Abstract
ABSTRACTThe 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.
Highlights
Successful propagation of cells relies on the fidelity of chromosome replication and segregation and the processes that compact and organize the chromosome
Structural maintenance of chromosome (SMC) complexes, which are present in most organisms, have been implicated in both bacterial chromosome organization and segregation, their precise functional role and biochemical action remain unclear
Topoisomerase IV associates with MukBEF in live E. coli
Summary
Successful propagation of cells relies on the fidelity of chromosome replication and segregation and the processes that compact and organize the chromosome. Biochemical experiments in vitro have shown that the MukB hinge interacts with the C-terminal domain of ParC, the catalytic subunit of TopoIV [18,19,20], leading to the proposal that these proteins collaborate in chromosome disentanglement [20] This interaction stimulates TopoIV-mediated relaxation of negatively supercoiled DNA, but not positively supercoiled DNA in vitro. Any MukBEF-stimulated negative supercoil relaxation would compound the in vivo unlinking problem because it would act to increase the overall linkage between duplex strands in the chromosome Other studies in both prokaryotes and eukaryotes have implicated functional interactions between SMC complexes and type II topoisomerases and have suggested that these are important for decatenation, it is not always clear whether these interactions influence the activity of the topoisomerase in regulating supercoiling, which is important for chromosome organization [5, 26,27,28]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
