低温对耐寒与不耐寒玉米线粒体膜的影响

线粒体是一种比较敏感的细胞器。很多环境因素的影响都能迅速引起线粒体发生变化。不耐寒植物受低温影响后线粒体膜的功能也会发生一定的改变。我们前曾报道玉米黄化幼苗经低温处理后(4℃,24小时)线粒体的耗氧率显著增加,呼吸控制率和氧化磷酸化效率明显下降,抗氰氧化途径基本失活。与此同时,电镜观察表明,线粒体的超细微结构也发生明显变化。本文比较低温对耐寒与不耐寒玉米线粒体膜的影响。实验的初步结果表明,与不耐寒品种不同,耐寒玉米品种的黄化幼苗经同样低温条件处理后其线粒体的氧化活性,呼吸控制,氧化磷酸化效率以及抗氰氧化途径都没有发生明显的变化。这提示,玉米经低温处理后线粒体膜的变化与耐寒性能之间具有一定的内在联系,从而根据线粒体膜的变化情况可能为鉴别玉米品种耐寒性提供一些生化指标。     

CBF:平衡植物低温应答与生长发育的关键

低温是影响植物生长发育以及植被分布的重要环境因子。目前,低温信号研究中比较清楚的是CBF依赖的低温信号途径。该文总结了近年来有关CBF的研究成果,详细介绍了CBF家族成员在植物耐寒性中的重要作用,着重分析与讨论CBF介导的低温调控网络及一系列复杂调控机制。理解CBF的复杂作用机制有助于了解植物中CBF介导的冷信号如何平衡耐寒性与生长发育,进而有助于耐寒作物的培育。     

植物应答低温胁迫机制的研究进展

低温是植物生长过程中遇到的主要环境胁迫因子之一,而植物响应低温胁迫是一个多因素协同作用的过程,涉及到复杂的基因表达调控网络。尤其是低温下植物体内生理生化、细胞骨架结构及基因表达调控等方面的改变及相关机制,一直受到研究者的普遍关注。该文主要从细胞学及分子生物学等角度入手,将低温胁迫下植物对低温的响应及可能机制进行综述,着重对植物通过细胞内部细胞器结构与功能的改变来抵御或适应低温,尤其对细胞骨架,以及低温信号转导受体及中间体、下游胁迫相关基因的表达及其在细胞内部的调控及应答机制等方面的作用进行探讨,为耐低温植物新品种的培育及农业生产实践提供理论指导。     

低温对耐寒与不耐寒玉米线粒体膜的影响

线粒体是一种比较敏感的细胞器。很多环境因素的影响都能迅速引起线粒体发生变化。不耐寒植物受低温影响后线粒体膜的功能也会发生一定的改变。我们前曾报道玉米黄化幼苗经低温处理后(4℃,24小时)线粒体的耗氧率显著增加,呼吸控制率和氧化磷酸化效率明显下降,抗氰氧化途径基本失活。与此同时,电镜观察表明,线粒体的     

低温诱导蛋白及其与植物的耐寒性研究进展

低温诱导蛋白是植物在温度逆境条件下诱导产生的一系列蛋白,以抗冻蛋白、脱水蛋白、热激蛋白和热稳定蛋白较多,而且低温诱导蛋白质一旦在体内形成,植物体就会尽快地适应外界环境,表现出较强的抗逆性.本文对几种主要的低温诱导蛋白——抗冻蛋白、脱水蛋白、热激蛋白和热稳定蛋白的特性及其与植物耐寒性的关系研究进行综述,以期为进一步阐明植物耐寒的分子机制以及提高植物耐寒力研究提供新的思路.     

