植物抗病基因工程的研究进展及前景展望

近年来,随着植物抗病基因(尤其是抗病毒基因)的分离,植物抗病机制的分子生物学和植物抗病基因工程的研究轰轰烈烈地展开并取得重大突破。本文针对植物抗病基因工程的原理、抗病基因、转化方法等方面的进展进行了综述,并对抗病基因工程的应用前景做了展望。     

哺乳动物防御素的研究进展及其应用前景

防御素是生物界广泛存在的一类具微生物抗性的低分子短肽,其中哺乳动物防御素的抗性谱最为广泛,具有很高的应用潜力.概述了哺乳动物防御素的组成分布、立体结构、基因表达调控、抗菌谱等的国内外研究进展,并展望了其在医学和植物抗病基因工程中的应用前景.     

林木抗病基因工程研究进展

植物抗病性是当前植物病理学中研究的热点和难点之一。着重讨论植物抗病机制、抗病基因的转化方法及其在林木抗病基因工程中的应用情况,并对现阶段林木抗病基因工程中存在的主要问题和应用前景进行了讨论。     

植物防御素基因家族结构、功能及调控研究进展

防御素(Defensins,DF)是抗菌肽家族中最古老的一员。植物防御素是一类包含45-54个氨基酸残基,含有半胱氨酸(Cys)稳定的αβ模序,与哺乳动物及昆虫抗菌肽的亲缘关系非常密切,大部分植物防御素在体内被合成为具有信号肽序列的前体,并被分泌到胞外空间。植物防御素在宿主防御系统中起重要作用,不仅可抑制一系列的植物、动物及人类体内的细菌、真菌,还可杀伤一些肿瘤细胞及病原虫。植物防御素广泛存在于植物的花、茎、叶、果实、根和种子中,具有抗菌、抗肿瘤、抑制酶活、作为离子阻断剂和增加耐受性的功能,可作为新型的杀菌剂或新型的抗生素类药物。随着抗生素导致的耐药菌株的出现,对植物防御素的研究就显得尤为重要,特别是因其有效的抗真菌活性,植物防御素在作物抗病基因工程方面具有巨大的潜力。主要介绍了其发现、结构、类型、逆境调控功能、作用机制以及外源表达植物防御素基因等方面的最新研究进展,有助于将来对植物防御素基因家族进行更深入和全面的研究。     

植物抗白粉病的分子机理

随着分子生物学与其相关技术的飞速发展 ,已克隆到了一系列的植物抗病基因和防御基因 ,加深了对植物与病原微生物相互作用分子机制的了解 ,促进了植物抗病分子机理的研究。国内外许多学者对大麦抗白粉病的分子机制进行了较系统的研究 ,在拟南芥菜中也找到了许多抗白粉病基因 ,这些结果对研究其它植物抗白粉病机制提供了线索。本文就该方面的研究进展加以阐述 ,并讨论了抗病机理在抗病育种工作中的应用前景。     

植物抗病分子机制研究进展

植物抗病机制是目前研究的热点。在长期的进化过程中,植物形成了一系列复杂有效的防御机制来抵御、破坏病原物的侵染。植物抗病基因在植物抗性反应中起着重要的作用,植物一旦监测到病原物马上起始防御反应,并伴随着植物体内一系列细胞和生理生化的变化。近年来,基因沉默作为一个重要的细胞内防御外源核酸的机制,越来越受到科学家重视。综述了植物抗病基因和基因沉默机制在植物抗病反应中的重要作用,并对研究植物抗病机制的前景做了展望。     

植物抗病基因工程的研究进展

对植物抗病基因工程的原理、抗病基因、转化方法等方面的进展进行了综述,并对抗病基因工程的应用前景做了展望。     

植物抗细菌病害基因工程的研究进展及应用前景

植物抗细菌病害基因工程的主要方法包括 :抑制细菌致病和毒性因子 ,激活植物本身抗病机制 ,导入植物防御基因 ,导入非植物抗菌蛋白的编码基因 ,利用细胞调亡反应控制病害的发生。本文综述了这方面的研究进展及应用前景。     

转录因子在植物抗病基因工程中的研究进展

转录因子与顺式作用元件结合,可调控下游一系列基因的表达。通过基因工程手段使一个抗病转录因子基因在植物中超表达就相当于转入了多个抗病基因,从而提高综合抗病能力,因此,转录因子已成为近年来的研究热点。综述了乙烯应答元件结合因子(Ethylene-responsive element binding factors,AP2/EREBP)、MYB、WRKY、碱性亮氨酸拉链家族和homeodomain蛋白5种植物抗病相关转录因子的结构、功能特性、调控机制以及它们在植物抗病基因工程方面的研究成果,并展望了其应用前景。     

哺乳动物β-防御素研究进展

哺乳动物β-防御素是一类小分子的阳离子抗菌多肽。其具有独特的抗菌机制和广谱抗菌、抗病毒活性,同时还可作为免疫调节剂来激活与调节机体的免疫系统,在医药开发或是动植物抗病育种等方面都具有很好的应用前景。就哺乳动物β-防御素的分布、分子结构、生物学活性、作用机制及其应用等方面的研究作一综述。     

