Intracellular Invasion by Streptococcus mutans: Significance in Disease

摘要 人类病原体变形链球菌是与龋齿相关的主要病原体， 感染性心内膜炎（IE）的常见病原体。最近有证据表明，S。 产生胶原蛋白结合蛋白Cnm的变形杆菌可能与更严重的IE病例相关， 其他口腔外病变的发展，包括出血性中风和伊加肾病。而 产cnm菌株在口腔中并不普遍， 感染cnm+ S.变形菌增加龋齿的发病率和严重程度。此外，在我们的研究中， 实验室研究表明，S.产生Cnm的变异菌株可以更有效地粘附在富含胶原蛋白的细胞上， 存在于离体心脏瓣膜和牙齿（即牙本质）中的组织，并且能够侵入口腔上皮和心脏 体外培养内皮细胞。利用兔IE和大鼠龋模型，我们证实了Cnm是一种重要的 毒力因子表明S.产生Cnm的变异株可以是高毒性的。获得的数据 到目前为止，表明Cnm赋予的性质使S.口腔中的变形菌持久性和毒力 腔，也可以促进全身传播和感染。利用分子和生物化学 方法中，我们发现Cnm被一种新鉴定的糖基化复合物后修饰， 它被命名为PGF。Pgf系统的丧失通过影响其稳定性而损害Cnm功能。我们还发现 Cnm具有淀粉样变性的特性，尽管Cnm形成淀粉样蛋白对胶原结合和胶原降解的影响是不明显的。 相关功能目前尚不清楚。我们认为，糖基化状态和淀粉样形成的形式， Cnm可通过促进蛋白质稳定性（糖基化）或通过赋予新的糖基化活性来促进其功能。 特性（淀粉样蛋白形成）到Cnm。本申请的首要假设是cnm+ S。变形 菌株更适合在人体组织中定植，包括口腔软组织和暴露的牙本质和牙齿 根，因此可能与更具侵略性和复发性的龋齿病例有关，例如 发现于儿童早期龋齿。我们推测cnm+ S.变异人与大多数 侵蚀性龋齿病例，以及无法用常规风险解释的龋齿复发 高含蔗糖食品摄入量和口腔卫生不良等因素。在这个应用程序中，我们将 使用生物化学，分子，糖蛋白质组学和结构方法来表征Cnm， 糖蛋白和淀粉样蛋白，以及大鼠龋齿模型，以揭示Cnm在口腔中的意义。 定植、持久性和致龋性。这项研究的结果将揭示其他方面的相关 感染不同的S.变异，这可能会导致目前的范式的变化，评估 龋齿风险，并最终帮助设计新的抗菌治疗。

ABSTRACT The human pathogen Streptococcus mutans is a major etiological agent associated in dental caries and a common causative agent of infective endocarditis (IE). Evidence has recently emerged that strains of S. mutans producing the collagen binding protein Cnm may be associated with more severe cases of IE and with the development of other extra-oral pathologies including hemorrhagic stroke and IgA nephropathy. While Cnm-producing strains are not prevalent in the oral cavity, crosss-sectional clinical studies have linked infection with cnm+ S. mutans with increased caries incidence and severity. Moreover, studies conducted in our laboratory have showed that S. mutans strains producing Cnm can more efficiently adhere to collagen-rich tissues present in heart valves and in teeth (i.e. dentin) ex vivo, and are able to invade oral epithelial and heart endothelial cells in vitro. Using the rabbit IE and rat caries models, we confirmed that Cnm is an important virulence factor indicating that S. mutans strains that produce Cnm can be hypervirulent. The data obtained thus far indicate that the properties conferred by Cnm enables S. mutans persistence and virulence in the oral cavity that could also facilitate systemic dissemination and infection. Using molecular and biochemical approaches, we found that Cnm is posttranslationally modified by a newly identified glycosylation complex that was named Pgf. Loss of the Pgf system impaired Cnm function by affecting its stability. We also found that Cnm has amylodoigenic properties albeit the impact of amyloid formation by Cnm on collagen-binding and related functions is presently unknown. We propose that the glycosylation state and amyloidogenic form of Cnm may contribute to its function by promoting protein stability (glycosylation) or by conferring novel properties (amyloid formation) to Cnm. The overarching hypothesis of this application is that cnm+ S. mutans strains are better equipped to colonize human tissues, including oral soft tissues and exposed dentin and teeth roots, and, therefore may be associated with more aggressive and reincident cases of caries, such as those found in early childhood caries. We postulate that oral infection by cnm+ S. mutans are linked to the most aggressive cases of caries, as well as to caries recurrence that cannot be explained by conventional risk factors such as high intake of sucrose-containing foodstuffs and poor oral hygiene. In this application, we will use biochemical, molecular, glycoproteomic and structural approaches to characterize Cnm as both a glycoprotein and an amyloidogenic protein, and the rat caries model to uncover the significance of Cnm in oral colonization, persistence and cariogenicity. The findings of this study will reveal additional facets associated with infection by different strains of S. mutans, which could lead to changes in current paradigms for assessing caries risk and ultimately help devise novel antimicrobial therapies.