昆虫抗冻耐寒能力的测定与分析方法

昆虫抗冻耐寒能力因其理论意义和实践价值成为当前生物学和生态学的重要研究内容。尤其昆虫抗冻耐寒能力的测定与分析是昆虫低温生物学的热点问题。本文从昆虫生态,及生理生化层面阐述了昆虫抗冻策略和耐寒机制类型。进一步介绍了昆虫抗冻耐寒能力的测定与分析方法:一方面,以昆虫种群为对象,分析低温对种群存活的胁迫作用,如低温实验中种群的存活率,致死中温度或致死中时间,冷伤害上限温度,冷害低温总和,以及低温冷伤害的死亡速率等。另一方面,以昆虫个体为对象,测定个体为适应低温环境而采取响应机制,如昆虫个体过冷却点、含水量、能量物质、抗冻小分子物质和抗冻蛋白含量等。在未来,从微观上看随着低温生物学拓展到基因组、转录组、蛋白质组及代谢组层次的研究,从宏观上看随着越冬代昆虫种群数量动态及其迁飞转移行为规律与栖息地微环境气候和区域性景观格局特征等的关系研究,有利于更全面地和深入地了解昆虫类群的抗冻策略或耐寒机制,从而为更系统地建立昆虫抗冻耐寒能力的测定与分析方法体系提供可靠指标。     

雌雄福寿螺耐寒能力的差异

为了解福寿螺耐寒性是否存在性别差异,对华南地区的雌雄福寿螺进行了过冷却研究及低温胁迫下存活率调查,并对入冬前后其体内耐寒性生理生化物质进行测定.结果表明:雌雄螺的过冷却点均值分别为-6.83℃和-6.26℃,雌螺过冷却点显著低于雄螺;进入冬季后,福寿螺自由水含量明显下降,结合水、甘油、脂肪、糖原含量均出现明显上升,除甘油外,其余各耐寒性生理指标均存在极显著的性别差异;未经过冷驯化的雌雄福寿螺经过10℃处理7d后存活率均在90%以上,继续在5℃下培养7 d后存活率出现明显的性别差异和壳高差异.野外调查发现在福寿螺自然种群中雌螺多于雄螺.表明福寿螺雌螺的耐寒能力强于雄螺.这对进一步探索福寿螺的生态适应性、自然性比及其耐寒性机制有一定的参考价值.     

雌雄福寿螺耐寒能力的差异

为了解福寿螺耐寒性是否存在性别差异,对华南地区的雌雄福寿螺进行了过冷却研究及低温胁迫下存活率调查,并对入冬前后其体内耐寒性生理生化物质进行测定.结果表明: 雌雄螺的过冷却点均值分别为-6.83 ℃和-6.26 ℃,雌螺过冷却点显著低于雄螺;进入冬季后,福寿螺自由水含量明显下降,结合水、甘油、脂肪、糖原含量均出现明显上升,除甘油外,其余各耐寒性生理指标均存在极显著的性别差异;未经过冷驯化的雌雄福寿螺经过10 ℃处理7 d后存活率均在90%以上,继续在5 ℃下培养7 d后存活率出现明显的性别差异和壳高差异.野外调查发现在福寿螺自然种群中雌螺多于雄螺.表明福寿螺雌螺的耐寒能力强于雄螺.这对进一步探索福寿螺的生态适应性、自然性比及其耐寒性机制有一定的参考价值.     

转基因技术在能源植物育种中的应用

由于能源危机与环境污染问题日益严重,能源植物以其安全、环保、可再生和低成本等特性,成为能源开发的一个热点。随着转基因技术的不断进步,利用转基因技术培育高产、优质、高效新型能源植物新品种也取得了相应的成果。本文简要介绍了能源植物的概念和分类,概述了转基因技术在提高植物总生物量、降低植物木质素的含量、在植物中过表达纤维素降解酶、以及提高油料植物含油量等方面的应用现状,并探讨了该技术在能源植物遗传改良中的应用前景,以期为后续的能源植物新品种培育等研究和应用提供参考。     

