Impact of regulatory cross-talk on the pathophysiology of emergent acapsular group A streptococcus

项目总结 人类主要病原体A组链球菌(GAS)的透明质酸胶囊一直是经典的 被认为对传染性是必不可少的。根据emm基因的变异，GAS被分为emm类型 编码细胞表面、抗吞噬的M蛋白。最近人们认识到，来自 缺乏包膜的EMM型正在日益引起严重的人类感染，以至于大约30%的 侵袭性气体分离株是无囊化的。无囊性气体是如何导致感染的目前尚不清楚，尽管 据推测，无囊化气体菌株产生了一种高水平的关键细胞毒素--链球菌溶血素O(Slo)。 与这一猜测相反，我们发现多种囊状EMM类型不会产生高水平的SLO， 因此，我们研究的长期目标是阐明无囊性气体的机制基础。 病理生理学。通过比较基因组学方法，我们已经鉴定出无囊性气体EMM 类型有不同的MGA和EMM启动子的组成，它们编码关键的转录 调节因子多基因激活剂(MGA)和M蛋白。此外，我们发现，控制 毒力因子(CovRs)双组分基因调控系统特异性影响MGA和EMM转录水平 相对于被包裹的气体，形成无囊状。这些和其他发现为这项提案奠定了基础 旨在验证MGA和EMM启动子组成对无囊化病毒感染性至关重要的假设。 通过允许CovRs的调节，这对GAS感知和响应宿主信号至关重要。在……里面 具体目标1，我们将确定无囊性气体的MGA和EMM启动子的差异与 作为回应，包裹的气体改变了囊状emm4和emm89菌株的细胞表面成分。 人类抗菌肽LL-37。我们还将评估MGA和EMM启动子组成的影响 包囊气体对人上皮细胞和人细胞外基质成分的黏附作用 人血，对吞噬的抵抗力，以及在小鼠模型中的毒力。在具体目标2中，我们将确定 如果在无囊气体的MGA和EMM启动子中观察到的变化允许COVR直接调节 也是CoVS失活影响MGA和EMM转录水平的机制。我们会调查的 COVR与MGA和EMM启动子的体内相互作用及关键启动子区域的鉴定 到使用基于荧光素酶的报告系统的基于COVR的调控。我们还将评估COVR是否直接 抑制无胶囊气体中的EMM，独立于MGA。完成本文概述的研究将 极大地增强了对无囊性气体产生的分子机制的理解 它们是气体感染的主要原因，但相对于被包裹的菌株，人们仍然知之甚少。

PROJECT SUMMARY The hyaluronic acid capsule of the major human pathogen group A Streptococcus (GAS) has classically been considered essential to infectivity. GAS is divided into emm types based on variation in the emm gene that encodes for the cell-surface, anti-phagocytic M protein. It has recently been appreciated that GAS strains from emm types lacking capsule are increasingly causing serious human infections to the point where some 30% of invasive GAS isolates are acapsular. How acapsular GAS causes infection is not currently known although it has been postulated that acapsular GAS strains elaborate high levels of a key cytotoxin, streptolysin O (Slo). Contrary to this conjecture, we have found that multiple acapsular emm types do not produce high levels of Slo, and thus the long term goal of our research is to elucidate the mechanistic basis of acapsular GAS pathophysiology. Through a comparative genomics approach, we have identified that acapsular GAS emm types have distinct compositions of the promoters of mga and emm which encode the key transcriptional regulator multi-gene activator (Mga) and M protein, respectively. Moreover, we have discovered that the control of virulence (CovRS) two-component gene regulatory system specifically impacts mga and emm transcript levels in acapsular relative to encapsulated GAS. These, and other, findings set the stage for this proposal which seeks to test the hypothesis that mga and emm promoter composition are critical to the infectivity of acapsular GAS by allowing for regulation by CovRS, which is critical for GAS to sense and respond to host signals. In specific aim 1, we will determine how variation in mga and emm promoters of acapsular GAS compared to encapsulated GAS alters the composition of the cell surface of acapsular emm4 and emm89 strains in response to the human antimicrobial peptide LL-37. We will also assess the impact of mga and emm promoter composition on acapsular GAS adherence to human epithelial cells and human extracellular matrix components, survival in human blood, resistance to phagocytosis, and virulence in a mouse model. In specific aim 2, we will determine if the changes observed in the mga and emm promoters of acapsular GAS allow for direct regulation by CovR and is the mechanism by which CovS inactivation influences mga and emm transcript levels. We will investigate in vivo interaction of CovR with mga and emm promoters and attempt to identify promoter regions that are key to CovR-based regulation using a luciferase based reporter system. We also will assess whether CovR directly represses emm in acapsular GAS independent of Mga. Completion of the research outlined herein will significantly augment understanding of the molecular mechanisms underlying the emergence of acapsular GAS which are a major cause of GAS infection yet remain poorly understood relative to encapsulated strains.