Novel Multichannel Environmental Monitoring Network Controlling Virulence of Ralstonia solanacearum

控制青枯雷尔斯顿菌毒力的新型多通道环境监测网络

• 批准号: 9727921
• 负责人: Mark Schell
• 金额: $ 30.03万
• 依托单位: University of Georgia Research Foundation Inc
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-01 至 2003-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9727921&HistoricalAwards=false
• 关键词: Novel; Multichannel; Environmental; Monitoring; Network

Ralstonia (Pseudomonas) solanacearum causes a lethal wilting disease of over 200 different plants worldwide. Secreted plant cell-wall-degrading exoenzymes and the EPS I exopolysaccharide, both contribute to its successful colonization of a plant's vascular system after root invasion. Because EPS I blocks water flow in the xylem, it is also a primary cause of wilting and killing. Biosynthesis and export of EPS I, a long unbranched polymer of 3 amino sugars, are encoded by the 16-kb eps operon. Transcription of the eps operon, as well as genes encoding some exoenzymes and other virulence genes, is controlled by a large, interactive regulatory network of over 12 proteins that is responsive to multiple environmental signals. This network is comprised of two distinct two-component regulatory systems (VsrAD and VsrBC), the unique signal integrator protein XpsR and the unusual Phc signal transduction system. Preliminary data suggest that the Phc module employs an atypical phosphorelay cascade that responds to 3-hydroxypalmitic acid methyl ester (3-OH PAME), a new type of volatile, quorum-indicating molecule. Phc and 3-OH PAME in turn regulate phcA, encoding a global, LysR-type transcriptional regulator that controls reversible switching between two very different physiological states, one adapted for virulence in plants, the other for saprophytic survival. A major goal of this project is to understand at the molecular level how the Phc module responds to 3-OH PAME, and how the Phc components subsequently control phcA function. Using two different approaches, a genetic one (transposon mutagenesis) or a biochemical one (DNA-affinity chromatography of R. solanacearum extracts on columns with the eps regulatory region covalently bound), missing genes that mediate regulation of eps by XpsR and VsrC), will be searched for, cloned, and characterized. These studies of the R. solanacearum virulence regulatory network are important because one of the most critical factors for successful pathogenesis is the ability to coordinate production of virulence and pathogenicity factors in response to environmental signals, and because the ability of the R. solanacearum network to process and integrate multiple signal inputs is largely unparalleled. These studies should give insight into the ways multiple environmental cues are processed by pathogens to adjust virulence gene expression. Finally, genes required by R. solanacearum for rapid, efficient colonization of plants by cloning and analysis of genes encoding several vsrD-regulated exoproteins will be searched for. In parallel, in vivo expression technology will be adapted and applied to R. solanacearum. This technology identifies and isolates genes that are highly expressed. by a pathogen only during colonization of a host. Either or both of these approaches will provide a way to access new, important genes, and provide insight into a pathogen's strategies for successful colonization of plants.

青枯雷尔氏菌（Ralstonia）（假单胞菌）引起全世界200多种不同植物的致死性枯萎病。分泌的植物细胞壁降解胞外酶和EPS I胞外多糖都有助于其在根侵入后成功地定殖于植物的维管系统。因为EPS I阻断木质部中的水流，所以它也是萎蔫和死亡的主要原因。EPS I是3个氨基糖的长链无支链聚合物，其生物合成和输出由16-kb的eps操纵子编码。eps操纵子的转录，以及编码一些外切酶和其他毒力基因的基因，是由超过12种蛋白质组成的一个大型的、相互作用的调控网络控制的，该网络对多种环境信号做出响应。该网络由两个不同的双组分调节系统（VsrAD和VsrBC）、独特的信号整合蛋白XpsR和不寻常的Phc信号转导系统组成。初步数据表明，Phc模块采用非典型的磷酸化级联反应，响应3-羟基棕榈酸甲酯（3-OH PAME），一种新型的挥发性，群体指示分子。Phc和3-OH PAME反过来调节phcA，编码一种全局的LysR型转录调节因子，该转录调节因子控制两种非常不同的生理状态之间的可逆切换，一种适应植物中的毒力，另一种适应荒漠化生存。 该项目的一个主要目标是在分子水平上了解Phc模块如何响应3-OH PAME，以及Phc组件如何随后控制phcA功能。利用两种不同的方法，一种是遗传方法（转座子诱变），另一种是生化方法（R.将搜索、克隆和表征在具有共价结合的eps调节区的柱上的青枯菌提取物），缺失通过XpsR和Vsrc介导eps调节的基因）。本文对R.青枯菌毒力调控网络是重要的，因为成功致病的最关键因素之一是能够协调毒力因子和致病因子的产生以响应环境信号，并且因为青枯菌毒力调控网络的能力是重要的。青枯菌网络处理和整合多个信号输入在很大程度上是无与伦比的。这些研究应该让我们深入了解病原体处理多种环境线索以调节毒力基因表达的方式。 最后，研究了R.将通过克隆和分析编码几种vsrD调节的外蛋白的基因来寻找青枯菌用于快速、有效地定殖植物的方法。与此同时，体内表达技术将被改造并应用于R。青枯菌属该技术鉴定并分离高度表达的基因。只有在宿主定植期间才能被病原体感染。这两种方法中的一种或两种都将提供一种获得新的重要基因的方法，并提供对病原体成功定殖植物的策略的深入了解。