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1.
作为活体营养专性寄生真菌,条形柄锈菌(小麦条锈病)在侵染过程中通过形成吸器向寄主细胞释放效应蛋白,干扰寄主的防卫反应,促进其侵染与致病。因此,条形柄锈菌效应蛋白的鉴定与功能研究对揭示其毒性机理具有重要意义。本实验室前期完成了条形柄锈菌CYR31生理小种吸器转录组分析,从中鉴定得到一个吸器特异诱导表达分泌蛋白Hasp68,利用农杆菌侵染在烟草细胞中瞬时表达该基因,能够抑制小鼠促细胞凋亡蛋白Bax诱导的细胞程序性死亡,鉴定为条形柄锈菌候选效应蛋白。Hasp68基因全长318bp,编码105_aa,N-端包含20_aa的信号肽,无保守结构域。BlastX分析表明Hasp68为条形柄锈菌特有效应蛋白,在其他真菌中无同源蛋白,且在条形柄锈菌16个菌系中呈较低的序列多态性,表明其在条形柄锈菌的进化过程中相对保守。借助荧光假单胞菌EtHAn的三型分泌系统,在小麦细胞中过表达Hasp68能够抑制由非致病细菌引起的PTI(PAMP-triggered immunity)相关胼胝质的积累;同时,也能抑制小麦与无毒条形柄锈菌互作中ETI(effector-triggered immunity)相关的活性氧爆发和过敏性坏死反应,表明效应蛋白Hasp68具有抑制寄主免疫反应的功能。利用酵母双杂交系统筛选Hasp68在小麦中的互作蛋白,发现其与组织蛋白酶B(cathepsin B)TaCTSB互作,双分子荧光技术进一步验证二者在烟草细胞中共表达存在互作,初步揭示了效应蛋白Hasp68的互作靶标。  相似文献   

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水稻抗稻瘟病天然免疫机制及抗病育种新策略   总被引:3,自引:0,他引:3  
何峰  张浩  刘金灵  王志龙  王国梁 《遗传》2014,36(8):756-765
稻瘟病是水稻最严重的病害之一,由子囊菌(Magnaporthe oryzae)引起。利用抗病品种是防治稻瘟病最经济、最有效的措施。近年来,稻瘟病已发展为研究植物与病原真菌分子互作机制的模式系统,在水稻与稻瘟菌互作和寄主抗性分子生物学、基因组学和蛋白组学等领域取得了一系列重要的研究成果。文章综述了近年来水稻抗稻瘟病两种天然免疫机制,即病原菌相关分子模式诱导和效应蛋白诱导的抗病机制研究的最新进展,讨论了GWAS、TALLEN、CRISPR和HIGS等基因组研究新方法和新技术在水稻抗病育种中的应用,并对目前稻瘟病抗性机制研究和抗病育种中的问题和挑战进行了探讨和展望。  相似文献   

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  总被引:1,自引:0,他引:1  
A Pseudomonas syringae pv. pisi effector protein, AvrRPS4, triggers RPS4 -dependent immunity in Arabidopsis. We characterized biochemical and genetic aspects of AvrRPS4 function. Secretion of AvrRPS4 from Pst DC3000 is type III secretion-dependent, and AvrRPS4 is processed into a smaller form in plant cells but not in bacteria or yeast. Agrobacterium -mediated transient expression analysis of N-terminally truncated AvrRPS4 mutants revealed that the C-terminal 88 amino acids are sufficient to trigger the hypersensitive response in turnip. N-terminal sequencing of the processed AvrRPS4 showed that processing occurs between G133 and G134. The processing-deficient mutant, R112L, still triggers RPS4 -dependent immunity, suggesting that the processing is not required for the AvrRPS4 avirulence function. AvrRPS4 enhances bacterial growth when delivered by Pta 6606 into Nicotiana benthamiana in which AvrRPS4 is not recognized. Transgenic expression of AvrRPS4 in the Arabidopsis rps4 mutant enhances the growth of Pst DC3000 and suppresses PTI (PAMP-triggered immunity), showing that AvrRPS4 promotes virulence in two distinct host plants. Furthermore, full virulence activity of AvrRPS4 requires both proteolytic processing and the KRVY motif at the N-terminus of processed AvrRPS4. XopO, an Xcv effector, shares the amino acids required for AvrRPS4 processing and the KRVY motif. XopO is also processed into a smaller form in N. benthamiana , similar to AvrRPS4, suggesting that a common mechanism is involved in activation of the virulence activities of both AvrRPS4 and XopO.  相似文献   