Plant defensins

Plant defensins are small, basic peptides that have a characteristic three-dimensional folding pattern that is stabilized by eight disulfide-linked cysteines. They are termed plant defensins because they are structurally related to defensins found in other types of organism, including humans. To date, sequences of more than 80 different plant defensin genes from different plant species are available. In Arabidopsis thaliana, at least 13 putative plant defensin genes (PDF) are present, encoding 11 different plant defensins. Two additional genes appear to encode plant defensin fusions. Plant defensins inhibit the growth of a broad range of fungi but seem nontoxic to either mammalian or plant cells. Antifungal activity of defensins appears to require specific binding to membrane targets. This review focuses on the classification of plant defensins in general and in Arabidopsis specifically, and on the mode of action of plant defensins against fungal pathogens.     

The Tomato Defensin TPP3 Binds Phosphatidylinositol (4,5)-Bisphosphate via a Conserved Dimeric Cationic Grip Conformation To Mediate Cell Lysis

Defensins are a class of ubiquitously expressed cationic antimicrobial peptides (CAPs) that play an important role in innate defense. Plant defensins are active against a broad range of microbial pathogens and act via multiple mechanisms, including cell membrane permeabilization. The cytolytic activity of defensins has been proposed to involve interaction with specific lipid components in the target cell wall or membrane and defensin oligomerization. Indeed, the defensin Nicotiana alata defensin 1 (NaD1) binds to a broad range of membrane phosphatidylinositol phosphates and forms an oligomeric complex with phosphatidylinositol (4,5)-bisphosphate (PIP2) that facilitates membrane lysis of both mammalian tumor and fungal cells. Here, we report that the tomato defensin TPP3 has a unique lipid binding profile that is specific for PIP2 with which it forms an oligomeric complex that is critical for cytolytic activity. Structural characterization of TPP3 by X-ray crystallography and site-directed mutagenesis demonstrated that it forms a dimer in a “cationic grip” conformation that specifically accommodates the head group of PIP2 to mediate cooperative higher-order oligomerization and subsequent membrane permeabilization. These findings suggest that certain plant defensins are innate immune receptors for phospholipids and adopt conserved dimeric configurations to mediate PIP2 binding and membrane permeabilization. This mechanism of innate defense may be conserved across defensins from different species.     

Antiviral Mechanisms of Human Defensins

Defensins are an effector component of the innate immune system with broad antimicrobial activity. Humans express two types of defensins, α- and β-defensins, which have antiviral activity against both enveloped and non-enveloped viruses. The diversity of defensin-sensitive viral species reflects a multitude of antiviral mechanisms. These include direct defensin targeting of viral envelopes, glycoproteins, and capsids in addition to inhibition of viral fusion and post-entry neutralization. Binding and modulation of host cell surface receptors and disruption of intracellular signaling by defensins can also inhibit viral replication. In addition, defensins can function as chemokines to augment and alter adaptive immune responses, revealing an indirect antiviral mechanism. Nonetheless, many questions regarding the antiviral activities of defensins remain. Although significant mechanistic data are known for α-defensins, molecular details for β-defensin inhibition are mostly lacking. Importantly, the role of defensin antiviral activity in vivo has not been addressed due to the lack of a complete defensin knockout model. Overall, the antiviral activity of defensins is well established as are the variety of mechanisms by which defensins achieve this inhibition; however, additional research is needed to fully understand the role of defensins in viral pathogenesis.     

Structural characterization of plant defensin protein superfamily

Plant defensins represent a major innate immune protein superfamily with strong inhibitory effects on infectious diseases of humans, antifungal/antibacterial activities, proteinase and insect amylase inhibitory activities. They are generally defined by their conserved cysteine scaffold with α-helix and triple strand anti parallel β-sheet connected to the scaffold. With the genome of more plant species being fully sequenced, significant information about newly sequenced defensin proteins has been revealed. In this paper, we identify members of defensin protein families across plant species and use protein-modeling-based structural reconstitution to reveal specific three dimensional hidden features of plant defensins mediating defense responses and other interesting biological activities in plants. Our data revealed that plant defensins are structurally similar to their insect counterparts despite the low amino acid sequence similarity between these two organisms. The molecular and structural relationship among plant defensins and defensins from other species is discussed.     

Differential effects of five types of antipathogenic plant peptides on model membranes

The effects of five antipathogenic plant peptides, wheat α-thionin, potato PTH1 defensin, barley LTP2 lipid transfer protein, and potato tuber DL1 and DL2 defensins, have been tested against phospholipid vesicles (liposomes). Wheat thionin very actively induces aggregation and leakage of negatively charged vesicles. LTP2 displays the same activities, although to a limited extent. Under certain conditions PTH1 and DL2 induce vesicle aggregation, but not leakage. Potato defensin DL1 failed to show any effect on liposomes. The same peptides have been assayed against a plant pathogenic bacterium, both the membrane-active and -inactive compounds having efficient antibacterial action.     

