Chemotactic Cues in the Symbiosis Between Vibrio fischeri and Euprymna scolopes

费氏弧菌和圆角藻共生中的趋化信号

• 批准号: 8776534
• 负责人: Kiel Nikolakakis
• 金额: $ 5.15万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8776534
• 关键词: Address; Agar; Animal Model; Animal Organ; Area; Bacteria; Behavior; Biological Assay; Biological Models; Blood capillaries; Cells; Chemotaxis; Complex; Cues; Data; Environment; Escherichia coli; Event; Fluorescence Resonance Energy Transfer; Generations; Genes; Goals; Hawaiian population; Health; Human; Individual; Infection; Investigation; Life; Ligands; Light; Methods; Microbe; Microscopy; Migration Assay; Modeling; Nutrient; Organ; Organism; Output; Pathogenesis; Pathway interactions; Postdoctoral Fellow; Process; Proteins; Recruitment Activity; Regulation; Reporter; Research; Role; Seawater; Signal Pathway; Signal Transduction; Squid; Staging; Symbiosis; System; Techniques; TimeLine; Tissues; Training; Tube; Vibrio; Vibrio fischeri; bacterioplankton; base; capillary; cell motility; design; environmental change; extracellular; insight; interest; methyl-accepting chemotaxis proteins; migration; novel; public health relevance; receptor; research study; response; small molecule; success; tool

DESCRIPTION (provided by applicant): Host-microbe interactions, both beneficial and pathogenic, are a rapidly developing area of research with important implications for understanding the normal microbiota, as well as its disturbance by infection. The monospecific symbiosis between the bioluminescent bacteria Vibrio fischeri and the Hawaiian bobtail squid, Euprymna scolopes, provides us with an ideal model with which to study the earliest events of symbiotic initiation. This proposal will focus on a key bacterial signaling pathway, chemotaxis, which allows a bacterium to recognize and respond to changes in its environment. Characterization of bacterial chemotaxis in the context of a beneficial host-microbe association will provide a more complete picture of how microbes interact with and respond to the complex environments presented by host tissues, while migrating from the ambient seawater into a host organism. The overall goals of this research are to characterize the methyl-accepting chemotaxis proteins (MCPs) which act as the ligand receptors in the bacterial chemotaxis machinery, as well as the regulation of these receptors. MCPs have been well studied to date in E. coli. However, E. coli only uses a set of five of these receptors, while the majority of bacteri are known to have many more. V. fischeri is predicted to encode 43 MCPs, and their expression is known to be relevant to colonization of the host. The colonization of E. scolopes by V. fischeri has been shown to occur in several distinct stages, and involves the bacterium migrating through several environments and tissues as it moves from the bacterioplankton to the interior light organ of the animal. This research will further clarify the specific ligands recognized by V. fischeri, and which of these are involved during the initiation of symbiosis. The first aim will address the receptor-ligand interactions, and will be pursued using established techniques such as soft-agar motility assays and capillary-tube migration assays, as well as a FRET-based assay that allows investigation of specific ligand-protein interactions in a receptorless- background strain of E. coli. The second aim will investigate the regulation of these MCPs, and will utilize both transcriptional profiling by RNAseq and fluorescent reporter methods, in experiments designed to determine whether distinct stages of the colonization involve specific receptors. The tools used and developed during the course of this project will also be applicable to other studies, both in the Ruby lab and in other labs studying host-microbe interactions. This project will also provide significant training to the post-doctoral fellow in several areas, includng advanced microscopy and transcriptional profiling techniques.

描述(申请人提供)：宿主-微生物相互作用，无论是有益的还是致病的，都是一个快速发展的研究领域，对于理解正常的微生物区系以及感染对其干扰具有重要意义。生物发光细菌费氏弧菌和夏威夷短尾乌贼之间的单种共生为我们研究共生启动的最早事件提供了一个理想的模型。这项提案将重点放在一个关键的细菌信号通路--趋化作用上，它使细菌能够识别并响应环境的变化。在有益的宿主-微生物联合的背景下表征细菌的趋化性将提供一个更完整的图景，说明微生物在从周围海水迁移到宿主生物体时如何与宿主组织呈现的复杂环境相互作用并做出反应。本研究的总体目标是研究在细菌趋化机制中作为配体受体的甲基接受趋化蛋白(MCPs)，以及这些受体的调节。到目前为止，在大肠杆菌中已经对MCPS进行了很好的研究。然而，大肠杆菌只使用其中的五种受体，而大多数细菌已知有更多的受体。费氏弧菌编码43个MCP，它们的表达与寄主的定殖有关。费氏拟青霉对E.scolope的定植 已被证明发生在几个不同的阶段，涉及细菌在从浮游细菌移动到动物内部光器官的过程中通过几个环境和组织迁移。本研究将进一步阐明V。 它们中的哪一种参与了共生的启动。第一个目标将解决受体-配体相互作用，并将使用现有的技术，如软琼脂运动分析和毛细管迁移分析，以及基于FRET的分析，允许研究无受体-背景菌株的特定配体-蛋白质相互作用。第二个目标将调查这些MCP的调节，并将利用RNAseq转录图谱和荧光报告方法，在设计的实验中确定不同的定植阶段是否涉及特定的受体。在这个项目过程中使用和开发的工具也将适用于其他研究，无论是Ruby实验室还是其他研究宿主-微生物相互作用的实验室。该项目还将在几个领域为博士后研究员提供重要的培训，包括先进的显微镜和转录图谱技术。