The Mechanisms of Heme Toxicity and Detoxification in Staphylococcus aureus

描述（由申请人提供）：金黄色葡萄球菌是一种已知感染几乎所有器官并导致多种疾病的人类病原体。抗生素抗性S.金黄色葡萄球菌菌株正在迅速出现;因此，正在寻找新的药物靶点。宿主用来限制入侵病原体生长的机制之一是四吡咯分子血红素内必需营养铁的螯合。S.金黄色葡萄球菌采用复杂的血红素摄取系统以获得这种铁源;然而，过量血红素的存在对这种生物体和许多其它生物体具有高度毒性。血红素毒性的性质尚未完全确定，但认为至少部分是由于氧化损伤 由血红素分子内的铁原子的反应性质引起。S.金黄色葡萄球菌具有血红素解毒系统，一种被预测为HrtAB的转运蛋白，尽管对其作用机制知之甚少。在本申请中呈现的初步数据中，我们在血红素敏感hrtA突变体的背景下进行转座子筛选，筛选具有增加的血红素抗性的菌株。鉴定的所有突变体都针对甲萘醌（MK）生物合成途径。这些数据表明，MK生物合成途径的一个组成部分加强血红素应激。有趣的是，S。MK生物合成缺陷的金黄色葡萄球菌菌株（称为小菌落变体（SCV））通常从经历持续感染的患者中分离。虽然多种因素可能有助于选择MK缺陷型S。金黄色葡萄球菌菌株中，似乎对血红素（脊椎动物宿主中丰富的分子）的毒性作用的抗性可能起作用。基于这些数据，我们预测，血红素胁迫的条件是直接相关的环境所经历的S。在人类感染期间。因此，我们建议深入了解血红素胁迫的来源，在S。金黄色葡萄球菌和该病原体利用的机制，以克服血红素的毒性作用是必不可少的，以推进我们的知识，葡萄球菌的生理学在宿主环境。为了解决这个问题，我们提出了以下目标：目标1。确定MK生物合成增强S.金黄色。目标2.定义了S.金黄色葡萄球菌减轻血红素毒性。先前的研究已经表明使用MK生物合成途径作为潜在的抗微生物靶标;然而，本申请中提供的证据表明，MK途径的破坏可能选择性地诱导形成持续性和血红素耐药性感染。相反，S.金黄色葡萄球菌可能代表更可行的药物靶点。通过对S.金黄色葡萄球菌和其他病原体，如炭疽杆菌，白喉棒状杆菌，单核细胞增生李斯特菌，我们将提供潜在的洞察力，有针对性的治疗设计，利用这些生物体的血红素敏感性。 公共卫生相关性：金黄色葡萄球菌是一个全球性的威胁人类健康，因为它能够引起许多疾病和多药耐药菌株正在迅速出现。我们的研究将进一步加深对S.金黄色葡萄球菌，一种可能在人类感染期间经历的病症，并定义了这种细菌用于科普血红素毒性的机制。从这些研究中获得的结果可能会提供洞察如何血红素敏感性的S。金黄色葡萄球菌可以被用作药物靶标。

DESCRIPTION (provided by applicant): Staphylococcus aureus is a human pathogen known to infect virtually every organ and cause numerous diseases. Antibiotic-resistant S. aureus strains are rapidly emerging; therefore, new drug targets are being sought. One of the mechanisms utilized by the host to restrict the growth of invading pathogens is the sequestration of the essential nutrient iron within the tetrapyrrole molecule heme. S. aureus employs sophisticated heme uptake systems in order to gain access to this iron source; however, the presence of excess heme is highly toxic to this and many other organisms. The nature of heme toxicity is not fully characterized but is thought to be, at least in part, due to oxidative damage induced by the reactive properties of the iron atom within the heme molecule. S. aureus possesses a heme detoxification system, a predicted transporter named HrtAB, although little is known about its mechanism of action. In the preliminary data presented in this application, we performed a transposon screen in the background of the heme-susceptible hrtA mutant, screening for strains with increased heme-resistance. All mutants identified targeted the menaquinone (MK) biosynthesis pathway. These data suggest that a component of the MK biosynthesis pathway potentiates heme stress. Interestingly, S. aureus strains deficient in MK biosynthesis, known as small colony variants (SCV), are commonly isolated from patients experiencing persistent infections. While multiple factors may contribute to the selection for MK-deficient S. aureus strains, it seems likely that resistance to the toxic effects of heme, an abundant molecule within the vertebrate host, could be playing a role. Based on these data, we predict that conditions of heme stress are directly relevant to the environment experienced by S. aureus during human infection. Therefore, we propose that a thorough understanding of the sources of heme stress in S. aureus and the mechanisms utilized by this pathogen to overcome the toxic effects of heme is essential to advancing our knowledge of staphylococcal physiology within the host environment. To address this issue we propose the following Aims: Aim 1. Determine the mechanism by which MK biosynthesis potentiates heme stress in S. aureus. Aim 2. Define the mechanism of the HrtAB system utilized by S. aureus to alleviate heme toxicity. Previous studies have suggested the use of the MK biosynthesis pathway as a potential antimicrobial target; however, evidence presented in this application indicates that disruption of the MK pathway might selectively induce the formation of persistent and heme-resistant infections. Instead, the heme detoxification system of S. aureus might represent a more viable drug target. By defining the regulation and function of the HrtAB heme detoxification system found in S. aureus and other pathogens such as Bacillus anthracis, Corynebacterium diphtheriae, and Listeria monocytogenes, we will provide potential insight into targeted therapeutic design exploiting the heme-susceptibility of these organisms. PUBLIC HEALTH RELEVANCE: Staphylococcus aureus is a global threat to human health because it is capable of causing numerous diseases and multi-drug resistant strains are rapidly emerging. Our proposed studies will further the understanding of heme stress in S. aureus, a condition likely experienced during human infection, and define the mechanisms utilized by this bacterium to cope with heme toxicity. Results obtained from these studies may provide insight into how the heme-susceptibility of S. aureus can be exploited as a drug target.