Laboratory of Microbiology and Infectious Diseases, Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka 567-047, Japan.
J Antimicrob Chemother. 2011 Feb;66(2):291-6. doi: 10.1093/jac/dkq420. Epub 2010 Nov 18.
Several putative and proven drug efflux pumps are present in Escherichia coli. Because many such efflux pumps have overlapping substrate spectra, it is intriguing that bacteria, with their economically organized genomes, harbour such large sets of multidrug efflux genes. To understand how bacteria utilize these multiple efflux pumps, it is important to elucidate the process of pump expression regulation. The aim of this study was to determine a regulator of the multidrug efflux pump in this organism.
We screened a genomic library of E. coli for genes that decreased drug susceptibility in this organism. The library was developed from the chromosomal DNA of the MG1655 strain, and then the recombinant plasmids were transformed into an acrB-deleted strain. Transformants were screened for resistance to various antibiotics including oxacillin.
We found that the multidrug susceptibilities of the acrB-deleted strain were decreased by the overexpression of small non-coding DsrA RNA as well as by the overexpression of known regulators of multidrug efflux pumps. Plasmids carrying the dsrA gene conferred resistance to oxacillin, cloxacillin, erythromycin, rhodamine 6G and novobiocin. DsrA decreased the accumulation of ethidium bromide in E. coli cells. Furthermore, expression of mdtE was significantly increased by dsrA overexpression, and the decreased multidrug susceptibilities modulated by DsrA were dependent on the MdtEF efflux pump.
These results indicate that DsrA modulates multidrug efflux through activation of genes encoding the MdtEF pump in E. coli.
大肠杆菌中存在几种假定的和已证实的药物外排泵。由于许多这样的外排泵具有重叠的底物谱，因此令人好奇的是，具有经济组织基因组的细菌却拥有如此大量的多药外排基因。为了了解细菌如何利用这些多种外排泵，阐明泵表达调控的过程很重要。本研究的目的是确定该生物体中多药外排泵的调节剂。
我们从大肠杆菌的基因组文库中筛选出降低该生物体药物敏感性的基因。该文库是从 MG1655 菌株的染色体 DNA 中开发的，然后将重组质粒转化到 acrB 缺失的菌株中。转化子通过对各种抗生素（包括苯唑西林）的抗性进行筛选。
我们发现，小非编码 DsrA RNA 的过表达以及已知多药外排泵调节剂的过表达降低了 acrB 缺失菌株的多药敏感性。携带 dsrA 基因的质粒赋予了苯唑西林、氯唑西林、红霉素、吖啶橙和新生霉素的抗性。DsrA 减少了大肠杆菌细胞中溴化乙锭的积累。此外，dsrA 过表达显著增加了 mdtE 的表达，dsrA 调节的降低多药敏感性依赖于 MdtEF 外排泵。
这些结果表明，DsrA 通过激活大肠杆菌中编码 MdtEF 泵的基因来调节多药外排。
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