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The soybean cyst nematode (SCN), Heterodera glycines, is one of the most destructive pathogens of soybeans. SCN is an obligate and sedentary parasite that transforms host plant root cells into an elaborate permanent feeding site, a syncytium. Formation and maintenance of a viable syncytium is an absolute requirement for nematode growth and reproduction. In turn, sensing pathogen attack, plants activate defence responses and may trigger programmed cell death at the sites of infection. For successful parasitism, H. glycines must suppress these host defence responses to establish and maintain viable syncytia. Similar to other pathogens, H. glycines engages in these molecular interactions with its host via effector proteins. The goal of this study was to conduct a comprehensive screen to identify H. glycines effectors that interfere with plant immune responses. We used Nicotiana benthamiana plants infected by Pseudomonas syringae and Pseudomonas fluorescens strains. Using these pathosystems, we screened 51 H. glycines effectors to identify candidates that could inhibit effector-triggered immunity (ETI) and/or pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). We identified three effectors as ETI suppressors and seven effectors as PTI suppressors. We also assessed expression modulation of plant immune marker genes as a function of these suppressors.  相似文献   

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王伟  唐定中 《植物学报》2021,56(2):142-146
植物先天免疫系统在抵御病原菌入侵过程中发挥至关重要的作用, 主要包括两个层次, 即病原菌相关分子模式和效应因子分别触发的PTI和ETI免疫反应。PTI和ETI分别由植物细胞膜表面模式识别受体(PRRs)和胞内免疫受体(NLRs)激活, 具有特异的激活机制, 但是两者激活的下游免疫事件相互重叠。PTI和ETI是否为泾渭分明的两道防线, 以及ETI与PTI下游事件为何如此相似, 一直是植物免疫领域最受关注的问题之一。最近, 中国科学院分子植物科学卓越创新中心辛秀芳团队与合作者利用拟南芥(Arabidopsis thaliana)与丁香假单胞杆菌(Pseudomonas syringae)互作系统对PTI和ETI在机制上的联系进行了研究。他们发现PRRs和共受体参与ETI, 而活性氧的产生是联系PRRs和NLRs所介导的免疫早期信号事件。他们还发现NLRs信号能够迅速增强PTI关键因子的转录和蛋白水平, PTI的增强在ETI免疫反应中不可或缺。该研究从机制上解析了植物免疫领域中长期悬而未决的PTI与ETI相似性之谜, 是该领域的一项突破性进展, 为未来作物分子设计育种提供了新的启示。  相似文献   

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【背景】西瓜食酸菌(Acidovorax citrulli,Ac)引起的细菌性果斑病是葫芦科植物重要的病害之一,通过Ⅲ型分泌系统(type Ⅲ secreted system,T3SS)分泌至植物体内的Ⅲ型效应蛋白(type Ⅲ effector,T3E)是该菌重要的致病因子,目前对Ac T3E的认识仍然非常有限。【目的】鉴定西瓜食酸菌候选的T3E Acidovorax outer protein AI (AopAI),分析其对Ac致病力的影响和干扰植物免疫的方式。【方法】利用生物信息学方法分析AopAI序列特征、AvrBs1无毒报告系统验证蛋白转运功能;通过荧光定量PCR技术分析aopAI基因表达的调控及其对植物病原相关分子模式(pathogen-associated molecular pattern,PAMP)激发的免疫反应(PAMP-triggered immunity,PTI)信号通路标记基因表达的影响;利用基因插入突变和基因功能互补方法,检测菌的致病力、植物组织过氧化氢和胼胝质积累量的变化;运用瞬时表达技术分析AopAI亚细胞定位和其抑制激发子诱导细胞死亡的能力。【结果】AopAI蛋白序列中不含跨膜螺旋区和信号肽,含有二磷酸腺苷(adenosine diphosphate,ADP)核糖基转移酶保守结构域;在T3SS核心基因hrpGhrpX突变体中aopAI基因表达量显著降低;表达AopAI及AvrBs1功能区(59-445 aa)的avrBs1突变体可诱导ECW-10R辣椒叶发生过敏性坏死反应,表明AopAI具有转运功能;aopAI基因突变体在黄瓜子叶上的致病力减弱,但与其互作的黄瓜子叶组织中过氧化氢和胼胝质的含量均显著增加;AopAI在本氏烟叶瞬时表达后,显示其定位于细胞膜和细胞核,还表现抑制激发子NIP诱导的叶细胞死亡,导致叶细胞的PTI信号通路标记基因GRAS2ACRE31的表达量显著降低。【结论】在西瓜食酸菌中具有一个定位于细胞核和细胞膜、有ADP核糖基转移酶结构域的T3E蛋白AopAI,该T3E是能够抑制NIP诱导的细胞死亡的毒性蛋白,通过抑制ACRE31调节的免疫途径降低植物过氧化氢和胼胝质的积累,以抑制植物PTI防御反应机制。  相似文献   

