The Role of Lactate in the ???Metabolic Evolution??? of Staphylococcus aureus

描述(申请人提供)：金黄色葡萄球菌几乎能够抵抗宿主先天免疫的方方面面，包括产生一氧化氮(No.)不耐药在其他细菌中不明显，包括密切相关的凝固酶阴性葡萄球菌(CNS)。我们已经发现，金黄色葡萄球菌对NO产生抵抗力的关键是这种微生物唤起以乳酸代谢为中心的代谢状态的能力，这种代谢状态本质上是对NO免疫的。金黄色葡萄球菌含有三种乳酸脱氢酶，其中一种(Ldh1)在中枢神经系统中不存在，在金黄色葡萄球菌耐药中起着关键作用。这项建议的重点是了解乳酸代谢在金黄色葡萄球菌毒力和免疫逃避中的作用。具体目标1.建立Lqo作为NOX循环的一部分，NOX循环是金黄色葡萄球菌毒力的重要途径。我们鉴定了编码金黄色葡萄球菌乳酸的一个基因：苯醌氧化还原酶(Lqo)，并证明了它在毒力中的作用。在这里，我们测试解释为什么金黄色葡萄球菌中的第四种乳酸利用酶对发病如此关键的假说。我们将建立NOX循环，这是一条依赖Ldh1和Lqo的代谢途径，实际上允许金黄色葡萄球菌使用宿主NO。用于能源生产。我们还将对Lqo进行结构和生化表征，Lqo是只在葡萄球菌中发现的一种新的酶家族的创始人。具体目的2.揭示乳酸代谢在金黄色葡萄球菌发病机制中的多重作用。与中枢神经系统不同，金黄色葡萄球菌在非应激条件下表现出对乳酸的“对映体偏好”，并产生大量的L异构体，这是乳酸参与氮氧化物循环的唯一形式。我们将确定导致ldh1峰值表达的调控机制，ldh1是金黄色葡萄球菌特有的乳酸脱氢酶，负责“对映体偏好”。我们还将测试旨在解释奇怪的观察结果的假设，即L-乳酸和D-乳酸的产生都是金黄色葡萄球菌毒力所必需的。我们观察到，参与金黄色葡萄球菌乳酸代谢的基因在中枢神经系统中要么不存在，要么存在差异调节。因此，金黄色葡萄球菌乳酸盐代谢相关基因的新获得和调控的改变为新出现的病原体的“代谢进化”提供了一个模型。这些渐进的进化变化为金黄色葡萄球菌提供了在具有免疫能力的宿主中茁壮成长所需的代谢灵活性，并且必须与该病原体出现的典型毒力因子的获得相吻合。

DESCRIPTION (provided by applicant): S. aureus is capable of withstanding nearly every facet of host innate immunity, including the production of nitric oxide (NO.) NO.-resistance is not apparent in other bacteria, including closely related coagulase negative staphylococci (CNS). We have found that the key to S. aureus NO.-resistance is the ability of this organism to evoke a metabolic state, centered around lactate metabolism, that is essentially immune to NO. S. aureus harbors three lactate dehydrogenases, one of which (Ldh1) is not present in CNS and plays a critical role in S. aureus NO.-resistance. The focus of this proposal is to understand the role of lactate-metabolism in S. aureus virulence and immune evasion. Specific Aim 1. Establish Lqo as part of the NOX-cycle, an essential pathway for S. aureus virulence. We identified a gene encoding the S. aureus lacate:quinone oxidoreductase (Lqo) and demonstrated its role in virulence. Here we test hypotheses explaining why a fourth lactate-utilizing enzyme in S. aureus is so critical to pathogenesis. We will establish the NOX cycle, a metabolic pathway relying on both Ldh1 and Lqo that actually allows S. aureus to use host NO. for energy production. We will also structurally and biochemically characterize Lqo, the founder of a new family of enzymes found only among the staphylococci. Specific Aim 2. Unravel the multiple roles for lactate metabolism in S. aureus pathogenesis. Unlike CNS, S. aureus exhibits lactate "enantiomer preference" during NO.-stress and produces copious amounts of the L-isomer, the only form of lactate able to participate in the NOX cycle. We will define the regulatory mechanisms resulting in peak expression of ldh1, the S. aureus-specific lactate dehydrogenase responsible for "enantiomer preference". We will also test hypotheses aimed at explaining the curious observation that both L- and D-lactate production are required for S. aureus virulence. We have observed that genes involved in S. aureus lactate metabolism are either not present or are differentially regulated in CNS. Thus, the new acquisition and altered regulation of genes involved in S. aureus lactate-metabolism provides a model for the "Metabolic Evolution" of an emerging pathogen. These incremental evolutionary changes provided S. aureus with the metabolic flexibility necessary to thrive within an immunocompetent host and must have coincided with the acquisition of canonical virulence factors for this pathogen to emerge.