Abstract
Structural maintenance of chromosomes (SMC) complexes contribute to chromosome organization in all domains of life. In Escherichia coli, MukBEF, the functional SMC homolog, promotes spatiotemporal chromosome organization and faithful chromosome segregation. Here, we address the relative contributions of MukBEF and the replication terminus (ter) binding protein, MatP, to chromosome organization-segregation. We show that MukBEF, but not MatP, is required for the normal localization of the origin of replication to midcell and for the establishment of translational symmetry between newly replicated sister chromosomes. Overall, chromosome orientation is normally maintained through division from one generation to the next. Analysis of loci flanking the replication termination region (ter), which demark the ends of the linearly organized portion of the nucleoid, demonstrates that MatP is required for maintenance of chromosome orientation. We show that DNA-bound β2-processivity clamps, which mark the lagging strands at DNA replication forks, localize to the cell center, independent of replisome location but dependent on MukBEF action, and consistent with translational symmetry of sister chromosomes. Finally, we directly show that the older ("immortal") template DNA strand, propagated from previous generations, is preferentially inherited by the cell forming at the old pole, dependent on MukBEF and MatP. The work further implicates MukBEF and MatP as central players in chromosome organization, segregation, and nonrandom inheritance of genetic material and suggests a general framework for understanding how chromosome conformation and dynamics shape subcellular organization.
Highlights
Structural maintenance of chromosomes (SMC) complexes contribute to chromosome organization in all domains of life
We show that nascent lagging strands and their templates are directed toward cell centers, a process that is required for the observed translational L-RL-R segregational symmetry and which is perturbed in the absence of MukBEF
To understand how anucleate E. coli cells form in the absence of MukBEF, we followed the successive cell cycles of ΔmukB cells with oriC and ter regions fluorescently labeled by fluorescence repressor–operator system (FROS) markers
Summary
Structural maintenance of chromosomes (SMC) complexes contribute to chromosome organization in all domains of life. Organisms have evolved a vast range of mechanisms to ensure timely replication and segregation of genetic material Despite this diversity, highly conserved structural maintenance of chromosomes (SMC) complexes play a central role in the organization of chromosomes in all domains of life. SMC complexes, MukBEF, MksBEF, and Smc-ScpAB, were initially identified in a screen for Escherichia coli mutants that generated anucleate cells as a consequence of a failure to segregate newly replicated chromosomes to daughter cells [6, 7]. We address the relative roles of two major players in chromosome organization–segregation and propose a model of how E. coli maintains chromosome conformation and orientation inside cells and how this organization is propagated over generations
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