CELL INTERACTION-REGULATED GENE EXPRESSION IN M XANTHUS

M Xanthus 中细胞相互作用调控的基因表达

• 批准号: 2184703
• 负责人: LEE R KROOS
• 金额: $ 12.07万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-01 至 1996-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2184703
• 关键词: DNA binding protein; Myxococcus; bacterial genetics; beta galactosidase; biological signal transduction; cell cell interaction; developmental genetics; gene deletion mutation; gene expression; genetic manipulation; genetic mapping; genetic promoter element; genetic transcription; genetic transduction; molecular cloning; mutant; nucleic acid probes; nucleic acid sequence; regulatory gene; southern blotting

The long-term objective of the proposed research is to understand how cell-cell interactions control developmental gene expression. The gram- negative bacterium, Myxococcus xanthus, provides a very attractive experimental system for these studies because it undergoes a simple multicellular development requiring cell-to-cell signaling, and it is amenable to both biochemical and genetic analyses. The proposed studies seek to elucidate the molecular mechanism(s) that connects a signaling interaction to gene activation. These studies are of broad significance because cell-to-cell communication controls both organismal development and many processes within mature multicellular organisms, as well as host/pathogen and host/symbiont interactions. In addition, it is becoming increasingly clear that interactions between microbes influence many important biological phenomena. At least four signaling interactions control Myxococcus development and the best-characterized of these is that mediated by csgA, which encodes a 17 Kda protein that serves as the C-signal molecule. A csgA mutant fails to normally express many developmentally regulated Myxococcus genes, which were originally identified by transposon-generated lacZ fusions. These C-dependent genes provide a starting point for the proposed investigation which is aimed at exploring the molecular mechanism(s) that connects C-signaling to gene activation. The regulatory regions of several C-dependent genes will be cloned and characterized by testing for promoter activity in vivo, then subjecting promoter regions to deletion analyses, transcriptional start site mapping, and DNA sequencing. Regulatory proteins that interact wit the regulatory regions of C-dependent genes will be identified using a mobility shift assay for sequence-specific DNA-binding proteins and by reconstructing C-dependent gene transcription in vitro. The regulatory proteins will be purified, the corresponding genes will be cloned and sequenced, and both gene and antibody probes will be developed for use in studies aimed at understanding how the regulatory proteins respond to C-signaling and control C-dependent gene expression.

拟议研究的长期目标是了解如何 细胞间相互作用控制发育基因表达。克- 阴性细菌，黄色粘球菌，提供了非常有吸引力的 这些研究的实验系统，因为它经历了一个简单的 多细胞发育需要细胞间信号传导， 适合生化和遗传分析。拟议的研究 寻求阐明连接信号传导的分子机制 与基因激活的相互作用。这些研究具有广泛意义 因为细胞间的通讯控制着生物体的发育 以及成熟多细胞生物体内的许多过程，以及 宿主/病原体和宿主/共生体相互作用。此外，它是 越来越清楚的是，微生物之间的相互作用会影响 许多重要的生物现象。 至少有四种信号相互作用控制粘球菌的发育和 其中最具特征的是由 csgA 介导，它编码 一种 17 Kda 的蛋白质，充当 C 信号分子。 csgA突变体 无法正常表达许多发育调节的粘球菌 最初由转座子产生的 lacZ 识别的基因 融合。这些C依赖性基因提供了一个起点 拟议的研究旨在探索分子 将 C 信号传导与基因激活联系起来的机制。这 几个C依赖性基因的调控区域将被克隆并 其特征是测试体内启动子活性，然后进行 启动子区域缺失分析、转录起始位点 作图和 DNA 测序。与相互作用的调节蛋白 C依赖性基因的调控区域将使用 序列特异性 DNA 结合蛋白的迁移率变化分析 体外重建 C 依赖性基因转录。监管 蛋白质将被纯化，相应的基因将被克隆， 测序，并将开发基因和抗体探针供使用 在旨在了解调节蛋白如何响应的研究中 C 信号传导和控制 C 依赖性基因表达。