Coordinate Regulation of Salmonella Virulence and Antimicrobial Resistance by MarR Transcription Factors

摘要马尔R（Multiple antibiotic resistance Repressor，多抗生素耐药抑制因子）蛋白包括转录的古老家族 在细菌中广泛保守的因子。在肠道病原体鼠伤寒沙门氏菌中，马尔R是一种 AcrAB-TolC药物外排泵的负调节因子，而另一种称为SlyA的马尔R转录因子是一种 水平获得的毒力基因表达所需的反沉默子。我们最近的研究表明 尽管其功能不同，但SlyA与马尔R共享通过芳香族化合物进行变构调节的能力， 羧酸分子，并可通过抑制marRAB操纵子影响mar表型。失活 TolC是AcrAB和其他RND家族外排泵的重要组成部分，表型为slyA突变， 减少沙门氏菌毒力基因的表达。这表明，外排调节的水平， 与SlyA相互作用的内源性代谢产物，进而控制马尔R的表达，完成调节 协调连接抗菌素耐药性、毒力和细菌代谢的回路。 该提案调查了沙门氏菌抗菌素耐药性和毒力是 通过马尔R和SlyA转录因子响应代谢信号而协同调节 具体目标如下： (1)鉴定调节马尔R/SlyA活性的内源性配体。初步数据表明 SlyA相互作用配体是芳族羧酸酯。特异性内源性配体将通过遗传学方法鉴定。 和代谢组学方法，并在亲和力和变构抑制方面进行表征。 (2)沙门氏菌抗菌肽中MarR-SlyA-TolC调控相互作用的功能表征 抗性和毒力。MarR-SlyA调控网络对沙门氏菌表型的贡献 将在体外和体内评估抗微生物剂耐药性和毒力。 (3)马尔R和SlyA调节子的比较分析。马尔/SlyA调节的基因和结合位点将 全面识别，以确定抑制和反沉默的监管要求， 并阐明转录网络进化的机制。 这些研究将为研究细菌毒力、药物敏感性和耐药性之间的中央调控网络提供重要的见解。 耐药性和代谢，这可以导致新的治疗靶点的预防或 治疗细菌感染。

SUMMARY. MarR (Multiple antibiotic resistance Repressor) proteins comprise an ancient family of transcription factors that are widely conserved in bacteria. In the enteric pathogen Salmonella Typhimurium, MarR is a negative regulator of the AcrAB-TolC drug efflux pump, while another MarR transcription factor called SlyA is a counter-silencer required for the expression of horizontally-acquired virulence genes. We have recently shown that despite its divergent function, SlyA shares with MarR the ability to undergo allosteric modulation by aromatic carboxylate molecules and can influence the mar phenotype by repressing the marRAB operon. Inactivation of TolC, an essential component of the AcrAB and other RND-family efflux pumps, phenocopies a slyA mutation, reducing the expression of Salmonella virulence genes. This suggests that efflux regulates the levels of an endogenous metabolite that interacts with SlyA, which in turn controls MarR expression, completing a regulatory circuit that coordinately links antimicrobial resistance, virulence, and bacterial metabolism. This proposal investigates the hypothesis that Salmonella antimicrobial resistance and virulence are coordinately regulated in response to metabolic signals by the MarR and SlyA transcription factors through the following specific aims: (1) Identification of endogenous ligand(s) that modulate MarR/SlyA activity. Preliminary data indicate that SlyA-interacting ligand(s) are aromatic carboxylates. Specific endogenous ligands will be identified by genetic and metabolomic methods and characterized with regard to affinity and allosteric inhibition. (2) Functional characterization of MarR-SlyA-TolC regulatory interactions in Salmonella antimicrobial resistance and virulence. The contribution of the MarR-SlyA regulatory network to Salmonella phenotypic antimicrobial resistance and virulence will be assessed in vitro and in vivo. (3) Comparative analysis of the MarR and SlyA regulons. MarR/SlyA-regulated genes and binding sites will be comprehensively identified to determine the regulatory requirements for repression and counter-silencing, respectively, and to elucidate the mechanisms of transcriptional network evolution. These studies will provide important insights into a central regulatory network linking bacterial virulence, drug resistance, and metabolism, which can lead to the identification of new therapeutic targets for the prevention or treatment of bacterial infections.