植物病原细菌Ⅲ型效应子的双向作用

病原细菌Ⅲ型效应子对寄主植物防卫反应的抑制和诱导决定了互作中植物抗(感)病性状。本文概述了植物病原菌Ⅲ型效应子抑制和诱导寄主植物防卫反应的双向作用,所涉及的抑制和诱导作用主要包括植物过敏反应、植物细胞壁介导的防卫反应以及植物抗病信号介导的抗性反应等。本文还对植物病原菌Ⅲ型效应子双向作用的研究和应用进行了展望。     

谷胱苷肽及其类似物诱导大豆活性氧和植保素的积累

 谷胱苷肽(Glutathione, GSH)是一种重要的抗氧化剂, 但它能诱导一些豆科等植物的防卫反应。本文采用大豆悬浮细胞体系分析了GSH及其衍生物诱导大豆活性氧和大豆植保素大豆素(glyceollin)的积累。GSH诱导大豆素和黄苷元的积累, 并有一定的剂量关系。N-乙酰半胱氨酸、巯基乙醇、二硫苏糖醇没有激发子活性, 但氧化型谷胱苷肽能激发大豆的防卫反应。在巯基被修饰的化合物中, S-对叠氮苯甲基谷胱苷肽和S-对氯苯甲基谷胱苷肽能够诱导大豆素和H₂O₂的产生, S-己基谷胱苷肽也能诱导一定量的H₂O₂积累。水杨酸和蛋白酶抑制剂DFP能增强芫菁素或者酵母细胞壁激发子诱导的大豆素积累, 但对GSH没有相似的增强效果。这些结果表明GSH中的巯基并不是激发所必需的, 大疏水基团能弥补巯基的作用。另外, GSH诱导大豆素积累信号传导途径可能与芫菁素的不同。     

植物水通道蛋白结构与功能及其识别与转导水稻黄单胞菌Hpa1信号的机制

 植物水通道蛋白PIP不仅担负细胞间或细胞内外水分子输导的基本功能,还参与植物-微生物互作与植物防卫反应,这种双重功能的调控机制目前还不清楚。水稻OsPIP1;2和拟南芥AtPIP1;4可以与水稻黄单胞III型泌出蛋白Hpa1互作,Hpa1定位于植物细胞的质外体,诱导过氧化氢在质外体产生及向原生质转运,进而影响植物防卫反应与对病原细菌的抗性。根据植物水通道蛋白拓扑结构与病原细菌Ⅲ型分泌系统工作模型,水稻OsPIP1;2与Hpa1互作的功能域是互作发生的分子基础。互作引发信号转导,调控过氧化氢信号从植物细胞的质外体向原生质转运与植物防卫反应。由于Hpa1对Ⅲ效应蛋白来说具有转位子的功能特征,OsPIP1;2-Hpa1还可能对水稻黄单胞菌Ⅲ型效应蛋白从细菌细胞向植物细胞转运发生调控作用。围绕这些设想进行研究,可以深入阐释水稻-黄单胞菌互作机制,同时为植物水通道蛋白功能调控提供新的见解。     

激发素与植物互作及抗病防卫信号传导

激发素是一类由疫霉属Phytophthora等真菌分泌的,可诱导茄科、十字花科等植物过敏性反应和系统获得抗病性的蛋白类激发子.激发素与植物的互作系统为研究植物抗病防卫信号传导途径提供了一个极好的模式.最近发现:激发素具有转移甾醇的活性,而形成甾醇和激发素复合物又是激发素与受体蛋白互作和激活抗病防卫信号传导途径的前提条件,也是对最近提出的病原激发子与受体互作"保卫假说"的支持.用125I标记法,找到了激发素的受体蛋白,由分子量约为160kD和50kD的2个亚基构成,被认为是Ca2 通道.在激发素激活的烟草抗病防卫信号传导途径中,蛋白质的磷酸化起着至关重要的作用,抑制蛋白激酶的活性,可阻止离子交换、活性氧进发、植保素合成等多种防卫反应;Ca2 参与诱导活性氧进发和植保素合成,活性氧进发与细胞死亡相关连,而植保素的合成与活性氧进发无关,过敏性反应对抗病防卫反应的建立不是必需的.激发素抗病基因工程在广谱抗病育种中具有广泛的应用前景.     

