ABSTRACT Drug repurposing efforts led to the discovery of bactericidal activity in auranofin, a gold-containing drug used to treat rheumatoid arthritis. Auranofin kills Gram-positive bacteria by inhibiting thioredoxin reductase, an enzyme that scavenges reactive oxygen species (ROS). Despite the presence of thioredoxin reductase in Gram-negative bacteria, auranofin is not always active against them. It is not clear whether the lack of activity in several Gram-negative bacteria is due to the cell envelope barrier or the presence of other ROS protective enzymes such as glutathione reductase (GOR). We previously demonstrated that chemical analogs of auranofin (MS-40 and MS-40S), but not auranofin, are bactericidal against the Gram-negative Burkholderia cepacia complex. Here, we explore the targets of auranofin, MS-40, and MS-40S in Burkholderia cenocepacia and elucidate the mechanism of action of the auranofin analogs by a genome-wide, randomly barcoded transposon screen (BarSeq). Auranofin and its analogs inhibited the B. cenocepacia thioredoxin reductase and induced ROS but did not inhibit the bacterial GOR. Genome-wide, BarSeq analysis of cells exposed to MS-40 and MS-40S compared to the ROS inducers arsenic trioxide, diamide, hydrogen peroxide, and paraquat revealed common and unique mediators of drug susceptibility. Furthermore, deletions of gshA and gshB that encode enzymes in the glutathione biosynthetic pathway led to increased susceptibility to MS-40 and MS-40S. Overall, our data suggest that the auranofin analogs kill B. cenocepacia by inducing ROS through inhibition of thioredoxin reductase and that the glutathione system has a role in protecting B. cenocepacia against these ROS-inducing compounds. IMPORTANCE The Burkholderia cepacia complex is a group of multidrug-resistant bacteria that can cause infections in the lungs of people with the autosomal recessive disease, cystic fibrosis. Specifically, the bacterium Burkholderia cenocepacia can cause severe infections, reducing lung function and leading to a devastating type of sepsis, cepacia syndrome. This bacterium currently does not have an accepted antibiotic treatment plan because of the wide range of antibiotic resistance. Here, we further the research on auranofin analogs as antimicrobials by finding the mechanism of action of these potent bactericidal compounds, using a powerful technique called BarSeq, to find the global response of the cell when exposed to an antimicrobial.

中文翻译：

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化学基因组分析揭示了金诺芬类似物在新洋葱伯克虫中的作用机制


摘要药物再利用的努力发现了金诺芬（一种用于治疗类风湿性关节炎的含金药物）的杀菌活性。金诺芬通过抑制硫氧还蛋白还原酶（一种清除活性氧 (ROS) 的酶）来杀死革兰氏阳性菌。尽管革兰氏阴性细菌中存在硫氧还蛋白还原酶，但金诺芬并不总是对它们有活性。目前尚不清楚几种革兰氏阴性细菌缺乏活性是否是由于细胞包膜屏障或其他 ROS 保护酶（例如谷胱甘肽还原酶 (GOR)）的存在所致。我们之前证明金诺芬的化学类似物（MS-40 和 MS-40S）（但不是金诺芬）对革兰氏阴性菌具有杀菌作用。洋葱伯克霍尔德菌复杂的。在这里，我们探讨了金诺芬、MS-40 和 MS-40S 在新洋葱伯克霍尔德杆菌并通过全基因组随机条形码转座子筛选 (BarSeq) 阐明金诺芬类似物的作用机制。金诺芬及其类似物抑制B. 新洋葱硫氧还蛋白还原酶并诱导 ROS，但不抑制细菌 GOR。对暴露于 MS-40 和 MS-40S 的细胞与 ROS 诱导剂三氧化二砷、二酰胺、过氧化氢和百草枯进行全基因组 BarSeq 分析，揭示了药物敏感性的常见和独特介质。此外，删除谷胱甘肽和谷胱甘肽编码谷胱甘肽生物合成途径中的酶的酶导致对 MS-40 和 MS-40S 的敏感性增加。总的来说，我们的数据表明金诺芬类似物会杀死B. 新洋葱通过抑制硫氧还蛋白还原酶诱导 ROS，谷胱甘肽系统具有保护作用B. 新洋葱对抗这些 ROS 诱导化合物。重要性 洋葱伯克霍尔德菌复合体是一组多重耐药细菌，可引起常染色体隐性遗传病（囊性纤维化）患者肺部感染。具体来说，新洋葱伯克霍尔德杆菌可引起严重感染，降低肺功能，并导致毁灭性的败血症、洋葱综合征。由于广泛的抗生素耐药性，这种细菌目前没有公认的抗生素治疗方案。在这里，我们通过使用名为 BarSeq 的强大技术来发现这些有效杀菌化合物的作用机制，进一步研究金诺芬类似物作为抗菌剂的作用，以发现细胞在接触抗菌剂时的整体反应。