质膜转运蛋白及其与植物耐盐性关系研究进展

植物细胞质膜有两种主要功能：⑴溶质运输（进出细胞）,溶质运输主要由转运蛋白完成;⑵信号传导,即接收信号并引发细胞生理生化响应。盐分过多对植物的伤害主要是离子毒害。质膜转运蛋白活性环境变化能做现迅速响应。本文简要叙述了植物细胞质膜转运蛋白类型、分子特性、生理功能及其活性调节。介绍了植物细胞质膜Ｈ＾＋－ＡＴＰａｓｅ、质膜氧化还原系统、质膜离子载体和离子通道对盐胁迫的响应及其这些响应与植物耐盐性之间的关     

质膜转运蛋白及其与植物耐盐性关系研究进展

植物细胞质膜有两种主要功能：(1)溶质运输(进出细胞),溶质运输主要由转运蛋白完成;(2)信号传导,即接收信号并引发细胞生理生化响应。盐分过多对植物的伤害主要是离子毒害。质膜转运蛋白活性对环境变化能做出迅速响应。本文简要叙述了植物细胞质膜转运蛋白类型、分子特性、生理功能及其活性调节。介绍了植物细胞质膜H+_ATPase、质膜氧化还原系统、质膜离子载体和离子通道对盐胁迫的响应及其这些响应与植物耐盐性之间的关系。     

植物跨膜离子转运蛋白与其耐盐性关系研究进展

盐胁迫下植物吸收过多的N a ,使植物体内的离子平衡受到破坏,为了维持其正常生长细胞内的各种离子就必须保持平衡,而这一过程主要是由位于质膜和液泡膜上的离子转运蛋白完成的,并在植物耐盐性方面起关键作用。本文主要对响应盐胁迫的几种跨膜转运蛋白如:K /N a 离子转运蛋白、N a /H 逆向转运蛋白以及与其相关的H -ATPase等,在植物耐盐分子生物学方面的研究进展进行综述。     

盐胁迫对豌豆根液泡膜H^＋—ATPase活性及含量的影响

为了阐明液泡膜H^ -ATPase在盐胁迫下的作用和适应性调节机制,对豌豆（Pisum sativum L.）植株进行不同盐浓度和不同盐胁迫时间（1－3d）的处理后,分别测定液泡膜H^ -ATPase的H^ 转运活性、水解性和蛋白含量（A亚基）的变化。结果表明,100mmol/L和200mmol/L NaCl 处理1dH^ -ATPase的水解活性没有变化,而250mmol/L NaCl处理1d引起水解活性降低约25％。100mmol/L NaCl处理2d内水解活性没有变化,而第3天活性下降约20％。但是上述盐胁迫均能提高液泡膜H^ -ATPase的质子转运活性,说明盐胁迫后H^ -ATPase的水解活性和质子转运活性的变化不成比例,盐胁迫可能导致偶联比率的改变。Western blot研究发现,上述盐胁迫对液泡膜H^ -ATPase（A亚基）的含量基本无影响,仅100mmol/L NaCl处理3d后A亚基的量略有下降,这些结果证明,盐胁迫能刺激提高豌豆根液泡膜H^ -ATPase的H^ 泵效率,且泵效率的提高是源于偶联比率的改变,而不是由于ATP水解活性的提高和蛋白含量的增加。     

NaCl胁迫对盐芥质膜和液泡膜ATPase活性的影响

以盐生植物盐芥和中生植物拟南芥幼苗为材料,研究了盐胁迫对它们叶片和根质膜、液泡膜H+-ATPase、Ca2+-ATPases和K+-ATPase活性以及H+-ATPase、Na+/H+ 逆向转运蛋白表达的影响.结果显示:在NaCl胁迫下,盐芥叶片和根质膜的H+-ATPase活性分别比对照显著升高41%～212%和35%～53%,液泡膜的H+-ATPase分别显著升高281%～373%和4%～38%,而拟南芥却比相应对照都显著降低;相同盐浓度胁迫下,盐芥叶片的H+-ATPase活性比根部高4～8倍,盐芥根也远高于拟南芥.在NaCl胁迫下,盐芥叶片和根的液泡膜H+-ATPase蛋白质β亚基含量变化与其酶活性变化趋势一致,质膜Na+/H+ 逆向转运蛋白的表达量与Na+含量变化趋势一致.盐胁迫下盐芥根中Ca2+-ATPases和K+-ATPase活性的增加与根中Ca2+和K+含量呈显著正相关.研究发现,在盐胁迫条件下,盐芥能有效增强H+-ATPase蛋白和Na+/H+逆向转运蛋白表达,显著提高其根系与叶片质膜和液泡膜的H+-ATPase、Ca2+-ATPase和K+-ATPase活性,维持细胞质中较高的Ca2+和K+水平,从而缓解盐胁迫的伤害,增强耐盐性.     