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EDS1 (Enhanced Disease Susceptibility 1) plays a crucial role in both effector-triggered immunity activation and plant basal defence. However, whether pathogen effectors can target EDS1 or an EDS1-related pathway to manipulate immunity is rarely reported. In this study, we identified a Phytophthora capsici Avirulence Homolog (Avh) RxLR (Arg-any amino acid-Leu-Arg) effector PcAvh103 that interacts with EDS1. We demonstrated that PcAvh103 can facilitate P. capsici infection and is required for pathogen virulence. Furthermore, genetic evidence showed that PcAvh103 contributes to virulence through targeting EDS1. Finally, PcAvh103 specifically interacts with the lipase domain of EDS1 and can promote the disassociation of EDS1–PAD4 (Phytoalexin Deficient 4) complex in planta. Together, our results revealed that the P. capsici RxLR effector PcAvh103 targets host EDS1 to suppress plant immunity, probably through disrupting the EDS1–PAD4 immune signalling pathway.  相似文献   

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In animals, major classes of Rho guanine nucleotide exchange factors (GEFs) possess a Dbl (diffuse B-cell lymphoma)- homology (DH) domain that functions as a GEF-catalytic domain. However, no GEFs with the DH domain had been identified in plants. Recently, we found that the rice homolog of human SWAP70, Oryza sative (Os) SWAP70, containing the DH domain, exhibited GEF activity toward the rice Rho GTPase OsRac1, and regulates chitin-induced production of reactive oxygen species and defense gene expression in rice.1 Arabidopsis contains a single SWAP70 gene. A T-DNA insertion mutant of Arabidopsis SWAP70 was morphologically wild type. Measurement of in planta growth of Pseudomonas syringae DC3000 hrcC, a mutant incapable of type III effector delivery, revealed enhanced growth of the pathogen in the atswap70 mutant, indicating that AtSWAP70 is required for PAMP-triggered immunity. In addition, the atswap70 mutation reduced the RPM1-mediated hypersensitive response. These results suggested that AtSWAP70 plays a role in both PAMP- and effector-triggered immunity in Arabidopsis.  相似文献   

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Diverse pathogen effectors convergently target conserved components in plant immunity guarded by intracellular nucleotide-binding domain leucine-rich repeat receptors (NLRs) and activate effector-triggered immunity (ETI), often causing cell death. Little is known of the differences underlying ETI in different plants triggered by the same effector. In this study, we demonstrated that effector RipAW triggers ETI on Nicotiana benthamiana and Nicotiana tabacum. Both the first 107 amino acids (N1-107) and RipAW E3-ligase activity are required but not sufficient for triggering ETI on Nbenthamiana. However, on Ntabacum, the N1-107 fragment is essential and sufficient for inducing cell death. The first 60 amino acids of the protein are not essential for RipAW-triggered cell death on either Nbenthamiana or N. tabacum. Furthermore, simultaneous mutation of both R75 and R78 disrupts RipAW-triggered ETI on Ntabacum, but not on Nbenthamiana. In addition, Ntabacum recognizes more RipAW orthologs than Nbenthamiana. These data showcase the commonalities and specificities of RipAW-activated ETI in two evolutionally related species, suggesting Nicotiana species have acquired different abilities to perceive RipAW and activate plant defences during plant–pathogen co-evolution.  相似文献   

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With the increasing availability of plant pathogen genomes, secreted proteins that aid infection (effectors) have emerged as key factors that help to govern plant–microbe interactions. The conserved CRN (CRinkling and Necrosis) effector family was first described in oomycetes by their capacity to induce host cell death. Despite recent advances towards the elucidation of CRN virulence functions, the relevance of CRN‐induced cell death remains unclear. In planta over‐expression of PcCRN83_152, a CRN effector from Phytophthora capsici, causes host cell death and boosts P. capsici virulence. We used these features to ask whether PcCRN83_152‐induced cell death is linked to its virulence function. By randomly mutating this effector, we generated PcCRN83_152 variants with no cell death (NCD) phenotypes, which were subsequently tested for activity towards enhanced virulence. We showed that a subset of PcCRN83_152 NCD variants retained their ability to boost P. capsici virulence. Moreover, NCD variants were shown to have a suppressive effect on PcCRN83_152‐mediated cell death. Our work shows that PcCRN83_152‐induced cell death and virulence function can be separated. Moreover, if these findings hold true for other cell death‐inducing CRN effectors, this work, in turn, will provide a framework for studies aimed at unveiling the virulence functions of these effectors.  相似文献   

