REGULATION OF DEVELOPMENT IN CAULOBACTER

新月柄杆菌是一种不对称分裂的革兰氏阴性杆菌， 细菌，已被证明是一个很好的模型，为研究细胞 分化 最显著的特征是有柄细胞周期 这个有机体是特定阶段的事件序列，导致 在分裂时产生一个新的能动的蜂群细胞。 我们的长期目标是 来理解控制精确序列的发育程序 并协调它们的时间和细胞定位 表面结构。 我们的研究集中在鞭毛的形成， 需要50多个基因组成一个调控层级 鞭毛基因在细胞周期中的周期性表达是由 一系列反式调节因子。 中提出的工作目标 该应用是表征反式作用因子和DNA 序列元素，它们与之相互作用，以确定观察到的 转录特异性的时间调节。 我们的具体目标 i）确定FlbD与DNA结合的位置和机制 调节σ 54启动子所需的序列元件; ii） 确定FlbD是否是反应调节的功能同源物， 蛋白质NtrC，是通过磷酸化修饰和修饰作用 作为转录因子的功能; iii）鉴定 flaO和flbF操纵子，其与flbD一起沿着被需要用于激活 σ 54启动子在层级的底部; iv）纯化RNA 聚合酶从柄杆菌属，以表征专门的西格玛因素， 以评估它们在决定启动子特异性中的作用; v） 研究DNA合成作为细胞周期信号的作用 鞭毛基因级联;和vi）检查钩蛋白的序列 将其定位到细胞极点，利用翻译融合进行组装， 报告基因

Caulobacter crescentus is an asymmetrically dividing, Gram negative bacterium that has proved to be an excellent model for the study of cell differentiation. The most striking feature of the stalked cell cycle of this organism is the stage specific sequence of events that leads to the generation of a new motile swarmer cell at division. Our long term goal is to understand the developmental program that controls the precise sequence of these events and coordinates their timing and localization of cell surface structures. Our studies have focused on flagellum formation which requires over 50 genes that are organized in a regulatory hierarchy. Periodic expression of the flagellar genes in the cell cycle is mediated by a cascade of trans-acting regulatory factors. The goal of work proposed in this application is to characterize the trans-acting factors and the DNA sequence elements with which they interact to determine the observed temporal regulation of transcriptional specificity. Our specific aims are to i) Determine the location and mechanism of FlbD binding to DNA sequence elements required for regulation of sigma 54 promoters; ii) Determine if FlbD, which is a functional homologue of response regulatory protein NtrC, is modified by phosphorylation and the role of modification for its function as a transcriptional factor; iii) Identify genes in the flaO and flbF operons that are required along with flbD for activation of sigma 54 promoters at the bottom of the hierarchy; iv) Purify RNA polymerases from Caulobacter to characterize specialized sigma factors in order to asses their role in determining promoter specificity; v) Investigate the role of DNA synthesis as a cell cycle signal that triggers the flagellar gene cascade; and vi) Examine the hook protein for sequences that target it to the cell pole for assembly using translation fusions to reporter genes.