蓝色温和凝胶双向电泳用于分析嗜盐菌质膜蛋白复合物

为了揭示细胞对盐胁迫渗透适应的分子机制,以新鉴定的中度嗜盐芽孢杆菌Bacillussp.I121为实验材料,分析了该嗜盐菌质膜上的盐胁迫响应蛋白.为此,通过蓝色温和凝胶双向电泳(BN/SDS-PAGE)对纯化的质膜组分进行了差异蛋白质组学研究.经MALDI-TOF/TOF质谱分析,鉴定了8个盐胁迫响应蛋白.盐胁迫诱导上调表达的蛋白质包括ABC型转运蛋白、3-磷酸甘油透性酶、嘧啶核苷转运蛋白和甲酸脱氢酶,下调表达的蛋白质包括琥珀酸脱氢酶(succinate dehydrogenase)铁硫亚基、黄素蛋白亚基、细胞色素b556亚基以及分子伴侣DnaJ的同源蛋白;酶活力测定结果表明胁迫条件下上述蛋白质的活性变化与表达量变化相一致.这些蛋白质中绝大多数属于高度疏水的跨膜蛋白,主要负责物质跨膜运输及能量代谢.上述结果表明,中度嗜盐菌Bacillus sp.I121可通过加快跨膜物质运输,同时抑制TCA循环完成盐胁迫条件下相容性溶质脯氨酸和四氢嘧啶的合成与积累.也进一步证明,蓝色温和凝胶双向电泳不仅可用于线粒体、叶绿体中蛋白质复合物的分析,也同样适用于细胞质膜上高度疏水蛋白复合物的比较研究.     

大豆耐盐机理及相关基因分子标记

大豆耐盐涉及多种生理代谢途径.耐盐大豆能够通过Cl-排除、控制Na 的吸收和转运、合成渗透调节物质、改变细胞膜膜脂组分及相关酶类的活性等多种形式来适应盐胁迫;野生大豆群体具有盐腺,从形态结构上适应盐逆境;大豆-根瘤菌共生体在盐胁迫下通过互作来提高整体的耐盐性.分子生物学技术应用于大豆耐盐研究,已获得了一些与耐盐相关基因连锁的分子标记.广泛搜集筛选大豆栽培种和野生种资源,利用分子生物学技术和基因工程提高大豆耐盐性,将成为未来大豆耐盐研究的主要内容.     

大豆耐盐机理及相关基因分子标记

大豆耐盐涉及多种生理代谢途径。耐盐大豆能够通过Cl-排除、控制Na+的吸收和转运、合成渗透调节物质、改变细胞膜膜脂组分及相关酶类的活性等多种形式来适应盐胁迫;野生大豆群体具有盐腺,从形态结构上适应盐逆境;大豆-根瘤菌共生体在盐胁迫下通过互作来提高整体的耐盐性。分子生物学技术应用于大豆耐盐研究,已获得了一些与耐盐相关基因连锁的分子标记。广泛搜集筛选大豆栽培种和野生种资源,利用分子生物学技术和基因工程提高大豆耐盐性,将成为未来大豆耐盐研究的主要内容。     

中度噬盐菌Bacillus sp.Ⅰ121的质膜蛋白质组学分析

膜蛋白是连接细胞与外界环境的重要载体,发挥着物质转运、信号传导等作用。通过对一种新分离出来的中度噬盐菌Bacillussp.Ⅰ121进行了质膜蛋白质组学分析,盐胁迫下该菌株主要靠积累脯氨酸来维持细胞渗透压。在对Bacillussp.Ⅰ121在不同盐浓度下的生长情况进行测定的基础上,我们选取了1%,5%,10%,15%（w/v）NaCl4个盐浓度进行细胞培养,并分别提取细菌的质膜,通过5%～20%的梯度凝胶SDS-PAGE电泳将疏水的膜蛋白进行分离,随后对盐胁迫响应的8条不同的蛋白条带进行了ESI-LC-MS/MS质谱分析。将所得结果在NCBI数据库中进行比对,最终成功鉴定出10种蛋白。这些蛋白涉及物质跨膜转运,膜的生物合成以及胁迫信号转导等功能。为膜蛋白在盐胁迫响应过程中的作用方面的研究开辟了新思路。     