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Phytophthora infestans is a notorious oomycete pathogen that causes potato late blight. It secretes numerous effector proteins to manipulate host immunity. Understanding mechanisms underlying their host cell manipulation is crucial for developing disease resistance strategies. Here, we report that the conserved RXLR effector Pi05910 of P. infestans is a genotype-specific avirulence elicitor on potato variety Longshu 12 and contributes virulence by suppressing and destabilizing host glycolate oxidase StGOX4. By performing co-immunoprecipitation, yeast-two-hybrid assays, luciferase complementation imaging, bimolecular fluorescence complementation and isothermal titration calorimetry assays, we identified and confirmed potato StGOX4 as a target of Pi05910. Further analysis revealed that StGOX4 and its homologue NbGOX4 are positive immune regulators against P. infestans, as indicated by infection assays on potato and Nicotiana benthamiana overexpressing StGOX4 and TRV-NbGOX4 plants. StGOX4-mediated disease resistance involves enhanced reactive oxygen species accumulation and activated the salicylic acid signalling pathway. Pi05910 binding inhibited enzymatic activity and destabilized StGOX4. Furthermore, mutagenesis analyses indicated that the 25th residue (tyrosine, Y25) of StGOX4 mediates Pi05910 binding and is required for its immune function. Our results revealed that the core RXLR effector of P. infestans Pi05910 suppresses plant immunity by targeting StGOX4, which results in decreased enzymatic activity and protein accumulation, leading to enhanced plant susceptibility.  相似文献   

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RXLR effectors encoded by Phytophthora species play a central role in pathogen–plant interactions. An understanding of the biological functions of RXLR effectors is conducive to the illumination of the pathogenic mechanisms and the development of disease control strategies. However, the virulence function of Phytophthora parasitica RXLR effectors is poorly understood. Here, we describe the identification of a P. parasitica RXLR effector gene, PPTG00121 (PpE4), which is highly transcribed during the early stages of infection. Live cell imaging of P. parasitica transformants expressing a full-length PpE4 (E4FL)-mCherry protein indicated that PpE4 is secreted and accumulates around haustoria during plant infection. Silencing of PpE4 in P. parasitica resulted in significantly reduced virulence on Nicotiana benthamiana. Transient expression of PpE4 in N. benthamiana in turn restored the pathogenicity of the PpE4-silenced lines. Furthermore, the expression of PpE4 in both N. benthamiana and Arabidopsis thaliana consistently enhanced plant susceptibility to P. parasitica. These results indicate that PpE4 contributes to pathogen infection. Finally, heterologous expression experiments showed that PpE4 triggers non-specific cell death in a variety of plants, including tobacco, tomato, potato and A. thaliana. Virus-induced gene silencing assays revealed that PpE4-induced cell death is dependent on HSP90, NPK and SGT1, suggesting that PpE4 is recognized by the plant immune system. In conclusion, PpE4 is an important virulence RXLR effector of P. parasitica and recognized by a wide range of host plants.  相似文献   

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Griffithsin (GRFT) is an antiviral lectin, originally derived from a red alga, which is currently being investigated as a topical microbicide to prevent transmission of human immunodeficiency virus (HIV). Targeting GRFT to the apoplast for production in Nicotiana benthamiana resulted in necrotic symptoms associated with a hypersensitive response (HR)‐like cell death, accompanied by H2O2 generation and increased PR1 expression. Mannose‐binding lectins surfactant protein D (SP‐D), cyanovirin‐N (CV‐N) and human mannose‐binding lectin (hMBL) also induce salicylic acid (SA)‐dependent HR‐like cell death in N. benthamiana, and this effect is mediated by the lectin's glycan binding activity. We found that secreted GRFT interacts with an endogenous glycoprotein, α‐xylosidase (XYL1), which is involved in cell wall organization. The necrotic effect could be mitigated by overexpression of Arabidopsis XYL1, and by co‐expression of SA‐degrading enzyme NahG, providing strategies for enhancing expression of oligomannose‐binding lectins in plants.  相似文献   

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The wide host range of Xylella fastidiosa (Xf) indicates the existence of yet uncharacterized virulence mechanisms that help pathogens to overcome host defences. Various bioinformatics tools combined with prediction of the functions of putative virulence proteins are valuable approaches to study microbial pathogenicity. We collected a number of putative effectors from three Xf strains belonging to different subspecies: Temecula-1 (subsp. fastidiosa), CoDiRO (subsp. pauca), and Ann-1 (subsp. sandyi). We designed an in planta Agrobacterium-based expression system that drives the expressed proteins to the cell apoplast, in order to investigate their ability to activate defence in Nicotiana model plants. Multiple Xf proteins differentially elicited cell death-like phenotypes in different Nicotiana species. These proteins are members of different enzymatic groups: (a) hydrolases/hydrolase inhibitors, (b) serine proteases, and (c) metal transferases. We also classified the Xf proteins according to their sequential and structural similarities via the I-TASSER online tool. Interestingly, we identified similar proteins that were able to differentially elicit cell death in different cultivars of the same species. Our findings provide a basis for further studies on the mechanisms that underlie both defence activation in Xf resistant hosts and pathogen adaptation in susceptible hosts.  相似文献   

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