title: Escherichia coli SeqA structures relocalize abruptly upon termination of origin sequestration during multifork DNA replication.
abstract: The Escherichia coli SeqA protein forms complexes with new, hemimethylated DNA behind replication forks and is important for successful replication during rapid growth. Here, E. coli cells with two simultaneously replicating chromosomes (multifork DNA replication) and YFP tagged SeqA protein was studied. Fluorescence microscopy showed that in the beginning of the cell cycle cells contained a single focus at midcell. The focus was found to remain relatively immobile at midcell for a period of time equivalent to the duration of origin sequestration. Then, two abrupt relocalization events occurred within 2-6 minutes and resulted in SeqA foci localized at each of the cell's quarter positions. Imaging of cells containing an additional fluorescent tag in the origin region showed that SeqA colocalizes with the origin region during sequestration. This indicates that the newly replicated DNA of first one chromosome, and then the other, is moved from midcell to the quarter positions. At the same time, origins are released from sequestration. Our results illustrate that newly replicated sister DNA is segregated pairwise to the new locations. This mode of segregation is in principle different from that of slowly growing bacteria where the newly replicated sister DNA is partitioned to separate cell halves and the decatenation of sisters a prerequisite for, and possibly a mechanistic part of, segregation.
authors.affiliation: Department of Cell Biology, Institute for Cancer Research, Oslo University Hospital, the Norwegian Radium Hospital, Oslo, Norway; School of Pharmacy, Faculty of Mathematics and Natural Sciences, University of Oslo, Oslo, Norway.
authors.forename: Solveig
authors.initials: S
authors.lastname: Fossum-Raunehaug
authors.affiliation: Department of Cell Biology, Institute for Cancer Research, Oslo University Hospital, the Norwegian Radium Hospital, Oslo, Norway.
authors.forename: Emily
authors.initials: E
authors.lastname: Helgesen
authors.affiliation: Department of Cell Biology, Institute for Cancer Research, Oslo University Hospital, the Norwegian Radium Hospital, Oslo, Norway.
authors.forename: Caroline
authors.initials: C
authors.lastname: Stokke
authors.affiliation: Department of Cell Biology, Institute for Cancer Research, Oslo University Hospital, the Norwegian Radium Hospital, Oslo, Norway; School of Pharmacy, Faculty of Mathematics and Natural Sciences, University of Oslo, Oslo, Norway.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Skarstad
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D019175:DNA Methylation; D004261:DNA Replication; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D011485:Protein Binding; D018741:Replication Origin
chemical_list: D001425:Bacterial Outer Membrane Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C089746:SeqA protein, E coli