Quorum Sensing in Pseudomonas aeruginosa

描述（由申请人提供）：酰基高丝氨酸内酯（HSL）群体感应发生在多种革兰氏阴性细菌物种中。 在过去的几年里，信号、信号发生器和信号受体的细节已经以快速的速度产生。 群体感应作为基因表达的全局调节器发挥作用，研究人员刚刚开始探索信号传导的这一方面。 铜绿假单胞菌是一个特别有用的模型来研究基因表达的整体群体控制。 基因组已经测序，有一个商业化的基因芯片，并正在创建映射插入突变体库。 已经鉴定了信号和信号基因，并且申请人已经在群体控制的“调节子”中鉴定了超过300个基因。“铜绿假单胞菌群体感应特别相关，因为它是这种新兴人类病原体毒力所必需的。 本项目的目标如下：一、对“早期”群体控制基因的诱导进行精细定位。 该分析将是检验转录激活因子的结合位点的强度定义诱导的精确时间的假设的一个步骤。 二.用发酵剂可控的LasR和RhIR表达模块工程化铜绿假单胞菌菌株，并使用构建体来测试以下假设：在高水平表达下，由LasR或RhIR直接调节的所有基因的时间应该是早的。 在低表达水平下，只有那些对转录因子具有最大亲和力的启动子才应响应信号添加。 三.研究信号受体（转录因子）在体外测试模型的机制，控制复杂的群体基因调控模式。 四.研究一个在稳定期特异表达的群体控制基因。 研究群体感应在生物膜形成中的作用。 铜绿假单胞菌群体感应系统提供了一个机会，了解一个简单的调控元件如何影响细菌培养物中数百个基因的有序表达。 该研究将有助于丰富对群体感应机制和生理意义的理解。

DESCRIPTION (provided by applicant): Acyl-homoserine lactone (HSL) quorum sensing occurs in a wide variety of Gram-negative bacterial species. Details of the signals, signal generators and signal receptors have been generated at a rapid pace in the past several years. Quorum sensing functions as a global regulator of gene expression and the investigators are just beginning to explore this aspect of the signaling. Pseudomonas aeruginosa is a particularly useful model to study global quorum control of gene expression. The genome has been sequenced, there is a commercial GeneChip, and libraries of mapped insertion mutants are being created. The signals and signal genes have been identified, and the applicants have identified over 300 genes in the quorum-controlled "regulon." P. aeruginosa quorum sensing is particularly relevant as it is required for virulence of this emerging human pathogen. The goals of this project are as follows: I. To fine map the induction of "early" quorum-controlled genes. This analysis will be a step in testing the hypothesis that the strength of the binding sites for the transcriptional activators defines the precise timing of induction. II. To engineer P. aeruginosa strains with investigator-controllable LasR and RhlR expression modules and use the constructs to test the hypothesis that at high levels of expression the timing of all genes directly regulated by LasR or RhlR should be early. At low levels of expression only those promoters with the greatest affinity for the transcription factors should respond to signal addition. III. To study the signal receptors (transcription factors) in vitro to test models for mechanisms, which control the complex quorum gene regulatory patterns. IV. To study one particular quorum-controlled gene that is specifically expressed in stationary phase. V. To examine the role of quorum sensing in biofilm development. The P. aeruginosa quorum sensing system provides an opportunity to understand how a simple regulatory element can affect an ordered expression of hundreds of genes in a bacterial culture. The proposed research will serve to enrich the understanding of the mechanisms and physiological significance of quorum sensing.