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

总结。MARR(多重抗生素耐药抑制因子)蛋白组成一个古老的转录家族 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，MARR是一种 AcrAB-TolC药物外排泵的负调控因子，而另一种称为SlyA的Marr转录因子是一种 水平获得的毒力基因表达所需的反沉默因子。我们最近展示了 尽管SlyA具有不同的功能，但它与Marr一样具有通过芳香族化合物进行变构调节的能力 并且可以通过抑制marrab操纵子来影响mar的表型。停用 TolC是AcrAB和其他RND家族外排泵的重要组成部分，它的表型复制了slyA突变， 降低沙门氏菌毒力基因的表达。这表明，外排调节着一种 与SlyA相互作用的内源性代谢物，SlyA反过来控制MarR的表达，完成一种调节 协调连接抗菌素耐药性、毒性和细菌新陈代谢的回路。 这项建议调查了沙门氏菌抗菌素耐药性和毒力是 MarR和SlyA转录因子对代谢信号的协同调控 通过以下具体目标： (1)调控MarR/SlyA活性的内源配体(S)的鉴定。初步数据显示， SlyA相互作用配体(S)是芳香族羧酸盐。特定的内源性配体将通过基因识别 和代谢组学方法，并以亲和力和变构抑制为特征。 (2)沙门氏菌抗菌药物中Marr-SlyA-TolC调控相互作用的功能特征 抗药性和毒性。Marr-SlyA调控网络对沙门氏菌表型的影响 抗菌素耐药性和毒力将在体外和体内进行评估。 (3)MARR和SlyA启动子的比较分析。MarR/SlyA调节的基因和结合位点将 全面确定压制和反沉默的监管要求， 并阐明转录网络进化的机制。 这些研究将为联系细菌毒性、药物的中央调控网络提供重要的见解 抵抗力和新陈代谢，这可以导致确定新的治疗靶点，以预防或 细菌感染的治疗。

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.