Transcriptional inactivation of a regulatory site for replication of Vibrio cholerae chromosome II

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Abstract
The bacterium Vibrio cholerae has two chromosomes. The origin of replication of chromosome I is similar to that of Escherichia coli . The origin-containing region of chromosome II ( oriCII ) resembles replicons of plasmids such as P1, except for the presence of an additional gene, rctA [Egan, E. S. & Waldor, M. K. (2003) Cell 114, 521–530]. The oriCII region that includes the initiator gene, rctB , can function as a plasmid in E. coli . Here we show that RctB suffices for the oriCII -based plasmid replication, and rctA in cis or trans reduces the plasmid copy number, thereby serving as a negative regulator. The inhibitory activity could be overcome by increasing the concentration of RctB, suggesting that rctA titrates the initiator. Purified RctB bound to a DNA fragment carrying rctA , confirming that the two can interact. Although rctA apparently works as a titrating site, it is nonetheless transcribed. We find that the transcription attenuates the inhibitory activity of the gene, presumably by interfering with RctB binding. RctB, in turn, repressed the rctA promoter and, thereby, could control its own titration by modulating the transcription of rctA . This control circuit appears to be a putative novel mechanism for homeostasis of initiator availability.