Molecular and Cellular Basis of Toxin Mediated Pathogenesis in Staphylococcus aur

描述(由申请人提供)：金黄色葡萄球菌是一个主要的公共卫生问题，每年在美国导致超过30万人住院(1-2)。细菌是导致化脓性皮肤损害、细菌性心内膜炎、菌血症和肺炎的主要原因。金黄色葡萄球菌感染的治疗因出现大量耐药菌株而变得复杂(2)。以前仅限于医院环境，由金黄色葡萄球菌引起的社区获得性感染继续出现，通常发生在其他健康的个人中(2)。金黄色葡萄球菌分泌大量毒力因子，促进其在敏感宿主中的存活。在这些因素中有许多分泌的毒素，它们直接与宿主细胞相互作用并杀死/破坏宿主细胞(3-5)。毒素表达的协调调控是金黄色葡萄球菌致病的关键。我们的数据表明，通过转录抑制来调节毒素的产生是白毒素LukED最佳表达的关键。当这种调控模式被扰乱时，由于LukED产量增加，金黄色葡萄球菌菌株在感染小鼠模型中变得超强毒力。本申请中描述的实验旨在(I)阐明LukED基因在金黄色葡萄球菌中表达的调控机制(S)，以及(Ii)确定LukED中毒在体外、体外和体内的功能影响。细菌遗传学、启动子结合分析、体外细胞毒性和存活分析、体内败血症感染模型和免疫学技术将是实现上述目标的关键。实验将研究体内协调表达的LukED基因的影响，毒素在细菌介导的哺乳动物细胞杀伤中的作用，以及毒素对体内免疫细胞募集/存活的影响。最终，我们将确定以下主要贡献 LukED在体内的致病机制，并将提供对感染所需的最佳毒素表达模式的洞察。重要的是，这项工作将通过研究金黄色葡萄球菌引起疾病的机制基础来促进国家卫生研究院的使命，最终目标是改进预防和治疗战略。

DESCRIPTION (provided by applicant): Staphylococcus aureus is a major public health concern and is responsible for over 300,000 hospitalizations in the United States each year (1-2). The organism is a leading cause of suppurative skin lesions, bacterial endocarditis, bacteremia, and pneumonia. Treatment of S. aureus infection is complicated by the emergence of numerous antibiotic resistant strains (2). Once previously limited to hospital settings, community acquired infections due to S. aureus continue to arise, often in otherwise healthy individuals (2). S. aureus secretes an arsenal of virulence factors that promote the organism's survival in susceptible hosts. Among these factors are a number of secreted toxins that directly interact with and kill/damage host cells (3-5). The coordinated regulation of toxin expression is critical to S. aureus pathogenesis. Our data demonstrate that regulation of toxin production by transcriptional repression is critical to optimal expression of the leukotoxin LukED. When such regulatory patterns are perturbed, S. aureus strains become hypervirulent in mouse models of infection due to increased LukED production. Experiments described in this application are designed to (i) elucidate the regulatory mechanism(s) of lukED gene expression in S. aureus and (ii) determine the functional impact of LukED intoxication in vitro, ex vivo, and in vivo. Bacterial genetics, promoter binding assays, in vitro cell toxicity and survival assays, in vivo models of septicemic infection, and immunological techniques will be critical to the execution of the above aims. Experiments will investigate the impact of coordinated lukED gene expression in vivo, the role of the toxin in bacterial mediated mammalian cell killing, and the toxin's effecton immune cell recruitment/viability in vivo. Ultimately, we will determine the major contributions of LukED to pathogenesis in vivo and will provide insight into the optimal toxin expression patterns required for infection. Importantly, this work will further the mission of the National Institutes f Health by examining the mechanistic underpinnings of disease caused by S. aureus, with an end goal of improving prevention and treatment strategies.