Dynamics of the bacterial type IX secretion system and its effect on subgingival biofilm formation by bacteria of the human oral microbiome

项目概要/摘要 细菌IX型蛋白质分泌系统（T9 SS），其是最近发现的蛋白质分泌系统。 系统，发现在细菌是人类龈下生物膜的一部分。到目前为止，已经表明， T9 SS在牙龈卟啉单胞菌[1，2]、嗜酸坦纳菌[3]和二氧化碳噬纤维菌属中有功能。 [4]的文件。牙龈卟啉单胞菌Arg-牙龈卟啉菌蛋白酶（Rgp）和Lys-牙龈卟啉菌蛋白酶（Kgp）通过T9 SS分泌。Rgp和Kgp 通过破坏宿主免疫反应和降解牙周膜， 宿主组织和血浆蛋白。T9 SS与旋转电机相关联，其动力学使得 分泌细胞表面粘附素、已知的毒力因子和酶。它也需要为 细菌通过一种叫做滑动运动的过程在表面上移动。细胞粘附，表面 导航和运动对于许多细菌生物膜的形成是重要的[5]。最近的报告 这表明细菌运动机制能够通过一种称为 机械感测[6，7]。人类病原体，如大肠杆菌和沙门氏菌 外表面通过鞭毛马达III型分泌系统机械[6]，而假单胞菌 铜绿假单胞菌和黄色粘球菌通过IV型皮利机制感知外表面[7，8]。我们 初步和已发表的结果表明，携带T9 SS的滑行细菌具有感知 外表面和粘性环境[9]。这种细菌没有鞭毛马达或 IV皮利。在人类口腔微生物组的细菌中，二氧化碳噬细胞菌属的成员，其是 存在于人类龈下生物膜中[10]，是回答重要问题的有吸引力的系统 与生物膜形成、表面传感和流体力学有关。这些细菌具有T9 SS， 它们在外表面上表现出滑动运动性。最近，基因工具被开发出来， 操纵[4]。我建议研究牙龈二氧化碳嗜纤维菌，这是一个重要的，但研究不足， 细菌存在于人类龈下生物膜[4，10]。我们发现C.齿龈 由一种新颖的旋转马达驱动。此外，我们还发现C.牙龈细胞利用T9 SS分泌的粘附素 聚集并形成群体。这种聚集体在外表面上表现出滑动运动性。我们的数据 这表明滑翔是由相同的旋转马达驱动的，该马达通过T9 SS为分泌提供动力。滑翔 运动性是T9 SS动力学的输出，将用于测定基因调控和 基因筛选的发展。这种筛选将针对鉴定所涉及的蛋白质 在口腔细菌的感觉传导中。总的来说，T9 SS的动力学和由T9 SS分泌的蛋白质的作用是相似的。 T9 SS将在龈下生物膜形成的背景下进行研究。

Project Summary/Abstract The bacterial Type IX protein secretion system (T9SS), which is a recently discovered protein secretion system, is found in bacteria that are a part of human subgingival biofilms. So far, it has been shown that T9SS is functional in Porphyromonas gingivalis [1, 2], Tannerella forsythia [3] and Capnocytophaga sp. [4]. P. gingivalis Arg-gingipain (Rgp) and Lys-gingipain (Kgp) are secreted via T9SS. Rgp and Kgp increase the risk of periodontal diseases by disrupting the host immune response and degradation of the host tissue and plasma proteins. T9SS is associated with a rotary motor and its dynamics enables secretion of cell-surface adhesins, known virulence factors and enzymes. It is also required for movement of bacteria over a surface by a process called gliding motility. Cell adhesion, surface navigation and motility are important for the formation of many bacterial biofilms [5]. Recent reports suggest that bacterial motility machineries enable sensing of external surfaces via a process called as mechanosensing [6, 7]. Human pathogens such as Escherichia coli and Salmonella enterica sense external surfaces via the flagellar motor-Type III secretion system machinery [6], while Pseudomonas aeruginosa and Myxococcus xanthus sense external surface via the Type IV pili machinery [7, 8]. Our preliminary and published results suggest that gliding bacteria with T9SS have the ability to sense external surfaces and viscous environments [9]. Such bacteria do not have the flagellar motor or the type IV pili. Amongst bacteria of the human oral microbiome, members of Capnocytophaga genus, which are present in human subgingival biofilms [10], are an attractive system for answering important questions related to biofilm formation, surface-sensing, and hydrodynamics. These bacteria have the T9SS and they exhibit gliding motility over external surfaces. Recently, genetic tools were developed for their manipulation [4]. I propose to study Capnocytophaga gingivalis, which is an important, yet understudied bacterium present in human subgingival biofilms [4, 10]. We discovered that the T9SS of C. gingivalis is powered by a novel rotary motor. Also, we found that C. gingivalis cells use adhesins secreted by T9SS to aggregate and form groups. Such aggregates exhibit gliding motility over external surfaces. Our data suggests that gliding is powered by the same rotary motor that powers secretion via T9SS. Gliding motility, which is the output of the dynamics of T9SS, will be used to assay for gene regulation and for development of genetic screens. Such screens will be targeted towards identification of proteins involved in sensory transduction by oral bacteria. Overall, dynamics of T9SS and role of proteins secreted by T9SS will be studied in the context of subgingival biofilm formation.