凡纳滨对虾TCP-1-Beta基因的克隆及其与耐寒性状的相关性

为研究凡纳滨对虾耐寒性状的分子机理,研究克隆了凡纳滨对虾耐寒相关基因TCP-1-Beta并对其与耐寒性状的关系进行了研究.根据电子克隆所得序列设计引物,采用RT-PCR方法克隆得到长1691 bp的凡纳滨对虾TCP-1-Beta基因序列,其中包括1608 bp的完整开放阅读框,编码535个氨基酸残基.然后,运用荧光定量PCR对TCP-1-Beta基因进行时空表达谱的分析:组织表达谱的结果显示,该基因在凡纳滨对虾肝胰腺组织中表达量最高;不同低温处理下的表达结果显示,在28℃和15℃处理下基因表达量无显著变化,13℃开始呈上调表达,11℃时表达量升高;13℃低温处理发现该基因在24h内表达量无显著变化,但36h后表达量显著升高.采用PCR-RFLP方法对216尾凡纳滨对虾TCP-1-Beta基因进行了SNP多态性检测,并将其与耐寒性状进行了关联分析.在该基因编码区第420碱基上发现G/A突变,方差分析结果表明该位点的不同基因型与耐寒力指标CDH值相关(P＜0.05),其中GG基因型的耐寒能力比AA基因型强.     

植物抗冻基因研究进展

低温是作物生长一大限制因素,许多作物不能在低温下生长或者容易受到冻害,所以通过基因工程的方法,转入抗冻基因,是扩大作物生长区域,稳定作物产量的有效办法。目前通过对植物抗冻基因在抗冻机理、研究技术、应用等方面的研究,已经取得了一定的成果。综述抗冻基因的来源、抗冻机理、研究技术,并做适度的展望。     

The mechanism by which fish antifreeze proteins cause thermal hysteresis

Antifreeze proteins are characterised by their ability to prevent ice from growing upon cooling below the bulk melting point. This displacement of the freezing temperature of ice is limited and at a sufficiently low temperature a rapid ice growth takes place. The separation of the melting and freezing temperature is usually referred to as thermal hysteresis, and the temperature of ice growth is referred to as the hysteresis freezing point. The hysteresis is supposed to be the result of an adsorption of antifreeze proteins to the crystal surface. This causes the ice to grow as convex surface regions between adjacent adsorbed antifreeze proteins, thus lowering the temperature at which the crystal can visibly expand. The model requires that the antifreeze proteins are irreversibly adsorbed onto the ice surface within the hysteresis gap. This presupposition is apparently in conflict with several characteristic features of the phenomenon; the absence of superheating of ice in the presence of antifreeze proteins, the dependence of the hysteresis activity on the concentration of antifreeze proteins and the different capacities of different types of antifreeze proteins to cause thermal hysteresis at equimolar concentrations. In addition, there are structural obstacles that apparently would preclude irreversible adsorption of the antifreeze proteins to the ice surface; the bond strength necessary for irreversible adsorption and the absence of a clearly defined surface to which the antifreeze proteins may adsorb. This article deals with these apparent conflicts between the prevailing theory and the empirical observations. We first review the mechanism of thermal hysteresis with some modifications: we explain the hysteresis as a result of vapour pressure equilibrium between the ice surface and the ambient fluid fraction within the hysteresis gap due to a pressure build-up within the convex growth zones, and the ice growth as the result of an ice surface nucleation event at the hysteresis freezing point. We then go on to summarise the empirical data to show that the dependence of the hysteresis on the concentration of antifreeze proteins arises from an equilibrium exchange of antifreeze proteins between ice and solution at the melting point. This reversible association between antifreeze proteins and the ice is followed by an irreversible adsorption of the antifreeze proteins onto a newly formed crystal plane when the temperature is lowered below the melting point. The formation of the crystal plane is due to a solidification of the interfacial region, and the necessary bond strength is provided by the protein "freezing" to the surface. In essence: the antifreeze proteins are "melted off" the ice at the bulk melting point and "freeze" to the ice as the temperature is reduced to subfreezing temperatures. We explain the different hysteresis activities caused by different types of antifreeze proteins at equimolar concentrations as a consequence of their solubility features during the phase of reversible association between the proteins and the ice, i.e., at the melting point; a low water solubility results in a large fraction of the proteins being associated with the ice at the melting point. This leads to a greater density of irreversibly adsorbed antifreeze proteins at the ice surface when the temperature drops, and thus to a greater hysteresis activity. Reference is also made to observations on insect antifreeze proteins to emphasise the general validity of this approach.     

