The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>

ABSTRACT The Escherichia coli structural maintenance of chromosome (SMC) complex, MukBEF, and topoisomerase IV (TopoIV) interact in vitro through a direct contact between the MukB dimerization hinge and the C-terminal domain of ParC, the catalytic subunit of TopoIV. The interaction stimulates cataly...

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Autores principales: Emilien Nicolas, Amy L. Upton, Stephan Uphoff, Olivia Henry, Anjana Badrinarayanan, David Sherratt
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Publicado: American Society for Microbiology 2014
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spelling oai:doaj.org-article:108c324e4f974d9c96c399b1d02fe71c2021-11-15T15:45:11ZThe SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>10.1128/mBio.01001-132150-7511https://doaj.org/article/108c324e4f974d9c96c399b1d02fe71c2014-02-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01001-13https://doaj.org/toc/2150-7511ABSTRACT The Escherichia coli structural maintenance of chromosome (SMC) complex, MukBEF, and topoisomerase IV (TopoIV) interact in vitro through a direct contact between the MukB dimerization hinge and the C-terminal domain of ParC, the catalytic subunit of TopoIV. The interaction stimulates catalysis by TopoIV in vitro. Using live-cell quantitative imaging, we show that MukBEF directs TopoIV to ori, with fluorescent fusions of ParC and ParE both forming cellular foci that colocalize with those formed by MukBEF throughout the cell cycle and in cells unable to initiate DNA replication. Removal of MukBEF leads to loss of fluorescent ParC/ParE foci. In the absence of functional TopoIV, MukBEF forms multiple foci that are distributed uniformly throughout the nucleoid, whereas multiple catenated oris cluster at midcell. Once functional TopoIV is restored, the decatenated oris segregate to positions that are largely coincident with the MukBEF foci, thereby providing support for a mechanism by which MukBEF acts in chromosome segregation by positioning newly replicated and decatenated oris. Additional evidence for such a mechanism comes from the observation that in TopoIV-positive (TopoIV+) cells, newly replicated oris segregate rapidly to the positions of MukBEF foci. Taken together, the data implicate MukBEF as a key component of the DNA segregation process by acting in concert with TopoIV to promote decatenation and positioning of newly replicated oris. IMPORTANCE Mechanistic understanding of how newly replicated bacterial chromosomes are segregated prior to cell division is incomplete. In this work, we provide in vivo experimental support for the view that topoisomerase IV (TopoIV), which decatenates newly replicated sister duplexes as a prelude to successful segregation, is directed to the replication origin region of the Escherichia coli chromosome by the SMC (structural maintenance of chromosome) complex, MukBEF. We provide in vivo data that support the demonstration in vitro that the MukB interaction with TopoIV stimulates catalysis by TopoIV. Finally, we show that MukBEF directs the normal positioning of sister origins after their replication and during their segregation. Overall, the data support models in which the coordinate and sequential action of TopoIV and MukBEF plays an important role during bacterial chromosome segregation.Emilien NicolasAmy L. UptonStephan UphoffOlivia HenryAnjana BadrinarayananDavid SherrattAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 5, Iss 1 (2014)
institution DOAJ
collection DOAJ
language EN
topic Microbiology
QR1-502
spellingShingle Microbiology
QR1-502
Emilien Nicolas
Amy L. Upton
Stephan Uphoff
Olivia Henry
Anjana Badrinarayanan
David Sherratt
The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
description ABSTRACT The Escherichia coli structural maintenance of chromosome (SMC) complex, MukBEF, and topoisomerase IV (TopoIV) interact in vitro through a direct contact between the MukB dimerization hinge and the C-terminal domain of ParC, the catalytic subunit of TopoIV. The interaction stimulates catalysis by TopoIV in vitro. Using live-cell quantitative imaging, we show that MukBEF directs TopoIV to ori, with fluorescent fusions of ParC and ParE both forming cellular foci that colocalize with those formed by MukBEF throughout the cell cycle and in cells unable to initiate DNA replication. Removal of MukBEF leads to loss of fluorescent ParC/ParE foci. In the absence of functional TopoIV, MukBEF forms multiple foci that are distributed uniformly throughout the nucleoid, whereas multiple catenated oris cluster at midcell. Once functional TopoIV is restored, the decatenated oris segregate to positions that are largely coincident with the MukBEF foci, thereby providing support for a mechanism by which MukBEF acts in chromosome segregation by positioning newly replicated and decatenated oris. Additional evidence for such a mechanism comes from the observation that in TopoIV-positive (TopoIV+) cells, newly replicated oris segregate rapidly to the positions of MukBEF foci. Taken together, the data implicate MukBEF as a key component of the DNA segregation process by acting in concert with TopoIV to promote decatenation and positioning of newly replicated oris. IMPORTANCE Mechanistic understanding of how newly replicated bacterial chromosomes are segregated prior to cell division is incomplete. In this work, we provide in vivo experimental support for the view that topoisomerase IV (TopoIV), which decatenates newly replicated sister duplexes as a prelude to successful segregation, is directed to the replication origin region of the Escherichia coli chromosome by the SMC (structural maintenance of chromosome) complex, MukBEF. We provide in vivo data that support the demonstration in vitro that the MukB interaction with TopoIV stimulates catalysis by TopoIV. Finally, we show that MukBEF directs the normal positioning of sister origins after their replication and during their segregation. Overall, the data support models in which the coordinate and sequential action of TopoIV and MukBEF plays an important role during bacterial chromosome segregation.
format article
author Emilien Nicolas
Amy L. Upton
Stephan Uphoff
Olivia Henry
Anjana Badrinarayanan
David Sherratt
author_facet Emilien Nicolas
Amy L. Upton
Stephan Uphoff
Olivia Henry
Anjana Badrinarayanan
David Sherratt
author_sort Emilien Nicolas
title The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
title_short The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
title_full The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
title_fullStr The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
title_full_unstemmed The SMC Complex MukBEF Recruits Topoisomerase IV to the Origin of Replication Region in Live <named-content content-type="genus-species">Escherichia coli</named-content>
title_sort smc complex mukbef recruits topoisomerase iv to the origin of replication region in live <named-content content-type="genus-species">escherichia coli</named-content>
publisher American Society for Microbiology
publishDate 2014
url https://doaj.org/article/108c324e4f974d9c96c399b1d02fe71c
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