The ratio between CcdA and CcdB modulates the transcriptional repression of the ccd poison-antidote system.
The ccd operon of the F plasmid encodes CcdB, a toxin targeting the essential gyrase of Escherichia coli, and CcdA, the unstable antidote that interacts with CcdB to neutralize its toxicity. Although work from our group and others has established that CcdA and CcdB are required for transcriptional repression of the operon, the underlying mechanism remains unclear. The results presented here indicate that, although CcdA is the DNA-binding element of the CcdA-CcdB complex, the stoichiometry of the two proteins determines whether or not the complex binds to the ccd operator-promoter region. Using electrophoretic mobility shift assays, we show that a (CcdA)2-(CcdB)2 complex binds DNA. The addition of extra CcdB to that protein-DNA complex completely abolishes DNA retardation. Based on these results, we propose a model in which the ratio between CcdA and CcdB regulates the repression state of the ccd operon. When the level of CcdA is superior or equal to that of CcdB, repression results. In contrast, derepression occurs when CcdB is in excess of CcdA. By ensuring an antidote-toxin ratio greater than one, this mechanism could prevent the harmful effect of CcdB in plasmid-containing bacteria.
Afif
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H
Laboratoire de Génétique des Procaryotes, Institut de Biologie et de Médecine Moléculaires, Université Libre de Bruxelles, 12 Rue des Professeurs Jeener et Brachet, 6041 Gosselies, Belgium.
Allali
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N
Couturier
M
M
Van Melderen
L
L
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D009875:Operator Regions, Genetic / Q000235:genetics / Q000502:physiology; D009876:Operon / Q000235:genetics; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics / Q000502:physiology; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D001427:Bacterial Toxins; C068355:CcdA protein, Bacteria; C068356:CcdB protein, Plasmid F; D004269:DNA, Bacterial