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

应力中 ??S 的表征

基本信息

批准号：
8115251
负责人：
Lindsey Neil Shaw
金额：
$ 32.74万
依托单位：
UNIVERSITY OF SOUTH FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8115251
关键词：
Acute suppurative arthritis due to bacteria Antibiotic Resistance Antibiotics Biochemical Biology Body Weight decreased Cell Wall Cells Clinical Community Hospitals Complex Coupled Cues DNA mapping Data Analyses Development Disease Ensure Environment Etiology Gene Expression Gene Fusion Genes Genetic Genetic Transcription Genus staphylococcus Glycopeptide Antibiotics Growth Human Hybrids Immune response Infection Investigation Knowledge Maps Mediating Membrane Proteins Metabolic Methods Microarray Analysis Modeling Molecular Molecular Profiling Multi-Drug Resistance Mus Nosocomial Infections Organism Other Genetics Pathogenesis Pathway interactions Peptide Hydrolases Perception Phenotype Physiological Primer Extension Process Protein Analysis Proteins Proteolysis Regulation Regulation of Proteolysis Regulatory Element Reporter Reporter Genes Resistance Resort Role Severities Sigma Factor Signaling Protein Staphylococcus aureus Stimulus Stress Switch Genes System Systemic infection Technology Therapeutic Virulence Virulent Yeasts combat design fight against high throughput screening human disease innovation insight methicillin resistant Staphylococcus aureus mortality mutant novel novel therapeutics pathogen promoter public health relevance response success tool

项目摘要

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.

描述（由申请人提供）：金黄色葡萄球菌是一种高毒力且广泛成功的病原体，据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，新靶标的识别对于我们阻止回到抗生素前时代的斗争至关重要。金黄色葡萄球菌作为病原体的成功很大程度上归功于其大量的毒力决定因素，或更具体地说，是调节这些和其他遗传成分的复杂的调控元件网络。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。我们最近发现了该调控网络的一个新组成部分：替代西格玛因子（CS），它不仅是金黄色葡萄球菌适应和生存所必需的，而且对其毒力也至关重要。使用化脓性关节炎的小鼠模型，我们发现感染 sigS 突变体的小鼠体重减轻、死亡率、感染严重程度、全身传播和免疫反应显着降低。因此，我们的具体假设是，CS 代表了金黄色葡萄球菌毒力调节网络的一个主要缺失组成部分。我们认为，本申请中提出的对其功能和控制的彻底分析最终将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型治疗方法。为此，我们建议： 1. 分析金黄色葡萄球菌中 sigS 诱导所需的机制。我们将使用启动子作图、DNA-蛋白下拉分析、高通量报告基因融合技术以及生长操作来确定金黄色葡萄球菌中诱导 CS 活性的确切机制。 2. 定义CS在金黄色葡萄球菌应激和毒力反应中的功能。我们将使用微阵列分析来准确确定哪些基因受CS控制；和表型微阵列来探测 CS 对金黄色葡萄球菌生物学的代谢和生理贡献。 3. 研究金黄色葡萄球菌中控制 CS 活性的具体途径。 ECF-sigma 因子通常通过抑制和蛋白水解介导的复杂的蛋白质相互作用级联以翻译后方式进行调节。我们将利用基因转录技术、特定蛋白质分析、酵母 2 杂交研究和诱变筛选来探测金黄色葡萄球菌 CS 调控网络的特定组成部分。随着金黄色葡萄球菌引起的感染在全球范围内呈上升趋势，尽一切努力了解其发病机制至关重要。这里提出的研究将为我们提供对金黄色葡萄球菌疾病分子机制的独特而具体的见解，通过绘制一个主要的新毒力调节剂的功能和控制。获得的知识也可能有助于合理设计新型抗葡萄球菌疗法。最后，考虑到葡萄球菌中 CS 的独立进化起源，我们的研究可能会提供有关原核调节网络的新信息。公共卫生相关性：金黄色葡萄球菌是一种高毒力且广泛成功的病原体，据推测是人类感染的最常见原因。随着多重耐药金黄色葡萄球菌分离株（例如 MRSA）的不断出现，迫切需要了解这种致命病原体引起疾病的机制。该提案旨在调查和了解新型毒力调节剂 CS 如何导致金黄色葡萄球菌致病。