Cold adaptation in the phytopathogenic fungi causing snow molds

Snow molds are psychrophilic or psychrotrophic fungal pathogens of forage crops, winter cereals, and conifer seedlings. These fungi can grow and attack dormant plants at low temperatures under snow cover. In this review, we describe the biodiversity and physiological and biochemical characteristics of snow molds that belong to various taxa. Cold tolerance is one of the important factors related to their geographic distribution, because snow molds develop mycelia under snow cover and because they should produce intra- and extracellular enzymes active at low temperatures for growth and infection. Basidiomycetous snow molds produce extracellular antifreeze proteins. Their physiological significance is to keep the extracellular environment unfrozen. The psychrophilic ascomycete Sclerotia borealis shows normal mycelial growth under frozen conditions, which is faster than that on unfrozen media at optimal growth temperature. This fungus does not produce extracellular antifreeze proteins, but osmotic stress tolerance enables the fungus to grow at subzero temperatures. In conclusion, different taxa of snow molds have different strategies to adapt under snow cover.     

Antifreeze proteins in higher plants

Overwintering plants produce antifreeze proteins (AFPs) having the ability to adsorb onto the surface of ice crystals and modify their growth. Recently, several AFPs have been isolated and characterized and five full-length AFP cDNAs have been cloned and characterized in higher plants. The derived amino acid sequences have shown low homology for identical residues. Theoretical and experimental models for structure of Lolium perenne AFP have been proposed. In addition, it was found that the hormone ethylene is involved in regulating antifreeze activity in response to cold. In this review, it is seen that the physiological and biochemical roles of AFPs may be important to protect the plant tissues from mechanical stress caused by ice formation.     

差示扫描量热法直接测定沙冬青抗冻蛋白的热滞效应

用差示扫描量热法直接测定了从沙冬青中提取的一种抗冻蛋白（ＡＦＰ）组分的低温热行为。结果表明,该组分的低温热行为远较文献报道的各种抗冻蛋白复杂。在降、升温过程中,在低和高温侧都给出两个放或吸热峰,两个峰表现出相互独立而又相互依存的热滞行为。低温峰的热滞活性远高于高温岭。我们认为,这种ＡＦＰ分子对水及冰晶很可能有两种不同的相互作用和影响。     

Targeted expression of redesigned and codon optimised synthetic gene leads to recrystallisation inhibition and reduced electrolyte leakage in spring wheat at sub-zero temperatures

Antifreeze proteins (AFPs) adsorb to ice crystals and inhibit their growth, leading to non-colligative freezing point depression. Crops like spring wheat, that are highly susceptible to frost damage, can potentially be made frost tolerant by expressing AFPs in the cytoplasm and apoplast where ice recrystallisation leads to cellular damage. The protein sequence for HPLC-6 α-helical antifreeze protein from winter flounder was rationally redesigned after removing the prosequences in the native protein. Wheat nuclear gene preferred amino acid codons were used to synthesize a recombinant antifreeze gene, rAFPI. Antifreeze protein was targeted to the apoplast using a Murine leader peptide sequence from the mAb24 light chain or retained in the endoplasmic reticulum using C-terminus KDEL sequence. The coding sequences were placed downstream of the rice Actin promoter and Actin-1 intron and upstream of the nopaline synthase terminator in the plant expression vectors. Transgenic wheat lines were generated through micro projectile bombardment of immature embryos of spring wheat cultivar Seri 82. Levels of antifreeze protein in the transgenic lines without any targeting peptide were low (0.06–0.07%). The apoplast-targeted protein reached a level of 1.61% of total soluble protein, 90% of which was present in the apoplast. ER-retained protein accumulated in the cells at levels up to 0.65% of total soluble proteins. Transgenic wheat line T-8 with apoplast-targeted antifreeze protein exhibited the highest levels of antifreeze activity and provided significant freezing protection even at temperatures as low as −7°C.     

