RUI: Acid Signals and Bacterial Drug Efflux Systems

植物产生与阿司匹林相关的芳香酸，可能限制或逆转土壤环境和肠道微生物群中的细菌耐药性。暴露于水杨酸选择大肠杆菌菌株已失去多药耐药性。适应度成本增加的机制尚不清楚，但可能涉及细菌细胞质子动力的耗竭，质子动力是药物外排泵的能量来源。在这个项目中，本科生研究人员将剖析有机酸如何逆转多药耐药的机制。“一年级第一”项目每年招收8名本科生（3年内招收24名）进行研究，由首席研究员和一组高级本科生指导。该招聘计划旨在吸引经济背景有限的学生，并鼓励他们在STEM领域从事研究工作。流式细胞术竞争分析将揭示大肠杆菌在水杨酸盐或茉莉酸盐存在下哪些药物外排基因产生适应成本。利用药物泵阻滞剂检测质子动力的作用，降低质子驱动泵的适配成本。通过比较水杨酸盐和茉莉酸盐及其甲酯的作用，将测试特定酸阴离子的作用。通过与植物、土壤和人类微生物组相关的水杨酸依赖的细菌物种的实验进化，包括植物共生的荧光假单胞菌、土壤细菌枯草芽孢杆菌和土壤和肠道发酵剂粪肠球菌，将耐药性逆转的假设扩展到大肠杆菌之外。用乙醚标记细胞的FACS分选可以识别出失去多药泵的突变体。水杨酸进化克隆的基因组和转录组将揭示每个物种独特的调节途径的重新布线。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Plants produce aromatic acids related to aspirin that may limit or reverse bacterial drug resistance within the soil environment and gut microbiome. Exposure to salicylic acid selects for Escherichia coli strains that have lost multidrug resistance. The mechanism of the increased fitness cost is unclear, but may involve depletion of the bacterial cell's proton motive force, the energy source that powers drug efflux pumps. In this project, undergraduate researchers will dissect the mechanism of how organic acids reverse multidrug resistance. The "First-years First" program will recruit eight incoming undergraduate students annually (24 over three years) to conduct research, with supervision by the principal investigator and a team of advanced undergraduates. The recruitment program is designed to attract students from economically limited backgrounds and encourage them to pursue research careers in STEM.Competition assays by flow cytometry will reveal which drug-efflux genes of E. coli incur fitness cost in the presence of salicylate or jasmonate. The role of proton motive force will be tested by using drug pump blockers, which may decrease the fitness cost of the proton-driven pumps. The role of specific acid anions will be tested by comparing the effects of salicylate and jasmonate with that of their methyl esters. The hypothesis of drug resistance reversal will be extended beyond E. coli by salicylate-dependent experimental evolution of bacterial species relevant to plant, soil and human microbiomes, including the plant commensal Pseudomonas fluorescens, the soil bacterium Bacillus subtilis, and the soil and gut fermenter Enterococcus faecalis. FACS sorting of ethidium-labeled cells will identify mutants that lose multidrug pumps. Genomes and transcriptomes of salicylate-evolved clones will reveal the rewiring of regulatory pathways unique to each species.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.