Proteolytic regulation of the Streptococcus pyogenes cell surface

项目摘要/摘要 化脓性链球菌(Spy；Group A)是致病菌中数量最多的一种 它要对疾病负责。最常见的是间谍是儿童急性咽炎(链球菌性咽喉炎)的原因，但它 也是死亡率排名前十的病原体，每年通过入侵导致50多万人死亡 感染和免疫疾病，包括坏死性筋膜炎、中毒性休克综合征和急性风湿病 发烧。尽管它们对公共卫生具有重要意义，但人们对这些严重疾病的发病机制知之甚少， 而且它们的治疗仍然具有挑战性。我们的长期目标是确定间谍毒力因素是如何起作用的 这些更严重的疾病的发展，以便有效的治疗对策 被开发出来。我们以前的研究表明，链球菌蛋白酶SpeB直接作用于宿主细胞因子 前-IL-1β在侵袭性感染过程中诱导炎症病理。我们假设额外的蛋白质 SpeB的靶标，来自宿主和微生物，有助于我们在 感染。在我们的初步数据中，我们已经表征了SpeB的生化活性，并鉴定了两种 它的底物协同导致发病机制的根本转变。在我们的第一个目标中，我们研究如何 SpeB调节M蛋白的活性，M蛋白是一种具有多种宿主靶标的多效性毒力因子。通过 消除某些宿主分子结合的特异性结构域或将M蛋白锚定到Spy表面， SpeB直接导致感染的病理并发症，如肾小球肾炎和中毒性休克 综合症。在我们的第二个目标中，我们定义了SpeB如何通过 COVR/COVS通过靶向其激动剂，宿主防御肽LL-37来调节双组分。这一机制 结果形成一个正反馈环路，以产生SpeB并抑制胶囊和其他细胞表面因子 这改变了抗菌素的耐药性，被宿主检测，并促进炎症和疾病并发症。 这两个目标都利用了我们开发的新的遗传和生化工具，用于靶向控制和特定的 SpeB及其底物的检测。这些创新使我们第一次能够将这些活动分开 多功能蛋白，并检测Spy在SpeB调控下的毒力策略的体内戏剧性变化。 这项拟议的研究的预期贡献是扩大了对间谍发病机制的理解 为严重间谍疾病的预防和治疗提供有吸引力的治疗目标的受监管的 对与NIAID任务高度相关的其他病原体的直系系统的洞察。

PROJECT SUMMARY/ABSTRACT Streptococcus pyogenes (Spy; group A Streptococcus) is the most prolific pathogen in the number of distinct diseases it is responsible for. Most commonly Spy is a cause of childhood acute pharyngitis (strep throat), but it is also a top-10-pathogen in mortality that results in more than half a million deaths annually through invasive infections and immune diseases that include necrotizing fasciitis, toxic shock syndrome, and acute rheumatic fever. Despite their public health importance, the pathogenesis of these severe diseases is poorly understood, and they remain challenging to treat. Our long-term goal seeks to determine how Spy virulence factors contribute to the development of these more severe forms of disease so that effective therapeutic countermeasures can be developed. Our previous studies show that the streptococcal protease SpeB acts directly on the host cytokine pro-IL-1β to induce inflammatory pathology during invasive infection. We hypothesize that additional proteins targeted by SpeB, from both host and microbe, contribute to the proinflammatory responses we see during infection. In our preliminary data, we have characterized the biochemical activity of SpeB and identified two of its substrates that cooperatively lead to fundamental shifts in pathogenesis. In our first aim, we examine how SpeB regulates the activities of M protein, a pleiotropic virulence factor with numerous host targets. By eliminating domains specific for the binding of some host molecules or anchoring of M protein to the Spy surface, SpeB directly contributes to pathological complications of infection like glomerulonephritis and toxic shock syndrome. In our second aim, we define the molecular determinates of how SpeB blocks signaling by the CovR/CovS two-component regulator by targeting its agonist, the host defense peptide LL-37. This mechanism results in a positive-feedback loop for SpeB production and repression of capsule and other cell surface factors that alter antimicrobial resistance, detection by the host, and promotes inflammation and disease complications. Both aims take advantage of new genetic and biochemical tools we developed for targeted control and specific detection of SpeB and its substrates. These innovations allow us for the first time to separate activities for these multifunctional proteins and examine in vivo dramatic switch in the virulence strategy of Spy regulated by SpeB. The expected contribution of the proposed research is an expanded understanding of how Spy pathogenesis is regulated that provides attractive therapeutic targets for the prevention and treatment of severe Spy disease and insights into orthologous systems of other pathogens highly relevant to the NIAID mission.