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Edited by Nancy E. Kleckner, Harvard University, Cambridge, MA, and approved May 17, 2011 (received for review January 18, 2011)
Abstract
Characterized bacteria, unlike eukaryotes and some archaea, initiate replication bidirectionally from a single replication origin contained within a circular or linear chromosome. We constructed Escherichia coli cells with two WT origins separated by 1 Mb in their 4.64-Mb chromosome. Productive bidirectional replication initiated synchronously at both spatially separate origins. Newly replicated DNA from both origins was segregated sequentially as replication progressed, with two temporally and spatially separate replication termination events. Replication initiation occurred at a cell volume identical to that of cells with a single WT origin, showing that initiation control is independent of cellular and chromosomal oriC concentration. Cells containing just the ectopic origin initiated bidirectional replication at the expected cell mass and at the normal cellular location of that region. In all strains, spatial separation of sister loci adjacent to active origins occurred shortly after their replication, independently of whether replication initiated at the normal origin, the ectopic origin, or both origins.
Footnotes
↵1X.W. and C.L. contributed equally to this work.
↵2Present address: Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115.
- ↵3To whom correspondence should be addressed. E-mail: david.sherratt{at}bioch.ox.ac.uk.
Author contributions: X.W., C.L., R.R.-L., and D.J.S. designed research; X.W., C.L., and R.R.-L. performed research; G.B. contributed new reagents/analytic tools; X.W., C.L., and R.R.-L. analyzed data; and X.W., C.L., and D.J.S. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
See Author Summary on page 10383.
This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1100874108/-/DCSupplemental.
Freely available online through the PNAS open access option.
Replication and segregation of an Escherichia coli chromosome with two replication origins
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