Mechanistic evaluation of resistance to sulfite toxicity in Salmonella

项目总结 亚硫酸盐是一种活性硫化物，在所有生命形式中普遍存在。它们是硫酸盐化的副产品 氨基酸代谢，几十年来一直被用来限制食品中的细菌生长。因为 亚硫酸盐可以与蛋白质、DNA和脂质反应，扰乱细胞过程，亚硫酸盐的解毒是必不可少的。 为了细胞的生存能力。有趣的是，亚硫酸盐是由激活的中性粒细胞对细菌感染做出反应而产生的， 这表明它们可能在宿主与病原菌的相互作用中发挥重要作用。这项工作的目的是 以肠道沙门氏菌为模型，确定亚硫酸盐对细菌感染结局的影响 病原体。肠道沙门氏菌是细菌性食源性胃肠炎的主要原因，也是最主要的原因。 在美国死于食源性疾病的人数。在我们发表的作品中，我们描述了一个 转录调控因子YeiE在沙门氏菌致病中的作用。我们证明了YIE对沙门氏菌的关键作用 在肠道中定居，因为它调节鞭毛的产生，鞭毛是病毒的关键毒力因素 肠源性沙门氏菌病。最近对Skazakii慢杆菌的研究表明，YeiE感觉到过量的亚硫酸盐 调节亚硫酸盐还原酶的表达，是中性粒细胞杀伤生存所必需的。Yeie同系物存在 在许多细菌病原体中，表明Yeie在寄主病原菌中对亚硫酸盐的感知具有重要作用 互动。我们假设YeiE调节亚硫酸盐应激反应，允许沙门氏菌 抵抗先天免疫控制。在目标1中，我们将确定亚硫酸盐还原在沙门氏菌病中的作用。我们 将建立YEIE调节的亚硫酸盐还原对小鼠胃肠炎感染模型的影响 败血症。由于激活的中性粒细胞产生亚硫酸盐和中性粒细胞炎症是宿主控制 对于沙门氏菌，重要的是要了解YeiE是如何在宿主来源的亚硫酸盐压力下调节生存的。在……里面 目的2，我们将建立叶娥对亚硫酸盐毒性抗性的调控机制。我们将建立 用染色质免疫沉淀-测序技术研究亚硫酸盐结合的YEIE调控基因 利用高密度转座子突变文库，获得了抵御亚硫酸盐胁迫所需的基因。组合分析 这两种不偏不倚的方法将使我们能够确定耶伊如何协调细菌对 亚硫酸盐胁迫。Yeie同源物分布在许多不同的细菌病原体中，所以我们的工作将有 应用于广泛的宿主-病原体相互作用。这项工作的成功完成将证明 细菌如何在体内抵抗亚硫酸盐应激，并将导致未来的工作，以研究亚硫酸盐作为交叉 王国信号分子及其对细菌感染免疫反应的影响。

PROJECT SUMMARY Sulfites are reactive sulfur species with ubiquitous distribution in all life forms. They are byproducts of sulfated amino acid metabolism and have been used for decades to limit bacterial growth in food products. Because sulfites can react with proteins, DNA, and lipids to disrupt cellular processes, detoxification of sulfites is essential for cellular viability. Interestingly, sulfites are produced by activated neutrophils in response to bacterial infection, indicating they may play an important role in host-pathogen interactions. The purpose of this work is to establish how sulfites impact the outcome of bacterial infection, using Salmonella enterica as a model pathogen. Salmonella enterica is a leading cause of bacterial foodborne gastroenteritis and is the leading cause of death from foodborne disease in the United States. In our published work, we characterized the role of a transcriptional regulator, YeiE, in Salmonella pathogenesis. We demonstrated that yeiE is critical for Salmonella to colonize the intestine because it regulates production of flagella, which are a key virulence factor required for enteric salmonellosis. Recent work in Cronobacter sakazakii demonstrated that YeiE senses excess sulfite to regulate expression of a sulfite reductase and is needed to survive neutrophil killing. YeiE homologs are present in many bacterial pathogens, suggesting an important role for sulfite sensing by YeiE in host-pathogen interactions. We hypothesize that YeiE regulates the sulfite stress response to allow Salmonella to withstand innate immune control. In Aim 1, we will determine the role of sulfite reduction in salmonellosis. We will establish the impact of YeiE-regulated sulfite reduction in murine infection models of gastroenteritis and sepsis. Since activated neutrophils produce sulfite and neutrophilic inflammation is a key to host control of Salmonella, it is important to understand how YeiE mediates survival in the face of host-derived sulfite stress. In Aim 2, we will establish the mechanism by which YeiE regulates resistance to sulfite toxicity. We will establish the genes regulated by sulfite-bound YeiE using chromatin immunoprecipitation-sequencing and will establish the genes needed to withstand sulfite stress using a high-density transposon mutant library. Combined analysis of these two unbiased approaches will allow us to establish how YeiE coordinates the bacterial response to sulfite stress. YeiE homologs are distributed amongst many diverse bacterial pathogens, so our work will have application to a broad range of host-pathogen interactions. Successful completion of this work will demonstrate how bacteria resist sulfite stress in vivo and will lead to future work to investigate the role of sulfite as a cross- kingdom signaling molecule and its impact on the immune response to bacterial infections.