Abstract
The pathway of copper entry into Escherichia coli is still unknown. In an attempt to shed light on this process, a lux-based biosensor was utilized to monitor intracellular copper levels in situ. From a transposon-mutagenized library, strains were selected in which copper entry into cells was reduced, apparent as clones with reduced luminescence when grown in the presence of copper (low-glowers). One low-glower had a transposon insertion in the comR gene, which encodes a TetR-like transcriptional regulator. The mutant strain could be complemented by the comR gene on a plasmid, restoring luminescence to wild-type levels. ComR did not regulate its own expression, but was required for copper-induction of the neighboring, divergently transcribed comC gene, as shown by real-time quantitative PCR and with a promoter-lux fusion. The purified ComR regulator bound to the promoter region of the comC gene in vitro and was released by copper. By membrane fractionation, ComC was shown to be localized in the outer membrane. When grown in the presence of copper, ∆comC cells had higher periplasmic and cytoplasmic copper levels, compared to the wild-type, as assessed by the activation of the periplasmic CusRS sensor and the cytoplasmic CueR sensor, respectively. Thus, ComC is an outer membrane protein which lowers the permeability of the outer membrane to copper. The expression of ComC is controlled by ComR, a novel, TetR-like copper-responsive repressor.
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Acknowledgments
We thank Thomas Weber for expert technical assistance and the National BioResource Project (NIG, Japan) for providing strain JD27538. This work was supported by Grant 3100A0_122551 from the Swiss National Foundation and grants from the International Copper Association and the Swiss State Secretary for Education & Research.
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Mélanie Mermod and David Magnani have contributed equally to this work
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Mermod, M., Magnani, D., Solioz, M. et al. The copper-inducible ComR (YcfQ) repressor regulates expression of ComC (YcfR), which affects copper permeability of the outer membrane of Escherichia coli . Biometals 25, 33–43 (2012). https://doi.org/10.1007/s10534-011-9510-x
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DOI: https://doi.org/10.1007/s10534-011-9510-x