Characterization of ??S in the stress & virulence responses of S. aureus

描述（由申请方提供）：金黄色葡萄球菌是一种高毒力和广泛成功的病原体，推测是人类疾病的最常见原因。随着多重耐药菌株的不断出现，识别新的靶标对于我们对抗回到前抗生素时代至关重要。S.金黄色葡萄球菌作为病原体的致病性在很大程度上是由于其大量的毒力决定因子，或者更具体地说，是由于调节这些和其他遗传组分的复杂的调节元件网络。从发病机制的角度来看，这种控制网络的存在可能是为了微调宿主环境中生物体的需求。我们最近发现了这个调控网络的一个新的组成部分：一个替代的西格玛因子（CS），这不仅是S。金黄色葡萄球菌的适应和生存，但也对其毒力至关重要。使用脓毒性关节炎的小鼠模型，我们发现感染sigS突变体的小鼠显示出显著降低的体重减轻、死亡率、感染的严重程度、全身传播和安装的免疫应答。因此，我们的特定假设是，CS代表了S的主要缺失成分。金黄色葡萄球菌毒力调控网络。我们认为，在本申请中提出的对其功能和控制的彻底分析将最终使我们能够更好地了解这种生物体的发病机制，并可能有助于合理开发新的治疗方法。为此，我们建议：1.分析S.金黄色。我们将使用启动子定位、DNA-蛋白质下拉分析和高通量报告基因融合技术，结合生长操作来确定S.金黄色。2.明确了CS在S.金黄色葡萄球菌应激和毒力应答。我们将使用微阵列分析来确定哪些基因是由CS控制的;并使用表型微阵列来探测CS对S的代谢和生理贡献。aureus生物学3.研究控制S.金黄色。ECF-sigma因子通常通过蛋白质相互作用的复杂级联以翻译后方式调节，由抑制和蛋白水解介导。我们将探索S。利用基因转录技术、特异性蛋白质分析、酵母双杂交研究和诱变筛选，研究了CS调控网络的金黄色葡萄球菌特异性组分。感染由S.金黄色葡萄球菌在全球范围内呈上升趋势，因此，必须尽一切努力了解其发病机制。本文的研究将为我们提供一个独特的和具体的洞察的分子机制，S。金黄色葡萄球菌病，通过映射的功能和控制的一个主要的新的毒力调节。这些知识也可能有助于合理设计新的抗葡萄球菌治疗方法。最后，鉴于CS在葡萄球菌中的独立进化起源，我们的研究可能会提供有关原核调控网络的新信息。 公共卫生相关性：金黄色葡萄球菌是一种高毒性和广泛成功的病原体，被推测是人类感染的最常见原因。随着多重耐药菌株的不断出现，金黄色葡萄球菌（如MRSA），迫切需要了解这种致命病原体引起疾病的机制。本研究旨在研究和了解一种新的毒力调节因子CS是如何影响沙门氏菌致病的。金黄色。

DESCRIPTION (provided by applicant): Staphylococcus aureus is a highly virulent and widely successful pathogen that is speculated to be the most common cause of human disease. With the continued emergence of multi-drug resistant isolates, the identification of novel targets is crucial in our fight against a return to the pre-antibiotic era. The success of S. aureus as a pathogen is largely due to its vast array of virulence determinants, or more specifically, the complex network of regulatory elements that modulate these and other genetic components. From the perspective of pathogenesis, this network of control presumably exists to fine-tune the requirements of the organism within the host environment. We have recently identified a novel component of this regulatory network: an alternative sigma factor (CS) which is required not only for S. aureus adaptation and survival, but is also vital for its virulence. Using a murine model of septic arthritis we found that mice infected with a sigS mutant displayed significantly decreased weight-loss, mortality, severity of infection, systemic dissemination and mounted immune responses. Thus our specific hypothesis is that CS represents a major missing component of the S. aureus virulence regulatory network. We contend that the thorough analysis of its function and control proposed in this application will ultimately enable us to better understand the pathogenesis of this organism, and may aid in the rational development of novel therapeutic treatments. To this end we propose to: 1. Analyze the mechanisms required for sigS induction in S. aureus. We will use promoter mapping, DNA- protein pulldown assays, and high-throughput reporter gene fusion technologies coupled with growth manipulation to determine the exact mechanisms responsible for the induction of CS activity in S. aureus. 2. Define the function of CS in the S. aureus stress and virulence responses. We will use microarray analysis to determine exactly which genes are controlled by CS; and phenotypic microarrays to probe the metabolic and physiological contributions made by CS to S. aureus biology. 3. Investigate the specific pathways that control CS activity in S. aureus. ECF-sigma factors are commonly regulated in a post- translational manner via a complex cascade of protein interaction, mediated by inhibition and proteolysis. We will probe S. aureus specific components of the CS regulatory network using gene-transcription technologies, specific protein analysis, yeast-2-hybrid studies and mutagenic screens. With infections caused by S. aureus on the rise globally it is vital that every effort is exerted to understand the mechanisms involved in its pathogenesis. The investigations proposed here will provide us with a unique and specific insight into the molecular mechanisms of S. aureus disease, by mapping the function and control of a major new virulence regulator. The knowledge derived may also aid in the rational design of novel anti-staphylococcal therapies. Finally, given the independent evolutionary origins of CS in the staphylococci, our investigations will likely deliver new information regarding prokaryotic regulatory networks. PUBLIC HEALTH RELEVANCE: Staphylococcus aureus is a highly virulent and widely successful pathogen that is speculated to be the most common cause of human infection. With the continued emergence of multi-drug resistant isolates of S. aureus (such as MRSA), there is an urgent need to understand the mechanisms by which this deadly pathogen causes disease. This proposal seeks to investigate and understand how a novel regulator of virulence, CS, contributes to disease causation in S. aureus.