蛋白质组学研究揭示的植物根盐胁迫响应机制

植物根是感知外界盐胁迫信号的首要器官。近年来,人们利用高通量的差异表达蛋白质组学技术,分析了水稻(Oryzasativa)、拟南芥(Arabidopsis thaliana)、大豆(Glycine max)、大麦(Hordeum vulgare)、小麦(Triticum aestivum)、木榄(Bruguieragymnorhiza)和匍匐翦股颖(Agrostis stolonifera)等植物根应答盐胁迫过程中蛋白质组的动态变化特征。通过整合植物根响应盐胁迫蛋白质组学研究结果,揭示了植物根部响应盐胁迫的多种调节机制,包括:利用多种信号通路与蛋白质磷酸化/去磷酸化感知并传递盐胁迫信号;通过膜蛋白与转运蛋白调节离子吸收/外排与区室化;通过抗氧化酶系统活性清除活性氧,并通过合成多种渗透调节物质与防御物质减轻细胞受到的伤害;通过改变参与糖类与能量代谢相关酶的表达调节能量代谢水平;通过细胞骨架动态重塑保持正常的细胞结构、物质运输与信息传递;通过转录、翻译与翻译后调控调节各种蛋白质的动态变化与相互作用;通过调控各种基础代谢与次生代谢水平保持细胞结构与代谢状态正常。     

Salt‐Treated Roots of Oryza australiensis Seedlings are Enriched with Proteins Involved in Energetics and Transport

Salinity is a major constraint on rice productivity worldwide. However, mechanisms of salt tolerance in wild rice relatives are unknown. Root microsomal proteins are extracted from two Oryza australiensis accessions contrasting in salt tolerance. Whole roots of 2‐week‐old seedlings are treated with 80 mM NaCl for 30 days to induce salt stress. Proteins are quantified by tandem mass tags (TMT) and triple‐stage Mass Spectrometry. More than 200 differentially expressed proteins between the salt‐treated and control samples in the two accessions (p‐value <0.05) are found. Gene Ontology (GO) analysis shows that proteins categorized as “metabolic process,” “transport,” and “transmembrane transporter” are highly responsive to salt treatment. In particular, mitochondrial ATPases and SNARE proteins are more abundant in roots of the salt‐tolerant accession and responded strongly when roots are exposed to salinity. mRNA quantification validated the elevated protein abundances of a monosaccharide transporter and an antiporter observed in the salt‐tolerant genotype. The importance of the upregulated monosaccharide transporter and a VAMP‐like protein by measuring salinity responses of two yeast knockout mutants for genes homologous to those encoding these proteins in rice are confirmed. Potential new mechanisms of salt tolerance in rice, with implications for breeding of elite cultivars are also discussed.     

野生大豆耐盐性研究进展

野生大豆对于拓宽大豆种质遗传基础和丰富大豆种质基因库具有重要意义.该文从野生大豆的资源概况及优良性状、耐盐机理和利用野生大豆提高栽培大豆耐盐性等三个方面,对近年来国内外有关野生大豆耐盐性的解剖结构、生理基础、分子生物学基础等方面的研究进展进行了系统综述,并提出野生大豆通过茎叶表皮上的"腺体"及对Na+和Cl-的排斥性,实现对盐渍环境的颉颃作用.较强的抗氧化能力、大豆异黄酮代谢和耐盐基因也是其适应盐渍环境的重要原因.今后应对野生大豆耐盐机理的遗传学基础进行深入研究,并通过种群保护以保障野生大豆的发掘鉴定和创新利用.     

Differential proteomic analysis of apoplastic proteins during initial phase of salt stress in rice

The plant cell apoplast, which consists of all the compartments beyond the plasma membrane, is implicated in a variety of functions during plant growth and development as well as in plant defence responses to stress conditions. To evaluate the role of apoplastic proteins in initial phase of salt stress, a 2-DE based differential proteomics approach has been used to identify apoplastic salt response proteins. Six salt response proteins have been identified, among them, an apoplastic protein OsRMC, which belongs to cysteine-rich repeat receptor like protein kinase subfamily but without the kinase domain, has shown drastically increased abundance in response to salt stress during the initial phase. Our results show, OsRMC negative regulates the salt tolerance of rice plants. These results indicated that plant apoplastic proteins may have important role in plant salt stress response signal pathway.Key words: rice, apoplast, proteomic, salt stress, receptor-like protein kinase, OsRMC     

花生萌发期耐盐性综合评价及耐盐种质筛选

为了解江苏花生种质资源的耐盐性,挖掘耐盐种质资源,本试验对47份花生种质材料进行耐盐性综合评价,以发芽势、发芽指数、活力指数、发芽率、鲜重、相对含水量、干重的相对值为鉴定指标,采用主成分分析、隶属函数法以及聚类分析方法,对其进行萌发期耐盐性综合评价及耐盐种质筛选。结果表明,花生种质材料萌发期的耐盐性强弱判定结果受多个指标影响,相对含水量和鲜重可以作为花生种质萌发期耐盐性的最佳鉴定指标,5 g/L Na Cl溶液可以作为花生萌发期耐盐性鉴定的合适浓度,47份花生材料划为5个耐盐级别,筛选出JP42、JP29、JP23、JP43、JP35、JP4等6份耐盐性强的种质,JP27和JP98为高度敏感材料,隶属函数法结合耐盐分级可以作为一种简便快速鉴定花生萌发期耐盐性的方法。     

