Regulation of Streptococcus gordonii glucosyltransferase

描述：哥氏葡萄球菌葡萄糖基转移酶（GTF）酶合成葡聚糖，可以影响发育中的牙菌斑生物膜中细菌的积累。正在进行的项目（DE 11090）的工作假设是，godonii中GTF活性的调节涉及复杂的分子相互作用，可能对细菌在口腔中的定植和存活具有生态意义。这个正在进行的项目的目标是研究环境信号和分子机制，调控葡萄糖基转移酶的表达，在共生物种，s.o donii。本补充申请中描述的拟议研究的具体目的是使用DNA微阵列识别和表征与GTF结构基因gtfG及其正调控决定因素rgg表达相关的协调调节遗传网络。亲本菌株的消失表达谱将与与GTF活性相关的基因（如rgg和gtfG）被特异性灭活的菌株进行比较。本实验还将检测这些菌株在特定环境条件下的基因表达，这些环境条件会影响GTF活性水平。微阵列方法学将允许一个基本上全基因组，高度敏感的搜索协调表达的基因。获得的结果将使用定量实时聚合酶链反应（PCR）进行验证。这些拟议研究的微阵列的构建将通过使用现有的DNA克隆集来促进，这些克隆集来自于gordonii测序项目中使用的亲本菌株Challis；这些分子工具在最初的父母资助申请时是不可用的。对于本补充申请，微阵列实验将集中于解决母基金的工作假设。拟议补充研究的结果将专门用于加速和补充正在进行的遗传和功能研究。这些研究旨在揭示葡萄球菌GTF表达调控的复杂机制，并阐明合成的葡聚糖对牙菌斑生物学的贡献。

DESCRIPTION: The S. gordonii glucosyltransferase (GTF) enzyme synthesizes glucans that can influence the accumulation of bacteria in the developing Dental plaque biofilm. The working hypothesis of the ongoing project (DE 11090) is that regulation of GTF activity in S. gordonii involves complex molecular interactor(s) that may have ecological implications for bacterial colonization and survival in the oral cavity. The goal of this ongoing project is to investigate the environmental signals and molecular mechanisms that regulate glucosyltransferase expression in the commensal species, S. gordonii. The Specific Aim of the proposed studies described in this Supplemental Application, is to use DNA microarrays to identify and characterize coordinately regulated genetic networks associated with expression of the GTF structural gene, gtfG and its positive regulatory determinant, rgg. Gone expression profiles of the parent strain will be compared to strains in which genes involved in GTF activity (e.g. rgg and gtfG) have been specifically inactivated. The proposed experiments will also examine gene expression in these bacterial strains under specific environmental conditions, which influence the level of GTF activity. Microarray methodology will allow an essentially genome-wide, highly sensitive search for coordinately expressed genes. The results obtained will be validated using quantitative real-time polymerase chain reaction (PCR). Construction of microarrays for these proposed studies will be facilitated by the use of existing DNA clone sets available from the parental strain Challis used in the S. gordonii sequencing project; these molecular tools were not available at the time of the original parent grant application. For this Supplemental Application, microarray experiments will be focused on addressing the working hypothesis of the parent grant. The results of the proposed supplemental studies will be used specifically to accelerate and supplement ongoing genetic and functional studies in the parent grant. These studies are designed to unravel the complex mechanisms involved in regulation of GTF expression in S. gordonii and elucidate the contributions of the synthesized glucans to the biology of Dental plaque.