Regulation of an antibiotic resistance efflux pump by quorum sensing and a TetR-family repressor inChromobacterium subtsugae

Abstract

AbstractThe soil bacteriumChromobacterium substugaeuses a single LuxI-R-type quorum-sensing system, CviI-R, to regulate genes in a cell density-dependent manner. CviI synthesizes the signalN-hexanoyl-homoserine lactone (C6-HSL) and CviR is a C6-HSL-responsive cytoplasmic transcription regulator. C6-HSL-bound CviR activates dozens of genes, for example thecdeAB-oprMcluster coding for an efflux pump conferring antibiotic resistance. ThecdeAB-oprMgenes are also regulated by an antibiotic-responsive transcription factor, CdeR, which represses expression of these genes. We are interested in understanding howC. subtsugaeintegrates different environmental cues to regulate antibiotic resistance. In this study, we sought to delineate the mechanism of regulation of thecdeAB-oprMgenes by CviR and CdeR. In recombinantE. coli,thecdeApromoter is activated by CviR and repressed by CdeR. We identify non-overlapping sequence elements in thecdeApromoter that are required for CviR activation and CdeR repression, respectively. We also examined the role of CdeR in modulatingcdeAactivation by C6-HSL inC. subtsugae. We show that CviR and CdeR can independently modulate transcription from thecdeApromoter inC. subtsugae, consistent with the conclusion that CviR and CdeR regulate thecdeAB-oprMgenes by interacting directly with different binding sites in thecdeApromoter. These results contribute to a molecular understanding of how thecdeAB-oprMgenes are regulated and provide new insight into howC. subtsugaeintegrates different environmental cues to regulate antibiotic resistance.ImportanceMany bacteria regulate antibiotic resistance in response to antibiotics and other cues from the environment. In many cases the regulatory mechanisms are best understood in the context of clinical isolates where mutations frequently emerge in resistance regulation pathways. However, an understanding of the role of antibiotic resistance regulators in integrating environmental information is less well understood. In the soil bacteriumChromobacterium subtsugae,an antibiotic-resistance gene cluster is regulated by population density and antibiotics through two different transcription factors; the quorum sensing signal receptor CviR and an antibiotic-responsive transcription factor CdeR. In this study, we show that these factors independently modulate the transcription of the antibiotic resistance genes and coordinate to ensure sensitive responses to changes in cell density. The results give new insight into antibiotic resistance regulation inC. subtsugaeand contribute to a broader understanding of how bacteria optimize the regulation of antibiotic resistance in response to changes in the environment.