The initiator protein DnaA contributes to keeping new origins inactivated by promoting the presence of hemimethylated DNA.
The Escherichia coli replication origin oriC and other regions with high numbers of GATC sites remain hemimethylated after replication much longer than regions with average numbers of GATC sites. The prolonged period of hemimethylation has been attributed to the presence of bound SeqA protein. Here, it was found that a GATC cluster inserted at the datA site, which binds large amounts of DnaA in vivo, did not become remethylated at all, unless the availability of the DnaA protein was severely reduced. Sequestration of oriC was also found to be affected by the availability of DnaA. The period of origin hemimethylation was reduced by approximately 30% upon a reduction in the availability of DnaA. The result shows that not only SeqA binding but also DnaA binding to newly replicated origins contributes to keeping them hemimethylated. It was also found that the number of SeqA foci increased in cells with a combination of DnaA-mediated protection and sequestration at the GATC::datA cluster.
Department of Cell Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Rikshospitalet, University of Oslo, 0310 Oslo, Norway.
Trond
T
Bach
Morigen
Kirsten
K
Skarstad
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D002453:Cell Cycle; D002876:Chromosomes, Bacterial / Q000378:metabolism; D004265:DNA Helicases / Q000378:metabolism*; D019175:DNA Methylation; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D010957:Plasmids; D018741:Replication Origin; D015534:Trans-Activators / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D029968:Escherichia coli Proteins; C089746:SeqA protein, E coli; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases