Contact-Dependent Signaling in Burkholderia

伯克霍尔德杆菌中的接触依赖性信号传导

• 批准号: 9321389
• 负责人: Peggy A Cotter
• 金额: $ 29.57万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9321389
• 关键词: Alleles; Animals; Antibiotics; Bacteria; Behavior; Binding; Binding Proteins; Biochemical; Burkholderia; C-terminal; Cell Death; Cells; Cleaved cell; Communities; Competitive Behavior; Complex; Contact Inhibition; Control Locus; Cooperative Behavior; Cytoplasm; DNA; Data; Development; Disease; Environment; Enzymes; Gene Expression; Genes; Genetic; Genetic Epistasis; Gram-Negative Bacteria; Growth; Human; Immunity; Immunoprecipitation; In Vitro; Insecta; Mass Spectrum Analysis; Measures; Mediating; Medical; Medical Device; Membrane; Microbial Biofilms; Microbiology; Modeling; Molecular; Mutagenesis; Nucleotides; Plant Roots; Production; Proteins; Proteobacteria; RNA; Reaction; Regulon; Role; Second Messenger Systems; Shellfish; Ships; Signal Transduction; Signaling Molecule; Signaling Protein; Site-Directed Mutagenesis; Structure; Surface; System; Therapeutic; Transducers; base; experimental study; hookah; killings; member; mutant; nuclease; pathogenic bacteria; polypeptide; prevent; reconstitution; transcriptome sequencing; transmission process

Abstract: Bacteria typically live in complex sociomicrobiological communities, often as biofilms, on surfaces as diverse as water pipes, ship hulls, plant roots, insects, shellfish, indwelling medical devices, and mucosal surfaces. Biofilm growth in humans can cause or exacerbate disease, and biofilm growth in environmental niches can facilitate transmission of pathogenic bacteria to humans and other animals. Understanding how bacteria recognize, cooperate and compete with their neighbors in diverse environments is critical for developing strategies to control microbiological community composition, to prevent biofilm development, and to eliminate pre-existing biofilms and their consequent diseases. Contact-Dependent Growth Inhibition (CDI) is a phenomenon in which bacteria use the toxic C-terminus of a large exoprotein to kill or inhibit the growth of neighboring bacteria upon cell-cell contact. Production of a small immunity protein protects bacteria against CDI. Using the Gram-negative bacterium Burkholderia thailandensis as a model, we have discovered that in addition to using CDI system proteins to kill their neighbors, bacteria can use these proteins for signal transduction, causing a change in gene expression that leads to the production of cooperative behaviors, such as biofilm formation, when neighboring bacteria are recognized as ‘self’. Here, we propose experiments to determine the molecular mechanisms underlying this Contact-Dependent Signaling (CDS) phenomenon. Our preliminary data support a model in which the C-terminus of the large BcpA exoprotein is delivered into neighboring bacteria where it forms a ‘CDS complex’ that catalyzes a reaction that increases or decreases the concentration of a signal molecule, which may be a small nucleotide second messenger. Experiments described in Aim 1 will define the CDS regulatory network and identify the signal molecule, which will help us identify the enzymatic activity of the CDS complex. Experiments described in Aim 2 will identify the members of the CDS complex. We will reconstitute the CDS complex in vitro using purified proteins and will measure its catalytic activity. We have discovered that a small protein called BcpO is required for CDS. We proposed experiments in Aim 3 to determine if BcpO localizes to the inner leaflet of the outer membrane, as predicted, and if that localization is important for its function. We will also identify proteins with which BcpO interacts so that we can understand how it functions in CDS.

抽象的： 细菌通常生活在复杂的社会生物学群落中，通常为生物膜，在表面上 像水管一样潜水，船体，植物根，昆虫，贝类，留置医疗设备以及 粘膜表面。人类生物膜的生长会引起或加剧疾病，生物膜生长 在环境中，生态位可以促进致病细菌向人类和其他 动物。了解细菌如何认识，教练和与邻居竞争 潜水员环境对于制定控制微生物社区的策略至关重要 组成，防止生物膜发育，并消除先前存在的生物膜及其 随之而来的疾病。接触依赖性生长抑制（CDI）是一种现象，其中 细菌使用大一色蛋白的有毒C末端杀死或抑制相邻的生长 细菌接触时细菌。产生小的免疫蛋白可保护细菌免受CDI的影响。 将革兰氏阴性细菌Burkholderia tailandensis作为模型，我们发现了 除了使用CDI系统蛋白质杀死邻居外，细菌还可以使用这些蛋白质 对于信号转导，导致基因表达的变化，导致产生 当相邻细菌被认为是生物膜形成之类的合作行为，例如 '自己'。在这里，我们提出了实验，以确定这一点的分子机制 接触依赖性信号（CD）现象。我们的初步数据支持一个模型 大型BCPA代蛋白的C末端被递送到邻近的细菌中 “ CDS络合物”催化反应增加或降低信号的浓度 分子，这可能是小型核苷酸第二信使。 AIM 1中描述的实验 将定义CDS调节网络并确定信号分子，这将帮助我们识别 CDS复合物的酶活性。 AIM 2中描述的实验将确定 CDS综合体的成员。我们将使用纯化的蛋白质在体外重建CDS络合物 并将测量其催化活性。我们发现一种称为BCPO的小蛋白质是 CD所需。我们在AIM 3中提出了实验，以确定BCPO是否本地化到内部 如所预测的，外膜的传单，以及该定位对于其功能很重要。我们 还将识别BCPO相互作用的蛋白质，以便我们可以理解其在 CD。