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Identification of the EmrR regulon in Chromobacterium violaceum

ABSTRACT: Antibiotic resistance can arise by several mechanisms, including mutation in transcription factors that regulate drug efflux pumps. In this work, we identified EmrR as a MarR family transcription factor involved in antibiotic resistance in Chromobacterium violaceum, a Gram-negative bacterium that occurs in soil and water and can act as a human opportunistic pathogen. Antibiogram and minimum inhibitory concentration (MIC) assays showed that the ΔemrR mutant presented increased resistance to the antibiotic nalidixic acid in respect to the wild-type strain. The emrR gene is near to a putative operon emrCAB, which encode the efflux pump EmrCAB. DNA Microarray analysis showed that EmrR represses the emrCAB operon and some other putative transporters. Northern blot assays validated that EmrR represses the emrCAB operon and this repression can be released by salicylate, but not other compounds such as nalidixic acid or ethidium bromide. Electrophoretic mobility shift assays (EMSA) showed that EmrR binds directly to the promoter regions of emrR, emrCAB and other genes to exert negative regulation. Therefore, in response to compounds as salicylate, EmrR derepresses the operon emrCAB causing overexpression of the efflux pump EmrCAB and increased resistance to nalidixic acid in C. violaceum.

ORGANISM(S): Chromobacterium violaceum Chromobacterium violaceum ATCC 12472

PROVIDER: GSE112521 | GEO | 2018/11/01

REPOSITORIES: GEO

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Project description:Indole is ubiquitously synthesized by plants and bacteria and functions as an inter species signaling molecule to modulate a wide variety of cellular activities. However, it is not clear how the indole signal is perceived and responded by plant growth promoting rhizobacteria (PGPR) at the rhizosphere. Here, we demonstrated that indole enhanced antibiotic tolerance of Pseudomonas fluorescens 2P24, a PGPR well known for its biocontrol capacity. By conducting quantitative proteomic analysis, we showed that indole influences the expression of multiple genes including the emhABC operon encoding the major multidrug efflux pump in P. fluorescens 2P24. The indole-induced antibiotic tolerance was not related to bacterial dormancy or slow growth, but depended on the emhABC operon and the divergently transcribed TetR-like regulator emhR. By binding to the semi-palindromic operator sequence, EmhR repressed the expression of emhABC. It was further revealed that indole bound to EmhR and weakened the interaction between EmhR and the operator. This is consistent with our finding that indole-induced expression of the EmhABC efflux pump is dependent on EmhR. Using homology modeling and molecular dynamics simulation, we found that indole binding resulted in significantly decreased distance between the two DNA-recognizing α3 helices within the EmhR dimer, which would possibly account for its compromised DNA binding capacity. EmhR was further shown to globally influence protein expressions, especially transporters and proteins involved in the denitrification pathway. This EmhR-dependent, indole-induced antibiotic tolerance is likely to be prevalent in the Pseudomonas species, as the EmhR homologue in Pseudomonas syringae was also shown to be responsible for the indole-induced antibiotic tolerance. Taken together, our results revealed an indole-sensing transcription factor EmhR responsible for indole-induced antibiotic tolerance in Pseudomonas species and have important implications on the general mechanism for the indole sensing and responses in rhizobacteria.

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Identification of the EmrR regulon in Chromobacterium violaceum

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