Regulation of Listeria virulence gene expression by glutathione and hydrogen sulf

描述(申请人提供)：单核细胞增生性李斯特菌是一种革兰氏阳性兼性胞内细菌，能够入侵多种宿主细胞类型并进行复制 在细胞质中。介导侵袭、空泡逃逸和在宿主细胞质中存活的蛋白质都受正调控因子PrFA的转录控制。PRFA属于cAMP受体蛋白(CRP)转录因子家族，结合小分子辅因子后被激活。到目前为止，还没有确定与PrFA相关的辅助因子，尽管人们认为针对宿主细胞胞浆的信号(S)触发了PrFA的激活，导致了毒力基因的表达。我们进行了一次正向遗传筛选，以确定允许LM识别其胞浆定位并激活毒力基因转录的因素。我们的细胞质检测非常成功，因为我们发现了许多在依赖PRFA的基因调控中存在严重缺陷的基因，但以前从未与毒力相关。选择了两个转座子突变体进行进一步研究，每个转座子突变体在静脉感染的小鼠的脾和肝脏的定植中都显示出3-5个对数缺陷。本申请中详细介绍的实验旨在剖析这些突变如何调控李斯特菌的毒力基因表达。

DESCRIPTION (provided by applicant): Listeria monocytogenes is a Gram positive facultative intracellular bacterium that is capable of invading a wide range of host cell types and replicating in the cytosol. Proteins which mediate invasion, vacuolar escape, and survival in the host cytosol are all transcriptionally controlled by the positive regulatory factor PrfA. PrfA belongs t the cAMP receptor protein (Crp) family of transcription factors, which are activated upon binding small molecule cofactors. To date, a cofactor has not been identified for PrfA, although it is believed that a signal(s) specific to the host cell cytosol triggers PrfA activation, leading to virulence gene expression. We performed a forward genetic screen to identify factors that allow Lm to recognize its cytosolic localization and activate transcription of virulence genes. Our screen for 'cytosol sensing' was extremely successful, as we identified many genes that have severe defects in PrfA-dependent gene regulation, but have never before been associated with virulence. Two transposon mutants were chosen for further investigation, and each displayed a 3-5 log defect in colonization of the spleens and livers of intravenously infected mice. Experiments detailed in this application aim to dissect how these mutations regulate virulence gene expression in Listeria.