How Listeria senses and responds to different host environments

项目摘要/摘要 单核细胞增多性李斯特菌是一种兼性的细胞内食物传播的病原体，它提供了一种极其 宿主-病原菌相互作用基础研究的服从模型。这项建议是基于一项 揭示细菌氧化还原感应在毒力基因上调中的关键作用的遗传筛选 在体内表达。我们发现单核细胞增多性乳杆菌谷胱甘肽合成酶在感染过程中上调。 并确定谷胱甘肽是主要毒力转录因子PrFA的变构激活剂。 基于这些发现，经过多个小组数十年的调查，我们成功地 通过在生长的细菌中简单地添加还原剂来概括细胞内毒力基因的表达 一种化学定义的合成介质。这一发现导致了另一种识别细菌突变的基因筛查 它们对高浓度的生长抑制特性更敏感或更耐受 还原剂。对从这个筛选中产生的抗性突变体的分析导致发现，L. 单核细胞增多症拥有一条以黄素为基础的胞外电子传输链，该链可以运输 两种胞外脂蛋白上存在着超过10万个电子/细菌/秒的黄素部分。 和FrdA，它可以通过激活FrdA将电子转移到铁和富马酸，它编码一种 富马酸还原酶。我们证明，无论使用铁离子还是铁离子，舰船都可以介导厌氧生长。 富马酸作为电子受体。对还原剂更敏感的突变体聚集在PERR，a 氧化还原感应转录因子，控制对氧化应激的转录反应。我们 假设氧化应激是由我们显示的正在产生超氧化物的舰队产生的 氧气的存在。在包括病原体在内的100种菲米库特物种中发现了大量的同源基因 和微生物区系的成员，我们认为它代表了一个多功能的电子转移中心 在各种革兰氏阳性细菌中，这为厌氧生长的细菌提供了选择性优势 肠子。我们还提出，舰队的有氧活动还有其他后果，可能会 对感染、毒力和免疫的细胞生物学产生深远影响。

Project Summary/Abstract Listeria monocytogenes is facultative intracellular food-borne pathogen that provides an extremely amenable model for basic studies on host-pathogen interactions. This proposal is based on the results of a genetic screen that revealed a critical role of bacterial redox sensing for the upregulation of virulence gene expression in vivo. We found that L. monocytogenes glutathione synthase was upregulated during infection and determined that glutathione is the allosteric activator of the major virulence transcription factor PrfA. Based on these findings, and after decades of investigation by multiple groups, we successfully recapitulated intracellular virulence gene expression by simply adding reducing agents to bacteria grown in a chemically defined synthetic media. This finding led to another genetic screen to identify bacterial mutants that were either more sensitive or resistant to the growth inhibitory property of high concentrations of reducing agents. Analysis of the resistant mutants that arose from this screen led to the discovery that L. monocytogenes possess a Flavin-based Extracellular Electron Transport chain (FLEET) that can transport over 100,000 electrons/bacterium/second to flavin moieties present on two extracellular lipoproteins, PplA and FrdA, which can transfer electrons to ferric iron and fumarate via activation of FrdA, which encodes a fumarate reductase. We show that FLEET can mediated anaerobic growth using either ferric iron or fumarate as electron acceptors. Mutants that were more sensitive to reducing agents clustered in perR, a redox-sensing transcription factor that controls the transcriptional response to oxidative stress. We hypothesize that the oxidative stress is being generated by FLEET that we show is producing superoxide in the presence of oxygen. FLEET orthologues were found in 100s of Firmicute species including pathogens and members of the microbiota and we propose that it represents a versatile electron transfer hub present in diverse Gram-positive bacteria, that provides a selective advantage to bacteria growing anaerobically in the intestine. We also propose that FLEET activity has other consequences aerobically that may have profound effects on the cell biology of infection, virulence, and immunity.