Regulation and Mechanism of M. xanthus Social Gliding

黄花鮟鱇群居性滑翔的调控与机制

基本信息

批准号：
7943643
负责人：
ZHAOMIN YANG
金额：
$ 14.27万
依托单位：
VIRGINIA POLYTECHNIC INST AND ST UNIV
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-30 至 2011-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7943643
关键词：
Address Affinity Affinity Chromatography Amino Acids Archaea Bacteria Bacterial Infections Bacterial Typing Binding Proteins Biochemical Biogenesis Cell surface Cells Chemoreceptors Chemotaxis Chronic Cytoplasmic Tail Defect Dependence Developmental Process Escherichia coli Exhibits Extracellular Matrix Fruit Genes Genetic Screening Goals Homologous Gene Human Immunoblotting Immunofluorescence Microscopy Lead Ligand Binding Mediating Methylation Microbial Biofilms Modeling Modification Mutation Myxococcus xanthus Nitrates Output Pathogenicity Pathway interactions Phosphorylation Pilum Production Property Proteins Regulation Sensory Signal Pathway Signal Transduction Structure Surface System Tars Tertiary Protein Structure Testing Virulence Yeasts cell motility design extracellular mutant novel pathogen periplasm pill response social yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): Type IV pill (Tfp) are important determinants of bacterial virulence and biofilm formation which is the cause of many opportunistic and chronic bacterial infections. Tfp also mediate a form of bacterial surface motility known as social (S) gliding in Myxococcus xanthus and twitching in other bacterial species. The function of Tfp in surface motility correlates with their function in bacterial pathogenicity and biofilm formation. Mutations in genes essential for Tfp biogenesis and function lead to simultaneous defects in bacterial virulence, biofilm formation and Tfp-mediated motility. Our long term goal is to use M. xanthus S-motility as a model to study Tfp-mediated functions and signaling. S-motility itself also warrants further studies on its own merit because it is important to the developmental process of M. xanthus multicellular fruiting bodies. Besides Tfp, M. xanthus S-motility requires another cell surface component known as extracellular fibrils. Previous studies showed that the dif genes, encoding homologues of bacterial chemotaxis proteins, are central to the biogenesis or production of fibrils. Recent findings suggest that the pil genes which are required for Tfp biogenesis and function are also involved in the regulation of fibril biogenesis in M. xanthus. We propose to do the following. First, we will verify the requirement of fibril biogenesis for Tfp. We will further construct double mutants to examine the epistatic relationships in the regulation of fibril biogenesis among dif and pil genes as predicted by our hypothesis. Second, we will use yeast two-hybrid (Y2H) system and phosphorylation studies to examine if the Dif chemosensory-like proteins interact with one another physically and biochemically as their counterparts in bacterial chemotaxis and to identify any novel interactions that may exist among Dif. Third, it is unknown whether DifA, a homologue of methyl-accepting chemoreceptor proteins (MCP), localizes to specific subcellular regions and how DifA and the Dif pathway respond to stimulation. We will study DifA localization and the dependence of correct DifA localization on dif and pil genes by immunofluorescence microscopy. We will additionally construct and use NarX-DifA fusions to examine DifA modification and the signaling properties of the Dif pathway in response to stimulation by nitrate. Finally, Y2H and genetic screens will be carried out to identify new genes important for fibril regulation and biogenesis, especially downstream of Dif proteins.

描述（由申请人提供）：IV 型药丸（Tfp）是细菌毒力和生物膜形成的重要决定因素，而生物膜形成是许多机会性和慢性细菌感染的原因。 Tfp 还介导一种细菌表面运动，在黄色粘球菌中被称为社交 (S) 滑动，在其他细菌物种中被称为抽搐。 Tfp 在表面运动中的功能与其在细菌致病性和生物膜形成中的功能相关。 Tfp 生物发生和功能所必需的基因突变导致细菌毒力、生物膜形成和 Tfp 介导的运动同时缺陷。我们的长期目标是使用 M. xanthus S 运动作为模型来研究 Tfp 介导的功能和信号传导。 S-运动本身也值得进一步研究，因为它对 M. xanthus 多细胞子实体的发育过程很重要。除了 Tfp 之外，M. xanthus S 运动还需要另一种称为细胞外原纤维的细胞表面成分。先前的研究表明，编码细菌趋化蛋白同源物的dif基因对于原纤维的生物发生或产生至关重要。最近的研究结果表明，Tfp 生物发生和功能所需的 pil 基因也参与 M. xanthus 中原纤维生物发生的调节。我们建议采取以下措施。首先，我们将验证原纤维生物发生对 Tfp 的要求。我们将进一步构建双突变体来检查 dif 和 pil 基因之间原纤维生物发生调节的上位关系，正如我们的假设所预测的那样。其次，我们将使用酵母双杂交 (Y2H) 系统和磷酸化研究来检查 Dif 化学感应样蛋白是否与细菌趋化性中的对应蛋白一样在物理和生化方面相互作用，并确定 Dif 之间可能存在的任何新的相互作用。第三，尚不清楚 DifA（甲基接受化学感受器蛋白 (MCP) 的同源物）是否定位于特定的亚细胞区域以及 DifA 和 Dif 通路如何响应刺激。我们将通过免疫荧光显微镜研究 DifA 定位以及正确的 DifA 定位对 diff 和 pil 基因的依赖性。我们还将构建并使用 NarX-DifA 融合体来检查 DifA 修饰和 Dif 途径响应硝酸盐刺激的信号传导特性。最后，将进行 Y2H 和遗传筛选，以确定对原纤维调节和生物发生重要的新基因，特别是 Dif 蛋白的下游。