A re-evaluation of the role of type IV antifreeze protein

A lipoprotein-like antifreeze protein (type IV AFP) has previously been isolated only from the blood plasma of the longhorn sculpin. However, the plasma antifreeze activity in all individuals of this species tested from Newfoundland and New Brunswick waters ranges from low to undetectable. A close relative of the longhorn sculpin, the shorthorn sculpin, does have appreciable antifreeze activity in its blood but this is virtually all accounted for by the α-helical, alanine-rich type I AFP, other isoforms of which are also present in the skin of both fishes. We have characterized a putative ortholog of type IV AFP in shorthorn sculpin by cDNA cloning. This 12.2-kDa Gln-rich protein is 87% identical to the longhorn sculpin’s type IV AFP. Recombinant versions of both orthologs were produced in bacteria and shown to have antifreeze activity. Immunoblotting with antibodies raised to type IV AFP shows this protein present in longhorn sculpin plasma at levels of less than 100 μg/mL, which are far too low to protect the blood from freezing at the temperature of icy seawater. This confirms the results of direct antifreeze assays on the plasmas. It appears that type IV AFP has the potential to develop as a functional antifreeze in these fishes but may not have been selected for this role because of the presence of type I AFP. Consistent with this hypothesis is the observation that the type IV AFP gene has not been amplified the way functional antifreeze protein genes have in all other species examined.     

& 转录因子CBF在植物抗寒中的重要作用

低温能够诱导植物许多基因的表达,从而使植物具有抗寒性,这种现象称为冷驯化。对于植物冷驯化的分子机理,目前研究的最多的是CBF转录因子调控的信号转导途径,其作用途径可归纳为：CBF(C-repeat Binding Factor)转录因子→CRT/DRE(C-repeat /Dehydration Responsive Element)基序→COR基因表达→植物抗寒性增加。研究CBF转录因子在抗寒中的作用机制,能为提高植物的抗寒性,培育抗寒作物品种提供新方向。      

动物抗冷性的基因表达机制

简要综述了包括抗冻蛋白作用机理在内的动物抗冷性的基因表达调节机制。通过在寒冷逆境下转录抗冻蛋白、纤维蛋白原等耐冻性相关蛋白基因,形成mRNA,然后在各种转录翻译调节因子的调节下表达形成蛋白质和酶,从而形成冰农耐性或冰冻避性或毁损修复反应。其中,转录后调节和上调节是动物常用的抗冷基因表达机制。小分子有机物质在提高动物的耐冻性中有非常的增效作用。     

植物耐冷性基因工程

温度决定物种的分布,同时还影响作物的产量和品质。植物耐冷的机制涉及到许多方面,包括膜脂组成的变化、可混溶溶质的积累、抗氧化酶活性的提高、低温相关基因的诱导表达等。由于植物的耐冷性状由多基因控制,采用传统的育种方法往往难以取得理想的结果,而植物基因工程技术的发展及应用则提供了另外可能的途径,可以通过转移耐冷性状形成的关键基因从而对植物进行改良。本文从膜脂组成、可混溶溶质、抗冻蛋白、抗氧化酶和诱导植物低温相关基因的转录因子等方面对植物耐冷性的基因工程研究进行了综述,以期为植物育种者和从事冷胁迫机制研究的工作者提供参考。