Mechanisms of antibiotic efflux in Campylobacter

细菌抗生素外排转运蛋白在赋予内在和获得性耐药性方面发挥着重要作用 到抗菌剂。空肠弯曲菌，美国食源性腹泻的主要原因 以及 NIAID B 类优先病原体清单中包含的一种病原体，具有多种抗生素外流 不同家庭的运输者。在上一次资助期间，我们确定了功能和规定 抗结瘤分裂 (RND) 系列的两个外排泵（CmeABC 和 CmeDEF） 弯曲杆菌。我们的研究结果表明，外排系统不仅会导致抗菌素耐药性， 而且还具有促进弯曲杆菌在肠道定植的重要生理功能 的动物。我们还发现转录因子 CmeR 抑制 cmeABC，并且胆汁盐 （通常存在于肠道中）通过抑制 CmeR 与 cmeABC 的发起人。我们最近的初步研究也强烈表明 CmeR 是一种多效性 调节因子并调节 MF（主要促进因子）和 MATE（多药和毒性因子）的表达 复合挤压）转运蛋白以及可能参与的 C4-二羧酸转运蛋白 弯曲杆菌对动物肠道缺氧环境的适应。这些发现表明 抗生素外排系统由 CmeR 与其他生理过程共同调节，发挥着重要作用 在抗菌素耐药性和促进弯曲杆菌适应环境变化中发挥作用。 尽管最近取得了这些进展，弯曲杆菌中大多数 CmeR 调节的外排转运蛋白 尚未进行功能表征，并且控制表达的分子机制 转运蛋白和 CmeR 调节的结构基础仍有待确定。关闭这些重要的 由于我们对弯曲杆菌主动外排系统的理解存在差距，我们计划实现 3 个具体目标 本次更新申请的目的是 1) 确定 MF 和 MATE 的监管机制和职能 空肠弯曲菌中的转运蛋白，2) 定义了 C4-二羧酸盐转运系统的调节和作用 促进弯曲杆菌适应限氧条件，3）阐明其结构 使用 X 射线晶体学研究 CmeR 调节和胆汁诱导机制的基础。的 拟议的研究利用我们实验室可用的独特资源并利用当代 分子、遗传和生化方法以及已建立的动物模型。一旦完成， 拟议的工作与上一个资助期间进行的研究一起将揭示新的 有关细菌中抗生素外排转运蛋白的功能和调节机制的信息。的 研究结果将有助于确定控制和治疗抗生素耐药性的潜在分子靶标 弯曲杆菌。

Bacterial antibiotic efflux transporters are important players in conferring intrinsic and acquired resistance to antimicrobials. Campylobacterjejuni, a leading bacterial cause of foodborne diarrhea in the United States and an agent included in the NIAID Category B Priority Pathogens list, harbors multiple antibiotic efflux transporters of different families. During the previous grant period, we determined the function and regulation of two efflux pumps (CmeABC and CmeDEF) of the resistance-nodulation-division (RND) family in Campylobacter. Our findings indicate that the efflux system not only contributes to antimicrobial resistance, but also has important physiological functions in facilitating Campylobacter colonization in the intestinal tract of animals. We have also found that CmeR, a transcriptional factor, represses cmeABC and that bile salts (normally present in the gut) induce the expression of cmeABC by inhibiting the binding of CmeR to the promoter of cmeABC. Our recent preliminary studies also strongly suggest that CmeR is a pleiotropic regulator and modulates the expression of the MF (major facilitator) and MATE (multidrug and toxic compound extrusion) transporters as well as the C4-dicarboxylate transporters potentially involved in Campylobacter adaptation to the oxygen-limited environment in animal intestine. These findings indicate that the antibiotic efflux system is co-regulated by CmeR with other physiological processes and plays important roles in antimicrobial resistance and in facilitating Campylobacter adaptation to environmental changes. Despite these recent advances, the majority of the CmeR-regulated efflux transporters in Campylobacter have not been functionally characterized, and the molecular mechanisms governing the expression of the transporters and the structural basis of CmeR regulation remain to be determined. To close these important gaps in our understanding of the active efflux system in Campylobacter, we plan to pursue 3 specific aims in this renewal application to 1) determine the regulatory mechanisms and functions of the MF and MATE transporters in C. jejuni, 2) define the regulation and role of the C4-dicarboxylate transport system in facilitating Campylobacter adaptation to oxygen-limited conditions, and 3) elucidate the structural basis of CmeR regulation and the mechanisms of bile induction using X-ray crystallography. The proposed studies take advantage of unique resources available in our laboratories and utilize contemporary molecular, genetic, and biochemical approaches as well as an established animal model. Once completed, the proposed work together with the studies conducted in the previous grant period will reveal novel information on the functions and regulatory mechanisms of antibiotic efflux transporters in bacteria. The findings will help to identify potential molecular targets for the control and treatment of antibiotic resistant Campylobacter.