寡糖诱导植物防卫反应的信号转导

寡糖作为一种生物类激发子,可以诱导植物产生防卫反应,提高植物的抗病性,从而抵御病原物的入侵。本文就寡糖的种类、特点及诱导植物防卫反应信号转导等方面作一评述。     

蜡质芽孢杆菌AR156对辣椒的防病促生机理研究

 蜡质芽孢杆菌AR156是南京农业大学与新沂中凯农用化工有限公司合作开发的生物农药。为了解AR156对辣椒的防病促生机制, 本文研究了AR156在温室条件下对青枯劳尔氏菌引起的辣椒青枯病的生防效果, 对辣椒的促生作用, 在辣椒根围的定殖能力, 诱导植物细胞防卫反应, 如活性氧积累和胼胝质沉积, 以及植物防御相关酶的活力。结果表明, 蜡质芽孢杆菌AR156对辣椒青枯病的温室防效高达73.31%。AR156的使用使辣椒植株干重增加 22.30%, 并能稳定的在辣椒根围定殖, 接种60 d后, 其定殖量为5×10⁵ cfu·g^-1FW。AR156预处理后挑战接种病原菌能诱导植株更迅速的产生细胞防卫反应, 并可显著提高超氧化物歧化酶(SOD)和过氧化物酶(POD)的活力。可见AR156菌株诱导的植物细胞防卫反应, 提高植物防御相关酶活和在辣椒根围稳定的定殖能力使其产生对病害的广谱抗性。研究还发现 AR156菌剂可增加辣椒叶片叶绿素的含量, 这可能是该菌剂促进辣椒生长的原因之一。     

非寄主互作中抗病信号NO及H_2O_2对水稻细胞过敏反应的诱导机制

意义与目的细胞过敏性死亡反应(过敏反应,hypersensitive reaponae,HR)是植物抗病的特征性反应之一。HR引起植物细胞快速死亡,有效地将病原菌限制在侵染位点,阻止其进一步侵染周围健康组织,同时激活植物其他抗病反应的发生。已有研究表明,植物细胞过敏反应和动物细胞程序性死亡具有相似特征,是一有序过程。即在信号分子诱导下,调控植物基因的表达,最终导致细胞快速死亡。目前对诱导植物细胞过敏反应的信号组分尚不清楚。利用细胞悬浮培养体系,已初步揭示活性氧产生,离子通道通透性的改变,一氧化氮(NO)的产生可诱导植物细胞过敏反应,表明…     

细胞骨架解聚药物对小麦与叶锈菌互作诱发的细胞过敏性反应的影响

 小麦(Triticum aestivum)品种洛夫林10和叶锈菌小种366组成不亲和组合,小麦叶片发生过敏性坏死反应(HR)是小麦抵抗叶锈菌侵染的重要因素。在接种前给小麦叶片分别预注射微管解聚药物磺草硝(oryzalin)和微丝解聚药物细胞松弛素D (cytochalasin D,CD),结果表明2种药物注射使得寄主因叶锈菌侵染诱导的细胞过敏性坏死数目明显减少,并且注射药物的浓度越大,寄主细胞发生HR的数量越少。说明肌动蛋白和微管蛋白的聚合状态是诱发小麦叶片发生HR防卫反应所必需的,细胞骨架在小麦抵抗叶锈菌侵染过程中可能起着重要作用。     

