Genetic and biochemical studies of competence regulation by the hdrRM operon

描述(申请人提供)：拟议的研究旨在研究新发现的高细胞密度响应性hdrRM操纵子对变形链球菌能力调控的遗传和生化机制。这个项目的目标将通过两个目标来实现：1)确定受hdrrm调控的能力级联的步骤；2)确定hdrrm调控基因的机制。1)为了分析hdrRM操纵子如何与能力级联基因相互作用，将进行转录、上位性和突变分析。这些研究将为hdrrm作为能力负调节因子的作用提供遗传学基础。2)为了研究HdrR作为能力调节因子的作用机制，我们将HdrR用于能力基因启动子的DNA结合分析。此外，HdrR和HDRM之间的体内蛋白质相互作用研究将表征这两种蛋白质之间可能的相互作用。这些数据有望为目标1中产生的遗传和转录数据提供生化证据，并确定HdrRM系统中的信号转导机制。该项目的目标是将能力调控作为hdrRM操纵子能够调控变形链球菌其他重要毒力因子的机制的模型。由于该操纵子受高细胞密度条件的强烈诱导，预计它将在牙菌斑的高细胞密度环境中对生存和致病起关键作用。 项目简介：龋齿是人类最常见的细菌疾病，与变形链球菌的过度生长密切相关。由于它的致病能力要求它在牙菌斑的高细胞密度条件下发挥作用，因此对在高细胞密度条件下发挥作用的基因调控因子的研究可能会产生新的治疗策略来控制变形链球菌的发病。

DESCRIPTION (provided by applicant): The proposed research Aims to investigate the genetic and biochemical mechanisms of competence regulation in Streptococcus mutans by the newly discovered high cell density responsive hdrRM operon. The goals of this project will be achieved in two Aims by: 1) Determining the steps of the competence cascade that are regulated by hdrRM 2) Determining the mechanism of gene regulation by HdrRM. 1) To analyze how the hdrRM operon interacts with the genes of the competence cascade, a combination of transcription, epistatic, and mutation analysis will be performed. These studies will provide the genetic basis for the role of hdrRM as a negative regulator of competence. 2) To characterize the mechanism of HdrR as a regulator of competence, HdrR will be used for DNA binding assays of competence gene promoters. In addition, in vivo protein interaction studies between HdrR and HdrM will characterize possible interactions between the two proteins. These data are expected to provide biochemical evidence for the genetic and transcriptional data generated in Aim 1 as well as determine the mechanism of signal transduction in the HdrRM system. The goal of this project is to use competence regulation as a model for the mechanism by which the hdrRM operon is able to regulate other important virulence factors of Streptococcus mutans. Since this operon is strongly induced by conditions of high cell density, it is expected that it will play a crucial role for both survival and pathogenicity in the high cell density environment of the dental plaque. Project Narrative: Dental caries is the most common human bacterial disease and is largely correlated with the overgrowth of Streptococcus mutans. Since its ability to cause disease requires it to function in the high cell density conditions of dental plaque, studies of gene regulators that function in high cell density conditions may yield novel treatment strategies to control the pathogenesis of S. mutans.