Cloning/Characterization of S. mutans Iron Stimulation

描述(由申请人提供)：铁是细菌病原体及其哺乳动物宿主所必需的微量营养素，因此对该元素的竞争是感染过程中不可或缺的组成部分。细胞外的亚铁在生物上对入侵的病原体是不可用的，因为它在很大程度上隔离在宿主蛋白质中，并在其自由状态下迅速氧化成不溶的铁状态。细菌通常通过启动铁清除系统和毒力决定因素来响应宿主的铁保持系统，以促进细菌的生存和传播。在革兰氏阳性菌中研究最广泛的金属离子依赖调节因子是白喉棒杆菌中的DNA结合蛋白DtxR，以及它在表皮葡萄球菌中的系统发育同源物SirR和金黄色葡萄球菌中的MntR。重要的是，这些金属调节因子发挥转录因子的作用，调节细菌毒力基因和与毒力和氧化应激相关的高亲和力金属离子吸收系统的表达。我们实验室对变形链球菌进行的研究揭示了一种DtxR同源基因，它调控着一系列基因，并且积累的证据与铁在变形链球菌生物膜形成、氧化应激反应和耐酸性中的作用是一致的。变形链球菌是人类牙菌斑的主要产酸成分。事实上，识别和表征参与这些和其他致病过程的基因是一个高度优先的问题，因为它们可能被证明是未来的药物靶标。这项研究计划的主要目标将集中在阐明铁依赖的基因调控及其在变形链球菌诱导的龋病发生中的作用。其具体目标包括：1)鉴定可能属于变形链球菌DLG调节子的铁反应基因；2)克隆和鉴定变形链球菌中其他不受DLG调控的铁依赖基因；3)检测变形链球菌DLG调控基因和铁反应基因在接近人类口腔的环境中的表达；4)建立DLG/铁调控基因表达与变形链球菌诱导的龋病之间的关系。

DESCRIPTION (provided by applicant): Iron is an essential micronutrient for bacterial pathogens and their mammalian hosts, and hence competition for this element is an integral component of the infectious process. Extracellular ferrous iron is biologically unavailable to invading pathogens because it is largely sequestered to host proteins, and in its free form rapidly oxidizes to the insoluble ferric state. Bacteria often respond to the iron withholding system of the host by turning on iron scavenging systems and virulence determinants to promote bacterial survival and dissemination. Among the most extensively studied metal ion-dependent regulators in Gram-positive bacteria is DtxR, a DNA-binding protein, in Corynebacterium diphtheriae, and its phylogenetic homologs SirR in Staphylococcus epidermidis and MntR in Staphylococcus aureus. Importantly, these metalloregulators function as transcription factors that modulate the expression of bacterial virulence genes and high affinity metal ion uptake systems related to virulence and oxidative stress. Work conducted in our laboratory on Streptococcus mutans, the principal acidogenic component of human dental plaque, revealed a DtxR homolog that regulates a repertoire of genes, and accumulating evidence is consistent with a role for iron in S. mutans biofilm formation, the oxidative stress response, and acid tolerance. Indeed, the identification and characterization of genes involved in these and other pathogenic processes is a high priority since they can prove to be future drug targets. The major goal of this research proposal will focus on elucidating iron-dependent gene regulation and its role in S. mutans-induced cariogenesis. The specific aims include 1) characterizing iron-responsive genes that belong to a putative S. mutans Dlg regulon; 2) cloning and characterizing other iron-dependent genes in S.mutans that are not subject to Dlg control; 3) examining the expression of S. mutans Dlg- and iron-responsive genes in environments that approximate the human oral cavity; and 4) establishing a correlation between Dlg/iron-regulated gene expression and S. mutans-induced cariogenesis.