Mechanisms of antibiotic efflux in Campylobacter

细菌抗生素外排转运体是导致固有和获得性耐药的重要因素 为了抗菌剂。空肠弯曲菌是美国食源性腹泻的主要细菌来源 和包括在NIAID B类优先病原体名单中的一种制剂，含有多个抗生素外排 不同家族的运输者。在之前的授权期内，我们确定了功能和规则 阻力结瘤分割器(RND)家族的两个外排泵(CmeABC和CmeDEF) 弯曲杆菌。我们的发现表明，外排系统不仅有助于产生抗菌素耐药性， 而且在促进弯曲杆菌在肠道内定植方面也具有重要的生理功能 关于动物的。我们还发现，CmeR是一种转录因子，它抑制cmeABC和胆盐 (通常存在于肠道)通过抑制CmeR与CmeABC的结合来诱导cmeABC的表达 CmeABC的启动子。我们最近的初步研究也有力地表明CmeR是一种多效性 调节和调节Mf(主要促进剂)和Mate(多药和有毒)的表达 复合挤压)转运体以及C4-二羧酸转运体可能参与 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方面的作用。 尽管最近取得了这些进展，但弯曲杆菌中大多数由CmeR调控的外排转运蛋白 还没有被功能表征，并且控制基因表达的分子机制 转运体和CmeR调控的结构基础仍有待确定。要结束这些重要的 我们对弯曲杆菌主动外排系统的了解存在差距，我们计划在以下三个方面实现具体目标 此续签申请用于1)确定MF和Mate的监管机制和功能 空肠弯曲菌中的转运蛋白，2)C4-二羧酸转运系统在体内的调节和作用 促进弯曲杆菌对缺氧条件的适应，以及3)阐明结构 用X射线结晶学研究CmeR调节的基础和胆汁诱导的机制。这个 建议的研究利用我们实验室现有的独特资源，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， 拟议的工作与前一批赠款期间进行的研究将揭示出新的 关于细菌中抗生素外排转运体的功能和调节机制的信息。这个 这些发现将有助于确定控制和治疗抗生素耐药性的潜在分子靶点。 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.