The bacterial condensin MukB compacts DNA by sequestering supercoils and stabilizing topologically isolated loops

Kumar, Rupesh; Grosbart, Malgorzata; Nurse, Pearl; Bahng, Soon; Wyman, Claire; Marians, Kenneth J.

doi:10.1074/jbc.m117.803312

Cited by 28 publications

(47 citation statements)

References 52 publications

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“…Indeed, gel-shift assays and fluorescence anisotropy experiments showed DNA binding affinity to the hinge domain of mouse condensin (Griese et al, 2010), yeast condensin (Piazza et al, 2014), bovine cohesin (Chiu et al, 2004) and prokariotic condensin (Hirano, 2016). Previous AFM studies also showed DNA binding by the hinge domain for MukB (Kumar et al, 2017) and for S. pombe condensin (Yoshimura et al, 2002), suggesting a conserved hinge-binding mechanism. Our observation that the hinge angle of the O-shaped condensin changes dynamically ( Figure S6) indicates that the hinge domain is very flexible.…”

Section: Dna Binding Sites Of the Condensin Complexmentioning

confidence: 96%

AFM images of open and collapsed states of yeast condensin suggest a scrunching model for DNA loop extrusion

Ryu

Katan

et al. 2019

Preprint

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Structural Maintenance of Chromosome (SMC) protein complexes are the key organizers of the spatiotemporal structure of chromosomes. The condensin SMC complex, which compacts DNA during mitosis, was recently shown to be a molecular motor that extrudes large loops of DNA.The mechanism of this unique motor, which takes large steps along DNA at low ATP consumption, remains elusive however. Here, we use Atomic Force Microscopy (AFM) to visualize the structure of yeast condensin and condensin-DNA complexes. Condensin is foundto exhibit mainly open 'O' shapes and collapsed 'B' shapes, and it cycles dynamically between these two states over time. Condensin binds double-stranded DNA via a HEAT subunit and, surprisingly, also via the hinge domain. On extruded DNA loops, we observe a single condensin complex at the loop stem, where the neck size of the DNA loop correlates with the width of the condensin complex. Our results suggest that condensin extrudes DNA by a fast cyclic switching of its conformation between O and B shapes, consistent with a scrunching model.

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Section: Dna Binding Sites Of the Condensin Complexmentioning

confidence: 96%

AFM images of open and collapsed states of yeast condensin suggest a scrunching model for DNA loop extrusion

Ryu

Katan

et al. 2019

Preprint

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“…1). It is also worth noting that MukBEF has been indicated [49][50][51][52] to have some ability to multimerize (beyond the dimer of dimer functional unit [32]). This could play a role in the interaction between MukBEF-associated ori and the broader MukBEF gradient.…”

Section: Accurate Partitioning During Growthmentioning

confidence: 99%

“…In particular, MukBEF has a major role in chromosome organisation and facilitates long-range DNA interactions [40]. Like other SMC complexes, MukBEF may act by extruding loops of DNA [40], and/or be involved in stabilising them [52]. Furthermore, MatP, which binds to matS sites in the terminus region, interacts with MukBEF, displacing it from that region of the chromosome [12] and thereby restricting long-range DNA interactions between the terminus region and other regions of the chromosome [40].…”

Section: Accurate Partitioning During Growthmentioning

confidence: 99%

Self-organised segregation of bacterial chromosomal origins

Murray

Sourjik

2018

Preprint

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In spite of much effort, many aspects of chromosome organisation and segregation in bacteria remain unclear. Even for Escherichia coli, the most widely studied bacterial model organism, we still do not know the underlying mechanisms.Like many other bacteria, the chromosomal origin of replication in E. coli is dynamically positioned throughout the cell cycle. Initially maintained at mid-cell, where replication occurs, origins are subsequently partitioned to opposite quarter positions. The Structural Maintenance of Chromosomes (SMC) complex, MukBEF, which is required for correct chromosome compaction and organisation, has been implicated in this behaviour but the mode of action is unknown.Here, we build on a recent self-organising model for the positioning of E. coli MukBEF, to propose an explanation for the positioning and partitioning of origins. We propose that a specific association of MukBEF with the origin region, results in a non-trivial feedback between the self-organising MukBEF gradient and the origins, leading to accurate positioning and partitioning as an emergent property. We compare the model to quantitative experimental data of origin dynamics and their colocalisation with MukBEF clusters and find excellent agreement. Overall, the model suggests thatMukBEF and origins act together as a self-organising system for chromosome segregation and introduces protein selforganisation as an important consideration for future studies of chromosome dynamics.The faithful and timely segregation of genetic material is essential for all cellular life. In eukaryotes the responsibility for chromosome segregation lies with a well-understood macromolecular machine, the mitotic spindle. In contrast, the mechanisms underlying bacterial chromosome segregation are much less understood but are just as critical for cellular proliferation. Nevertheless, a lot has been learned in recent years (see [1] for a review). In particular, the starting point for (bidirectional) chromosomal replication, the origin, has been found to have a crucial role in chromosome organisation and segregation. Not only is it duplicated and segregated first but its genomic position defines other macrodomains [2] and its spatial position may determine the overall organisation of the chromosome [3].The positions of genomic loci, including the origin, change as replication and segregation progress. Initiated at the chromosomal origin, replication proceeds bi-directionally down each arm of the chromosome. In slowly to moderately growing (1-2 hr generation time) Escherichia coli cells, the home positon of the origin, oriC (henceforth and in the model, ori), in new-born cells is at mid-cell [4][5][6]. After 10-15 min of 'cohesion', in at least part arising from interlinking of the two daughter chromosomes (precatenation) [7][8][9][10][11][12], duplicated origins separate and migrate, initially very rapidly, to opposite quarter positions, which become the new home positions for the remainder of the cell cycle [13,14]. Other genomic loci migrate sequentially to ...