电压依赖性阴离子通道VDAC3参与拟南芥先天免疫

 线粒体外膜蛋白电压依赖性阴离子通道(VDAC)是动物细胞凋亡调控系统中的关键组分,但其在植物中的功能还不明确。拟南芥VDAC家族有4个成员,它们都能被病原菌诱导,VDAC1在由病原菌所引起的超敏性细胞死亡中发挥作用已有报道,但VDAC家族其他成员是否参与植物免疫反应还未见报道。本研究以拟南芥相关T-DNA插入突变体为材料,研究了VDAC3基因在植物抗病反应中的功能。病原相关分子模式flg22诱导的活性氧产生和胼胝质沉积在VDAC3突变体中都显著增强,表明VDAC3参与了病原相关分子模式触发的免疫反应。此外,效应因子激发的超敏性细胞死亡在vdac3突变体中也更明显,显示VDAC3也参与了效应子触发的免疫反应。以上结果说明,VDAC3在多个层面参与植物的抗病反应。     

植物防卫基因研究进展

植物在与病原微生物共同进化的过程中，发展了一系列拮抗病原物侵染的复杂的防御反应体系。在植物与病原互作中，有大量的基因诱导表达，它们编码的蛋白质参与植物对病原物的防卫生反应，这类基因称之为防卫相关基因（defense related genes)，或简称为防卫基因（defense genes)。根据防卫基因表达产物及其功能，可大致分为次生物质合成基因、水解酶和病程相关基因、细胞壁修饰有关基因和清除活性氧的细胞内防卫酶系统基因四大类。从1902年人们发现病原物侵染而引起植物过敏反应起，人们对防卫反应进行了广泛深入的研究，到目前为止，防卫基因仍然是植保和遗传学家们研究的前沿和重点。     

N-terminal domain including conserved flg22 is required for flagellin-induced hypersensitive cell death in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Flagellin in Pseudomonas syringae is a potent elicitor of defense responses including hypersensitive cell death in dicot plants. The oligopeptides flg22 consisting of 22 conserved amino acids near the N-terminus of flagellins is reported to induce plant defense responses. Because glycosylation of the central domain of flagellin affects its elicitor activity, we investigated whether any peptide sequence in addition to flg22 is required for flagellin-induced hypersensitive reaction. A study of recombinant flagellin polypeptides indicated that the N-terminal domain including the conserved flg22 is required for flagellin-induced hypersensitive cell death in Arabidopsis thaliana.     

Glycosylation of flagellin from Pseudomonas syringae pv. tabaci 6605 contributes to evasion of host tobacco plant surveillance system

Pseudomonas syringae pv. tabaci (Pta) possesses a genetic region composed of two open reading frames (ORFs), fgt1 and fgt2, that are involved in glycosylation of flagellin. The deletion mutant Δfgt1 produced non-glycosylated flagellin, and exhibited reduced ability to cause disease in the host tobacco plant. Flagellin is known to induce plant defense responses, and the recognition of flagellin by Arabidopsis thaliana is mediated by a conserved N-terminal region, flg22, in flagellin and a leucine-rich repeat domain in the FLS2 receptor. Because flg22 localizes inside the flagellum, polymerized flagellum needs to be dissociated to be recognized. Therefore, the effect of glycosylation on flagella stability was investigated. The polymerized flagella from glycosylated flagellins were more resistant to heat treatment than those from non-glycosylated flagellins, suggesting that the glycosylation of flagellin contributes to the structural stability of flagella and prevents exposure of the flg22 region. Polymerized flagella from Pta Δfgt1 flagellin and depolymerized and glycosylated flagellin from Pta wild type induced cell death and callose deposition, and inhibited seedling growth in tobacco more effectively, whereas polymerized flagella from Pta wild-type flagellin caused a low level of these responses. These results suggest Pta might have evolved the flagellin glycosylation system to evade detection and defense response of a host by increasing flagella stability and suppressing their dissociation.     

