Mechanistic studies of rot-mediated regulation of Staphylococcus aureus virulence

描述(申请人提供)：金黄色葡萄球菌是医院和社区中威胁生命的细菌感染的主要原因之一。这种细菌几乎可以侵袭和侵袭人体的所有组织。金黄色葡萄球菌作为病原体的巨大成功部分是由于毒力因子的暂时表达，这些毒力因子是由一系列双组分调控系统(TCS)和螺旋-转角-螺旋蛋白执行的。遗传分析表明，金黄色葡萄球菌毒力因子调控的核心是转录因子毒素抑制因子(Rot)和金黄色葡萄球菌外膜蛋白表达双组分系统(SaeRS-TCS)。使用小鼠全身感染模型的实验表明，没有腐烂的金黄色葡萄球菌菌株是超强毒力的，而缺乏sae的菌株是完全减毒的，这突显了这些因素的重要性。 金黄色葡萄球菌的病理生理学调节剂。ROT被认为是通过抑制编码细胞毒素和蛋白酶的基因的启动子，以及激活编码表面蛋白的基因的启动子来协调毒力因子的差异表达。相反，SaeRS-TCS参与了启动子的激活，这些启动子编码了包括细胞毒素在内的大多数分泌因子。对临床相关菌株的Rot和SaeRS调节子的比较表明，很大一部分腐烂抑制基因需要SaeRS-TCS来激活它们。Rot调控金黄色葡萄球菌基因表达的确切分子机制尚不清楚。同样，Rot干扰SaeRS-TCS改变靶基因表达的机制仍有待阐明。此外，研究ROT的转录和翻译调控主要是在体外进行的，并不一定模拟该转录因子在体内对毒力因子调控的影响。因此，这一建议的具体目标是：(I)确定Rot介导的毒力因子调节机制，以及(Ii)阐明腐烂转录和翻译的调节，体内外。为了实现这些目标，将采用多学科方法，其中分子生物学、生物化学和细菌学技术将与体内和体外感染模型相结合。总的来说，本申请中提出的工作将阐明Rot如何调节毒力因子的表达 金黄色葡萄球菌的致病机制。由于目前耐药金黄色葡萄球菌的流行，了解金黄色葡萄球菌如何调控毒力因子表达的复杂性可能为开发基于靶向调控电路的新的抗葡萄球菌药物奠定基础，这些调控电路是金黄色葡萄球菌致病所需的。

DESCRIPTION (provided by applicant): Staphylococcus aureus is one of the leading causes of life-threatening bacterial infections in both hospitals and community settings. This bacterium can colonize and invade virtually all tissues in the human body. The tremendous success of S. aureus as a pathogen is in part due to the temporal expression of an arsenal of virulence factors, which is executed by a series of two component regulatory systems (TCS) and helix-turn-helix proteins. Genetic analyses have revealed that at the center of the regulation of virulence factors in S. aureus are the transcription factors repressor of toxins (Rot) and the S. aureus exoprotein expression two-component system (SaeRS-TCS). Experiments using a murine model of systemic infection have revealed that S. aureus strains lacking rot are hypervirulent, while strains lacking sae are fully attenuated, highlighting the importance of these regulators for S. aureus pathophysiology. Rot is hypothesized to coordinate the differential expression of virulence factors by repressing the promoters of genes that code for cytotoxins and proteases, and activating the promoters of genes that code for surface proteins. In contrast, the SaeRS-TCS is involved in the activation of the promoters that code for most secreted factors, including cytotoxins. Comparison of the Rot and SaeRS regulons in clinically relevant strains has revealed that a large portion of Rot-repressed genes require the SaeRS-TCS for their activation. The precise molecular mechanism by which Rot regulates gene expression in S. aureus is not understood. Similarly, the mechanism by which Rot interferes with the SaeRS-TCS to alter the expression of target genes remains to be elucidated. Additionally, studies examining transcriptional and translational regulation of rot have primarily been done in vitro and do not necessarily mimic the impact this transcription factor has on virulence factor regulation in vivo. Thus, the Specific Aims of this proposal seek to: (i) define the mechanism of Rot-mediated regulation of virulence factors, and (ii) elucidate the regulation of rot transcriptio and translation ex vivo and in vivo. To accomplish these Aims, a multi-disciplinary approach will be employed where molecular biology, biochemistry, and bacteriology techniques will be combined with in vivo and ex vivo infection models. Collectively, the work proposed in this application will shed light into how Rot regulates the expression of virulence factors required for S. aureus pathogenesis. Due to the current epidemic of antibiotic-resistant S. aureus strains, understanding the intricacies of how S. aureus regulates the expression of virulence factors is likely to provide the foundation for the development of new anti-Staphylococcal drugs based on targeting regulatory circuits that are required for the pathogenesis of this bacterium.