A CIPK protein kinase targets sucrose transporter MdSUT2.2 at Ser254 for phosphorylation to enhance salt tolerance

Soil salinity is one of the major abiotic stressors that negatively affect crop growth and yield. Salt stress can regulate antioxidants and the accumulation of osmoprotectants. In the study, a sucrose transporter MdSUT2.2 was identified in apple. Overexpression of MdSUT2.2 gene increased salt tolerance in the transgenic apple, compared with the WT control “Gala.” In addition, it was found that protein MdSUT2.2 was phosphorylated at Ser²⁵⁴ site in response to salt. A DUAL membrane yeast hybridization system through an apple cDNA library demonstrated that a protein kinase MdCIPK13 interacted with MdSUT2.2. A series of transgenic analysis in apple calli showed that MdCIPK13 was required for the salt‐induced phosphorylation of MdSUT2.2 protein and enhanced its stability and transport activity. Finally, it was found that MdCIPK13 improved salt resistance in an MdSUT2.2‐dependent manner. These findings had enriched our understanding of the molecular mechanisms underlying abiotic stress.     

Ectopic Expression of Arabidopsis Glycosyltransferase UGT85A5 Enhances Salt Stress Tolerance in Tobacco

Abiotic stresses greatly influence plant growth and productivity. While glycosyltransferases are widely distributed in plant kingdom, their biological roles in response to abiotic stresses are largely unknown. In this study, a novel Arabidopsis glycosyltransferase gene UGT85A5 was identified as significantly induced by salt stress. Ectopic expression of UGT85A5 in tobacco enhanced the salt stress tolerance in the transgenic plants. There were higher seed germination rates, better plant growth and less chlorophyll loss in transgenic lines compared to wild type plants under salt stress. This enhanced tolerance of salt stress was correlated with increased accumulations of proline and soluble sugars, but with decreases in malondialdehyde accumulation and Na⁺/K⁺ ratio in UGT85A5-expressing tobacco. Furthermore, during salt stress, expression of several carbohydrate metabolism-related genes including those for sucrose synthase, sucrose-phosphate synthase, hexose transporter and a group2 LEA protein were obviously upregulated in UGT85A5-expressing transgenic plants compared with wild type controls. Thus, these findings suggest a specific protective role of this glycosyltransferase against salt stress and provide a genetic engineering strategy to improve salt tolerance of crops.     

Abiotic Stress Resistance,a Novel Moonlighting Function of Ribosomal Protein RPL44 in the Halophilic Fungus Aspergillus glaucus

Ribosomal proteins are highly conserved components of basal cellular organelles, primarily involved in the translation of mRNA leading to protein synthesis. However, certain ribosomal proteins moonlight in the development and differentiation of organisms. In this study, the ribosomal protein L44 (RPL44), associated with salt resistance, was screened from the halophilic fungus Aspergillus glaucus (AgRPL44), and its activity was investigated in Saccharomyces cerevisiae and Nicotiana tabacum. Sequence alignment revealed that AgRPL44 is one of the proteins of the large ribosomal subunit 60S. Expression of AgRPL44 was upregulated via treatment with salt, sorbitol, or heavy metals to demonstrate its response to osmotic stress. A homologous sequence from the model fungus Magnaporthe oryzae, MoRPL44, was cloned and compared with AgRPL44 in a yeast expression system. The results indicated that yeast cells with overexpressed AgRPL44 were more resistant to salt, drought, and heavy metals than were yeast cells expressing MoRPL44 at a similar level of stress. When AgRPL44 was introduced into M. oryzae, the transformants displayed obviously enhanced tolerance to salt and drought, indicating the potential value of AgRPL44 for genetic applications. To verify the value of its application in plants, tobacco was transformed with AgRPL44, and the results were similar. Taken together, we conclude that AgRPL44 supports abiotic stress resistance and may have value for genetic application.     

Cloning and Expression Analysis of Salt Responsive Gene from Kandelia candel

Identification of gene expression patterns in mangroves grown under salinity will help to reveal the molecular mechanisms of salt tolerance. Here, 10 cDNAs of genes were isolated from Kandelia candel and identified by representational difference analysis of cDNA (cDNA RDA) under different NaCl concentrations. Of five genes expressed preferentially under salt condition, two were unknown, three were two kinds of low molecular mass heat-shock proteins (sHSPs) and ADP-ribosylation factor, respectively. The expressions of other five genes were repressed under NaCl stress, two encoded cyclophilins, three were tonoplast intrinsic protein, early light-induced protein and 60S ribosomal protein, respectively.