Expression-based genotyping of the rice blast resistance genes in the elite maintainer line Yixiang1B

Plants employ extracellular immune receptors to perceive conserved pathogen-associated molecular patterns (PAMPs), triggering the first layer of defense known as pattern-triggered immunity (PTI). The understanding of PTI is mainly based on studies focusing on leaves. Plants are vulnerable to attack by various root pathogens including plant-parasitic nematodes. Evidence is accumulating that phytonematodes utilize their secreted effectors to suppress PTI to enable infection. PTI assays used for characterizing nematode effectors are often conducted in a non-host plant or tissue, such as leaves, because of lacking of root assays. Thus, there is a need for PTI assays in roots of host plants. Here, we tested two bacterial PAMPs (flg22 and flgII-28) and two nonpathogenic bacteria (Pseudomonas fluorescens and P. syringae strain DC3000 ΔhrcQ-U) for their ability to induce PTI responses, including the induction of defense gene expression and callose deposition, in roots of tomato and potato. We found that flg22 and the two nonpathogenic bacteria are potent in inducing defense gene expression and callose deposition in tested roots, demonstrating for the first time induction of PTI in roots of solanaceous plants. Effectors GrCEP12 and Hs10A06 were previously indicated to be involved in PTI suppression. Consistently, upon elicitor treatment, roots of transgenic plants overexpressing GrCEP12 and Hs10A06, respectively, showed a reduced level of defense gene expression or no induction of callose deposition compared to control roots. Taken together, our established root PTI assays represent a valuable tool that will facilitate the study of phytonematodes and potentially other root pathogens in their manipulation of plant immunity.     

A recombinant flagellin fragment,which includes the epitopes flg22 and flgII-28, provides a useful tool to study flagellin-triggered immunity

Plants and animals independently evolved the ability to recognize flagellin (also called FliC), the building block of the bacterial flagellum, as part of their innate immune response. While animals recognize a relatively large region of FliC, most plants recognize one or two short epitopes of FliC: flg22 and flgII-28. However, since most research in plants has focused on flg22 and flgII-28 and not the actual FliC protein, the importance of any FliC region beyond the two epitopes in plant immunity is poorly understood. Here we report cloning, overexpression, and purification of a Pseudomonas syringae FliC fragment from amino acid 1 to 143, which includes both FliC epitopes and the adjacent alpha helices. Exposing Arabidopsis thaliana leaves to FliC_1–143 did not reveal any additional FliC recognition capabilities beyond flg22. However, while the kiwifruit species Actinidia arguta did not respond to either flg22 or flgII-28, treatment of A. arguta leaves with FliC_1–143 triggered a significant reactive oxygen response, indicating recognition. This result suggests that in some plant species, recognition of FliC requires regions of FliC beyond the two well-known epitopes and that FliC_1–143 represents a useful tool in the study of plant immunity.     

野油菜黄单胞菌效应蛋白AvrXccC与拟南芥BIK1在植物体内相互作用

 AvrXccC是野油菜黄单胞菌Xcc8004的一个III型分泌效应蛋白。前期研究发现AvrXccC在拟南芥rar1突变体上发挥毒性功能,并且抑制植物的先天免疫反应。但是,AvrXccC在植物体内的毒性靶标还不清楚。本研究发现AvrXccC与植物体内的蛋白激酶BIK1发生特异的相互作用。然而,在拟南芥原生质体中,AvrXccC 并不能抑制鞭毛蛋白诱导的BIK1迁移,表明AvrXccC不是通过抑制BIK1的磷酸化来发挥功能的。有趣的是,AvrXccC可以在体外直接被BIK1磷酸化,我们推测AvrXccC可能作为BIK1的底物从而干扰了鞭毛蛋白诱导的先天免疫反应。     

