CELL INTERACTION-REGULATED GENE EXPRESSION IN M XANTHUS

拟议研究的长期目标是了解如何 细胞-细胞相互作用控制发育基因表达。克- 阴性细菌，黄色粘球菌，提供了一个非常有吸引力的 因为它经历了一个简单的实验系统， 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 试图阐明连接信号传导的分子机制， 基因激活的相互作用。这些研究具有广泛的意义 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， 宿主/病原体和宿主/共生体相互作用。此外还 越来越清楚的是微生物之间的相互作用 许多重要的生物学现象。 至少四种信号相互作用控制粘球菌发育， 其中最具特征的是由csgA介导的，其编码 一种17 Kda的蛋白质，作为C信号分子。一种csgA突变体 不能正常表达许多发育调节的粘球菌 这些基因最初是通过转座子产生的lacZ 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。的 将克隆几个C依赖性基因的调控区， 其特征在于，在体内测试启动子活性，然后进行 启动子区域到缺失分析，转录起始位点 作图和DNA测序。调节蛋白质与 C依赖性基因的调控区将使用 序列特异性DNA结合蛋白的迁移率变动测定， 体外重建C依赖基因转录。监管 蛋白质将被纯化，相应的基因将被克隆， 测序，基因和抗体探针将被开发用于 在旨在了解调节蛋白如何响应 C-信号传导和控制C-依赖性基因表达。

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.