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“…Thus far, SMC proteins have not been implied in phase separation. While ATP-independent clustering of DNA and SMC proteins has been reported (11)(12)(13)(14), such observations were attributed to imperfect protein purification or non-physiological buffer conditions. For example, Davidson et al (7) reported in vitro DNA loop extrusion by the human cohesin complex when the complex concentration was limited to very low values (<0.8 nM, i.e., much lower that physiological concentrations of ~333 nM (15,16), and mentioned that the cohesin complexes were prone to aggregation at higher concentrations.…”

mentioning

confidence: 99%

Phase separation induced by cohesin SMC protein complexes

Ryu

Bouchoux

Liu

et al. 2020

Preprint

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Cohesin is a key protein complex that organizes the spatial structure of chromosomes during interphase. Here, we show that yeast cohesin shows pronounced clustering on DNA in an ATPindependent manner, exhibiting all the hallmarks of phase separation. In vitro visualization of cohesin on DNA shows DNA-cohesin clusters that exhibit liquid-like behavior. This includes mutual fusion and reversible dissociation upon depleting the cohesin concentration, increasing the ionic strength, or adding 1,6-hexanediol, conditions that disrupt weak interactions. We discuss how bridging-induced phase separation can explain the DNA-cohesin clustering through DNA-cohesin-DNA bridges. We confirm that, in vivo, a fraction of cohesin associates with chromatin in yeast cells in a manner consistent with phase separation. Our findings establish that SMC proteins can exhibit phase separation, which has potential to clarify previously unexplained aspects of in vivo SMC behavior and constitute an additional principle by which SMC complexes impact genome organization. One sentence summary:Yeast cohesin complex is observed to phase separate with DNA into liquid droplets, which it accomplishes by ATP-independent DNA bridging.Members of the structural maintenance of chromosome (SMC) protein family such as condensin, cohesin, and the Smc5/6 complex are key proteins for the spatial and temporal organization of chromosomes (1)(2)(3)(4). Recent in vitro experiments visualized real-time DNA loop extrusion mediated by condensin and cohesin (5-7). While loop extrusion by SMC proteins constitutes a fundamental building block in the organization of chromosomes, other factors may also contribute. In the last decade, it has become abundantly clear that phase separation plays a role in many processes in biological cells (8), including chromosome organization (9, 10). Thus far, SMC proteins have not been implied in phase separation. While ATP-independent clustering of DNA and SMC proteins has been reported (11)(12)(13)(14), such observations were attributed to imperfect protein purification or non-physiological buffer conditions. For example, Davidson et al. (7) reported in vitro DNA loop extrusion by the human cohesin complex when the complex concentration was limited to very low values (<0.8 nM, i.e., much lower that physiological concentrations of ~333 nM (15,16), and mentioned that the cohesin complexes were prone to aggregation at higher concentrations. Such findings raise the question whether such aggregate formation may be intrinsic and may have a physiological meaning.Here we report that interactions between the yeast cohesin SMC complex and DNA lead to pronounced phase separation. This clustering behavior is ATP independent but depends on DNA length. We find that single cohesin complexes are able to bridge distant points along DNA that act as nucleation points for recruiting further cohesin complexes -a behavior indicating bridging-induced phase separation (BIPS) (17,18), also known as polymer-polymer phase separation (PPPS) (19), a type of phase co...

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The bacterial condensin MukB compacts DNA by sequestering supercoils and stabilizing topologically isolated loops

Cited by 28 publications

References 52 publications

AFM images of open and collapsed states of yeast condensin suggest a scrunching model for DNA loop extrusion

AFM images of open and collapsed states of yeast condensin suggest a scrunching model for DNA loop extrusion

Self-organised segregation of bacterial chromosomal origins

Phase separation induced by cohesin SMC protein complexes

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