Antibacterial activity and mechanism of action of tick defensin against Gram-positive bacteria

Defensins are a major group of antimicrobial peptides and are found widely in vertebrates, invertebrates and plants. Invertebrate defensins have been identified from insects, scorpions, mussels and ticks. In this study, chemically synthesized tick defensin was used to further investigate the activity spectrum and mode of action of natural tick defensin. Synthetic tick defensin showed antibacterial activity against many Gram-positive bacteria but not Gram-negative bacteria and low hemolytic activity, characteristic of invertebrate defensins. Furthermore, bactericidal activity against pathogenic Gram-positive bacteria including Bacillus cereus, Enterococcus faecalis and methicillin-resistant Staphylococcus aureus was observed. However, more than 30 min was necessary for tick defensin to completely kill bacteria. The interaction of tick defensin with the bacterial cytoplasmic membrane and its ability to disrupt the membrane potential was analyzed. Tick defensin was able to disrupt the membrane potential over a period of 30-60 min consistent with its relatively slow killing. Transmission electron microscopy of Micrococcus luteus treated with tick defensin showed lysis of the cytoplasmic membrane and leakage of cellular cytoplasmic contents. These findings suggest that the primary mechanism of action of tick defensin is bacterial cytoplasmic membrane lysis. In addition, incomplete cell division with multiple cross-wall formation was occasionally seen in tick defensin-treated bacteria showing pleiotropic secondary effects of tick defensin.     

Recent advances in the research and development of human defensins

Human defensins are a family of cationic antimicrobial peptides with molecular weights of 4-5 kDa, containing a conserved six disulphide-linked cysteine motif. During the last two decades a series of endogenous alpha- and beta-human defensins were discovered. They exhibit a broad range of antimicrobial properties and are thought to be ideal therapeutic agents because of their potential ability to circumvent the problems of acquired resistance often observed with other antimicrobial therapies. Because of their appealing medical and pharmaceutical potential there has been an emphasis on human defensins in medical and molecular biology research in recent years. This paper aims to present a comprehensive review of recent advances in the study of human defensins including their discovery, classification, molecular properties, expression, mechanisms of action and potential medical applications. In addition, the advances in producing human defensins via genetic engineered cells are summarized from research works in our group (besides host cells including E. coli, B. subtilis and yeast systems, the cell-free protein synthesis system was also employed to express human beta-defensin-2) along with other related published works. The present challenges and prospects for the potential application of human defensins are also discussed.     

A novel role for defensins in intestinal homeostasis: regulation of IL-1beta secretion

Impaired expression of alpha-defensin antimicrobial peptides and overproduction of the proinflammatory cytokine IL-1beta have been associated with inflammatory bowel disease. In this study, we examine the interactions between alpha-defensins and IL-1beta and the role of defensin deficiency in the pathogenesis of inflammatory bowel disease. It was found that matrix metalloproteinase-7-deficient (MMP-7(-/-)) mice, which produce procryptdins but not mature cryptdins (alpha-defensins) in the intestine, were more susceptible to dextran sulfate sodium-induced colitis. Furthermore, both baseline and dextran sulfate sodium-induced IL-1beta production in the intestine were significantly up-regulated in MMP-7(-/-) mice compared with that in control C57BL/6 mice. To elucidate the molecular mechanism for the increased IL-1beta production in defensin deficiency in vivo, we evaluated the effect of defensins on IL-1beta posttranslational processing and release. It was found that alpha-defensins, including mouse Paneth cell defensins cryptdin-3 and cryptdin-4, human neutrophil defensin HNP-1, and human Paneth cell defensin HD-5, blocked the release of IL-1beta from LPS-activated monocytes, whereas TNF-alpha expression and release were not affected. Unlike alpha-defensins, human beta-defensins and mouse procryptdins do not have any effect on IL-1beta processing and release. Thus, alpha-defensins may play an important role in intestinal homeostasis by controlling the production of IL-1beta.     

Expression of BrD1, a plant defensin from Brassica rapa, confers resistance against brown planthopper (Nilaparvata lugens) in transgenic rices

Plant defensins are small (5-10 kDa) basic peptides thought to be an important component of the defense pathway against fungal and/or bacterial pathogens. To understand the role of plant defensins in protecting plants against the brown planthopper, a type of insect herbivore, we isolated the Brassica rapa Defensin 1 (BrD1) gene and introduced it into rice (Oryza sativa L.) to produce stable transgenic plants. The BrD1 protein is homologous to other plant defensins and contains both an N-terminal endoplasmic reticulum signal sequence and a defensin domain, which are highly conserved in all plant defensins. Based on a phylogenetic analysis of the defensin domain of various plant defensins, we established that BrD1 belongs to a distinct subgroup of plant defensins. Relative to the wild type, transgenic rices expressing BrD1 exhibit strong resistance to brown planthopper nymphs and female adults. These results suggest that BrD1 exhibits insecticidal activity, and might be useful for developing cereal crop plants resistant to sap-sucking insects, such as the brown planthopper.     

抗菌肽及抗菌肽转基因植物研究进展

抗菌肽是一类小分子多肽,在生物体内分布广泛,具有广谱的抗菌性,是生物体内天然防御系统的一部分。主要介绍了抗菌肽的性质,类型,作用机制及抗菌肽转基因植物的研究进展。