Abstract
Caulobacter crescentus
(CB15) initiates chromosome replication only in stalked cells and not in swarmers. To better understand this dimorphic control of chromosome replication, we isolated replication origins (
ori
s) from freshwater
Caulobacter
(FWC) and marine
Caulobacter
(MCS) species. Previous studies implicated integration host factor (IHF) and CcrM DNA methylation sites in replication control. However,
ori
IHF and CcrM sites identified in the model FWC CB15 were only conserved among closely related FWCs. DnaA boxes and CtrA binding sites are established CB15
ori
components. CtrA is a two-component regulator that blocks chromosome replication selectively in CB15 swarmers. DnaA boxes and CtrA sites were found in five FWC and three MCS
ori
s. Usually, a DnaA box and a CtrA site were paired, suggesting that CtrA binding regulates DnaA activity. We tested this hypothesis by site-directed mutagenesis of an MCS10
ori
which contains only one CtrA binding site overlapping a critical DnaA box. This overlapping site is unique in the whole MCS10 genome. Selective DnaA box mutations decreased replication, while selective CtrA binding site mutations increased replication of MCS10
ori
plasmids. Therefore, both FWC and MCS
ori
s use CtrA to repress replication. Despite this similarity, phylogenetic analysis unexpectedly shows that CtrA usage evolved separately among these
Caulobacter ori
s. We discuss consensus
ori
s and convergent
ori
evolution in differentiating bacteria.