Mechanisms of Virulence Gene Regulation in Streptococcus mutans

变形链球菌毒力基因调控机制

• 批准号: 7348333
• 负责人: Indranil Biswas
• 金额: $ 39.34万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-02-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7348333
• 关键词: Adherence; Affect; Animal Experiments; Attenuated; Bacterial Infections; Beta-glucuronidase; Binding; Binding Sites; Biochemical; Biochemical Genetics; Biological Assay; Carbohydrates; Cellular Stress Response; Characteristics; Cloning; Country; Cues; DNA; DNA Binding; DNA Microarray Chip; DNA Microarray format; DNA-Binding Proteins; Dental Plaque; Dental caries; Development; EMSA; Electrophoretic Mobility Shift Assay; Elements; Environmental Risk Factor; Family; Gene Expression; Gene Expression Regulation; Genes; Genome; Genomics; Glucans; Glucosyltransferase; Glucosyltransferases; Goals; Helix-Turn-Helix Motifs; Human; In Vitro; Localized; Methods; Microbial Biofilms; Modeling; Modification; Molecular; Mutagenesis; Numbers; Operon; Organism; Orphan; Pathogenesis; Pathway interactions; Phase; Phosphorylation; Phosphotransferases; Play; Printing; Production; Protein Biosynthesis; Proteins; Proteomics; Rattus; Regulation; Regulon; Reporter; Reporter Genes; Reporting; Research Personnel; Role; Sequence Homology; Signal Transduction; Solid; Streptococcus; Streptococcus Group B; Streptococcus mutans; Streptococcus pyogenes; Sucrose; Surface; System; Testing; Therapeutic; Variant; Virulence; Virulence Factors; Water; World Health Organization; base; caN protocol; design; foot; glucan-binding protein; glucosyltransferase D; improved; in vivo; mutant; novel; pathogen; programs; promoter; research study; response; sensor; tooth surface; yeast two hybrid system

Dental caries is one of the most common bacterial infections in humans and remains untreated in many underdeveloped countries. Based on biochemical, epidemiological and animal experiments, Streptococcus mutans is considered to be the principal etiological agent of dental caries. The ability to metabolize carbohydrates and to adhere to and form tenacious biofilms on the tooth surfaces are believed to be critically associated with the cariogenicity of this human pathogen. S. mutans synthesizes glucans from sucrose by three glucosyltransferases (Gtf), and adheres firmly to tooth surfaces with their cooperation. It also produces glucan-binding proteins (Gbps) which play major roles in virulence. The specific mechanisms governing regulation of exopolysaccharide synthesis in S. mutans have yet to be discovered. However, recently it was found that an orphan response regulator, GcrR, modulates the expression of at least one Gtf and one Gbp gene. Remarkably, inactivation of gcrR drastically reduces biofilm formation and cariogenesis in rat. Therefore, GcrR appears to be very important for pathogenesis of this organism. In addition, GcrR shows extensive sequence homology (>80%) with the pathogenic group A streptococcus (GAS) CovR, a response regulator that controls as much as 15% of the GAS genes including many important virulence factors. Given the high degree of similarity with CovR and its effect in biofilm and cariogenesis, one would expect that GcrR may be a global regulator of S. mutans. Therefore, we have chosen to focus on gene regulation by GcrR with the following Specific Aims. In Aim 1, we will identify the gcrR regulon by DNA microarray and in vitro DNA binding assays using purified GcrR protein. We will confirm our results with proteomics approach. In Aim 2, we will determine the mechanisms of gene regulation by GcrR. We will identify and characterize the GcrR binding motif(s) on the promoters of genes (such as gtfD, gbpC) regulated by GcrR. In Aim 3, we will study the regulation of gcrR expression. Since the cognate sensor kinase of GcrR is absent in the nearby gcrR locus, using both biochemical and genetic approaches we will identify the cognate sensor kinase to understand gcrR regulation. This (investigationwill promote our understanding of molecular mechanisms of gene regulation and signal transduction in S. mutans and facilitate the development of therapeutic approaches aimed at controlling formation of plaque biofilm and subsequent cariogenesis.

龋齿是人类最常见的细菌感染之一，并且在许多情况下仍未得到治疗。 不发达国家。根据生物化学、流行病学和动物实验， 变形菌被认为是龋齿的主要病原体。新陈代谢的能力 碳水化合物和粘附并在牙齿表面上形成坚韧的生物膜被认为是至关重要的 与这种人类病原体的致龋性有关。S.变形菌从蔗糖合成葡聚糖， 三种葡糖基转移酶（Gtf），并在它们的合作下牢固地粘附于牙齿表面。它也产生 葡聚糖结合蛋白（Gbps）在毒力中起主要作用。管理的具体机制 调节S.变异人还没有被发现。然而，最近， 发现孤儿反应调节因子GcrR调节至少一种Gtf和一种Gbp的表达， 基因值得注意的是，gcrR的失活大大减少了大鼠中的生物膜形成和龋齿发生。 因此，GcrR似乎对该生物体的发病机制非常重要。此外，GcrR显示， 与致病性A组链球菌（GAS）CovR具有广泛的序列同源性（>80%）， 控制多达15%的GAS基因的调节因子，包括许多重要的毒力因子。给定 与CovR的高度相似性及其在生物膜和龋齿发生中的作用，可以预期GcrR 可能是S.变异人因此，我们选择专注于GcrR的基因调控 有以下具体目标。目的一：利用基因芯片技术和体外培养技术对gcrR调节子进行鉴定 使用纯化的GcrR蛋白的DNA结合测定。我们将用蛋白质组学方法证实我们的结果。在 目的2，我们将确定GcrR的基因调节机制。我们将识别和描述 受GcrR调节的基因（例如gtfD、gbpC）启动子上的GcrR结合基序。在目标3中，我们 研究gcrR的表达调控。由于GcrR的同源传感器激酶在附近的细胞中不存在， gcrR基因座，使用生物化学和遗传方法，我们将确定同源传感器激酶， 了解gcrR法规。 这一研究将促进我们对基因调控和信号传导的分子机制的理解 S.并促进治疗方法的发展，旨在控制 牙菌斑生物膜的形成和随后的龋齿发生。