水稻VQ37基因的克隆及其在病原侵染下的表达

 VQ蛋白作为转录辅助蛋白,在植物的生长、发育和抗逆等生理过程中发挥重要的调节功能。本研究采用RT-PCR从水稻叶片中克隆了VQ37基因的完整cDNA序列。VQ37 cDNA长622 bp,具有长为546 bp的完整开放读码框,编码蛋白质长181个氨基酸,具有FxxxVHxVTG的VQ基序变体。系统进化分析表明,水稻VQ37与短花药野生稻(Oryza brachyantha)、大麦(Hordeum vulgare)和Dichanthelium oligosanthes等禾本科植物亲缘关系近;除水稻旁系同源物VQ39外,VQ37与短花药野生稻中的XP 015699121亲缘关系最近。原生质体瞬间表达实验证实VQ37定位在细胞核中。荧光定量PCR分析显示,VQ37基因的组织特异性表达和诱导表达结果与启动子顺式元件预测基本一致。VQ37在叶片中的表达丰度最高,其次是叶鞘、茎、穗、根和花,在胚和胚乳中无表达。VQ37受纹枯病菌(Rhizoctonia solani)和稻瘟病菌(Magnaporthe oryza)显著诱导,而不受白叶枯细菌(Xanthomonas oryzae pv. oryzae)诱导;与此一致的是,VQ37受真菌病原相关模式(PAMP)分子几丁质寡糖快速诱导,而不受细菌鞭毛蛋白flg22影响。茉莉酸甲酯和乙烯利能显著诱导VQ37的表达,而水杨酸对其表达无明显影响。上述结果提示,VQ37可能调控水稻对稻瘟病菌和纹枯病菌防御反应,这种调节作用可能依赖于茉莉酸/乙烯介导的信号途径,而与水杨酸信号途径无关。该研究为阐释水稻VQ37基因在水稻抗病反应中的调节功能提供了基础。     

C-terminal region of plant ferredoxin-like protein is required to enhance resistance to bacterial disease in Arabidopsis thaliana

Protein phosphorylation is an important biological process associated with elicitor-induced defense responses in plants. In a previous report, we described how plant ferredoxin-like protein (PFLP) in transgenic plants enhances resistance to bacterial pathogens associated with the hypersensitive response (HR). PFLP possesses a putative casein kinase II phosphorylation (CK2P) site at the C-terminal in which phosphorylation occurs rapidly during defense response. However, the contribution of this site to the enhancement of disease resistance and the intensity of HR has not been clearly demonstrated. In this study, we generated two versions of truncated PFLP, PEC (extant CK2P site) and PDC (deleted CK2P site), and assessed their ability to trigger HR through harpin (HrpZ) derived from Pseudomonas syringae as well as their resistance to Ralstonia solanacearum. In an infiltration assay of HrpZ, PEC intensified harpin-mediated HR; however, PDC negated this effect. Transgenic plants expressing these versions indicate that nonphosphorylated PFLP loses its ability to induce HR or enhance disease resistance against R. solanacearum. Interestingly, the CK2P site of PFLP is required to induce the expression of the NADPH oxidase gene, AtrbohD, which is a reactive oxygen species producing enzyme. This was further confirmed by evaluating the HR on NADPH oxidase in mutants of Arabidopsis. As a result, we have concluded that the CK2P site is required for the phosphorylation of PFLP to enhance disease resistance.     

β-葡寡糖诱导植物抗病性的研究进展

β-葡寡糖作为一种植物激发子,可高效诱导植物产生抗病性,因此被普遍认为是一种病原物相关的分子模式.其作用的发挥主要是通过与细胞膜上的受体相互识别,引起受体构象改变产生跨膜信号,再经过一系列的胞内信号传导,调控防卫基因的表达,积累次生代谢产物,诱导植物抗性来实现.诱抗活性不仅受到寡糖聚合度和化学修饰基团的影响,而且植物对于结构上有差异的β-葡寡糖激发子的识别也是大相径庭.本文就β-葡寡糖诱导植物产生抗病性的研究进展进行了综述.     

拟南芥的抗病信号传导途径

 拟南芥是研究植物与病原物相互作用的模式植物。植物感病和抗病取决于病原物无毒基因产物和寄主抗病基因产物的识别,以及随后的相关防卫反应的激活。在拟南芥的抗病过程中,水杨酸、茉莉酸、乙烯等信号分子都不同程度地参与着抗病过程中的不同环节,起着非常重要的作用。由于这些信号分子在对不同病原菌的抗性中的作用存在差异,因而将抗病信号传导分为依赖于水杨酸和依赖于茉莉酸/乙烯的途径。本文将着重讨论这些信号分子在植物系统获得抗性以及诱导系统抗性中的作用。