Impact of ß-glucan metabolism on the development of pathogenic biofilms

项目摘要/摘要 牙周炎是由细菌引起的牙齿支持组织的慢性炎症。 这可能会导致牙槽骨破坏和牙齿脱落。连翘坦纳氏菌(Tannerella forsythia(Tf)) 属于红色复合体，是牙周炎的主要病原体之一。T. 连翘已被证明与核梭杆菌的“桥细菌”共同聚集 (FN)在体外形成协同共生物膜并诱导小鼠牙槽骨丢失 感染了新城疫。这项提议的总体目标是确定 连翘-F。核糖体属间相互作用以更好地理解 牙周炎。我们的初步数据显示，一种Tf分泌β-葡聚糖酶(GLCA)，其 诱导表达是对FN反应的反应，在混合性发育中起重要作用 生物膜。我们发现这种酶能将？葡聚糖水解成葡萄糖，而葡萄糖是FN的营养物质。 提高其在TF-FN共生物膜中的生物量。我们的数据显示，葡萄糖可利用性的增加并没有 影响转铁蛋白的生物量，但却增加了甲基乙二醛(MGO)的产生，这是一种高活性的二苯甲基 对细菌和宿主细胞有毒性的化合物。 这项建议的目标是了解Tf-FN相互作用的机制基础 以及这两个物种之间的新陈代谢相互作用如何促进 牙菌斑的发展、生物失调和炎症。我们的工作假设是 Tf在Fn和可能的其他刺激下产生的β-葡聚糖酶，从饮食中释放葡萄糖。 葡聚糖是整个微生物群落的营养物质，也是MGO分泌的代谢前体 有利于微生物的生态失调。为了验证这一假设，我们提出了两个具体目标：目标1： 目的：探讨转录因子Glca操纵子调节FN感觉的分子机制。 我们将分析预测的ECF西格玛-反西格玛系统的机制和功能 驱动Tf Glca？-葡聚糖酶操纵子对Fn和其他刺激物作出反应，以及；目标2 为了确定FN抵抗Tf产生的甲基乙醛的机制以及如何 水解性葡聚糖影响微生物群落结构。在这里，我们将确定分子 FN解毒生物膜中Tf产生的氧化镁并促进牙菌斑形成的机制 发展。 这项研究的圆满完成将为今后的调查探索奠定基础 Tf葡聚糖酶-MGO轴在促进微生物生态失调中的独特能力及其后续发展 葡聚糖酶靶向抑制剂阻断协同的Tf-Fn关联和牙菌斑 牙周炎的治疗进展。

Project Summary/Abstract Periodontitis is a bacterially induced chronic inflammation of the tooth supporting tissues that may result in alveolar bone destruction and tooth loss. Tannerella forsythia (Tf), a member of the ‘red-complex’ bacteria group, is one of the major pathogens implicated in periodontitis. T. forsythia has been shown to co-aggregate with the ‘bridge-bacterium’ Fusobacterium nucleatum (Fn) to form synergistic co-biofilms in vitro and induce alveolar bone loss in mice when co- infected with Fn. The overall goal of this proposal is to determine the molecular mechanisms of T. forsythia -F. nucleatum intergeneric interactions to better understand the pathogenesis of periodontitis. Our preliminary data show that a Tf secreted ß-glucanase enzyme (GlcA) whose expression is induced in response to Fn sensing plays a significant role in the development of mixed biofilms. We showed that this enzyme hydrolyzes ß-glucans into glucose, which serves as a nutrient for Fn to promote its biomass in Tf -Fn co-biofilms. Our data showed that the increased glucose availability did not affect Tf biomass but it rather enhanced the production of methylglyoxal (MGO), a highly reactive dicabonyl compound toxic to bacterial and host cells. The goal of this proposal is to understand the mechanistic basis of Tf-Fn interactions and how metabolic interactions between these two species contribute to the development of the dental plaque, dysbiosis and inflammation. Our working hypothesis is that b-glucanase produced by Tf in response to Fn, and possible other stimuli, releases glucose from dietary b- glucans as a nutrient for the microbial community at large and as a metabolic precursor for MGO secretion to favor microbial dysbiosis. To interrogate this hypothesis, we propose two specific aims: Aim 1: To define the molecular mechanism of regulation of Tf GlcA operon in response to Fn sensing. We will analyze the mechanism and function of ECF sigma-anti sigma system that is predicted to drive Tf glcA ß-glucanase operon to respond stimulate with Fn and other stimuli, and; Aim 2 To determine the mechanisms by which Fn resists Tf-produced methylglyoxal and how hydrolyzed glucans impact microbial community structure. Here, we will determine the molecular mechanisms by which Fn detoxifies MGO produced by Tf in biofilms and promote dental plaque development. Successful completion of this study will form the foundation for future investigations exploring the unique ability of Tf glucanase-MGO axis in promoting microbial dysbiosis and subsequently develop glucanase targeting inhibitors to block the synergistic Tf-Fn associations and dental plaque development for the treatment of periodontitis.