的作用和分子机理也被揭示出来。ATP 结合转运器(ABC)和阳离子扩散促进器(CDF) 是植物体内两种主要膜转运器,通过它们和其它跨膜方式,重金属被分隔贮藏于液泡内。控制这些蛋白转运器合成的基因也已经被克隆,在植物中的表达证实其与重金属的体内运输和平衡有关。热休克蛋白(HSP)等蛋白类物质的产生是一种重要的体内平衡机制,其分子机理有待进一步研究。重金属耐性植物在这些环节产生了相关响应基因或功能蛋白质,分子克隆和转基因技术又使它们在污染治理上得到了初步的应用。     

Cellular Mechanisms in Higher Plants Governing Tolerance to Cadmium Toxicity

Cadmium (Cd) is an inorganic mineral in the earth's crust. Cadmium entry into the environment occurs through geogenic and anthropogenic sources. Industrial activities including mining, electroplating, iron and steel plants, and battery production employ Cd during their processes and often release Cd into the environment. When disseminated into soil, Cd can be detrimental to agro-ecosystems because it is relatively mobile and phytotoxic even at low concentrations. Cadmium's phytotoxicity is due to reductions in the rate of transpiration and photosynthesis and chlorophyll concentration resulting in retardation of plant growth, and an alteration in the nutrient concentration in roots and leaves. In response to Cd toxicity, plants have developed protective cellular mechanisms such as synthesis of phytochelatins and metallothioneins, metal compartmentalization in vacuoles, and the increased activity of antioxidant enzymes to neutralize Cd-induced toxicity. While these direct protective mechanisms can help alleviate Cd toxicity, other indirect mechanisms such as microelements (zinc, iron, manganese, and selenium) interfering with Cd uptake may decrease Cd concentration in plants. This comprehensive review encompasses the significance of Cd, portals of contamination and toxicity to plants, and implications for crop production. Various mitigation strategies with the beneficial effects of zinc, iron, manganese, and selenium in activating defence mechanisms against Cd stress are discussed. Furthermore, this review systematically identifies and summarises suitable strategies for mitigating Cd-induced toxicity in plants.     

Differential cadmium stress tolerance in five indian mustard (Brassica juncea L.) cultivars: An evaluation of the role of antioxidant machinery

The presence of Cadmium (Cd) in the agricultural soils affects horticultural cultivars and constrains the crop productivity. A pot experiment was performed using five cultivars of mustard (Brassica juncea L.) to evaluate the difference in their response to Cd toxicity under greenhouse conditions. The pots containing reconstituted soil were supplied with different concentration of CdCl₂ (0, 25, 50, 100 or 150 mg Cd kg⁻¹ soil). Increasing concentration of Cd in the soil resulted in decreased growth, photosynthesis and yield. Maximum significant reduction in growth, photosynthesis and yield were observed with 150 mg Cd kg⁻¹ soil in all the cultivars. Our results indicate that the cultivar Alankar is found to be more tolerant to Cd stress, recording higher plant dry mass, net photosynthesis rate, associated with high antioxidant activity and low Cd content in the plant leaves and thus less oxidative damage. Cultivar RH30 experienced maximum damage in terms of reduction in growth, photosynthesis, yield characteristics and oxidative damage and emerged as sensitive cultivar. The data of tolerance index of Alankar were found to be higher among all tested mustard cultivars which indicate its higher tolerance to Cd. Better coordination of antioxidants protected Alankar from Cd toxicity, whereas lesser antioxidant activity in RH30 resulted in maximum damage. Cultivars of mustard were ranked with respect to their tolerance to Cd: Alankar > Varuna > Pusa Bold > Sakha > RH30, respectively.Key words: antioxidants, cadmium, growth, mustard cultivars, photosynthesis, stress, yield