Cryo-EM structure of MukBEF reveals DNA loop entrapment at chromosomal unloading sites

uumlrmann, F. B; Funke, Louise F. H.; Chin, Jason W.; oumlwe, J. L

doi:10.1101/2021.06.29.450292

Cited by 15 publications

(42 citation statements)

References 91 publications

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“…Early molecular models for loop extrusion suggested that the SMC ring topologically embraces the DNA upon loading and during DNA loop extrusion. 31,40 More recently, also a pseudotopological 11,41,13 and nontopological loading 13,42 of the DNA were considered. The data from the current study provide unambiguous evidence for a nontopological mode for DNA loop extrusion, where DNA must bind externally to the SMC complex.…”

mentioning

confidence: 99%

SMC complexes can traverse physical roadblocks bigger than their ring size

Pradhan

Barth

Kim

et al. 2021

Preprint

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The ring-shaped structural-maintenance-of-chromosomes (SMC) complexes condensin and cohesin extrude loops of DNA as a key motif in chromosome organization. It remains, however, unclear how these SMC motor proteins can extrude DNA loops in chromatin that is bound with proteins. Here, using in vitro single-molecule visualization, we show that nucleosomes, RNA polymerase, and dCas9 pose virtually no barrier to DNA loop extrusion by yeast condensin. Strikingly, we find that even DNA-bound nanoparticles as large as 200 nm, much bigger than the SMC ring size, can be translocated into DNA loops during condensin-driven extrusion. Similarly, human cohesin can pass 200 nm particles during loop extrusion, which even occurs for a single-chain version of cohesin in which the ring-forming subunits are covalently linked and cannot open up to entrap DNA. These findings disqualify all common loop-extrusion models where DNA passes through the SMC rings (pseudo)topologically, and instead point to a nontopological mechanism for DNA loop extrusion.

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mentioning

confidence: 99%

SMC complexes can traverse physical roadblocks bigger than their ring size

Pradhan

Barth

Kim

et al. 2021

Preprint

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“…ParC inhibits the MukBF ATPase (21), implying, given the folded conformation of MukB (23,24) and our crosslinking results, that ParC might interpose itself between the MukB hinge and neck regions and that this might disrupt the ATPase cycle, further suggesting that isolated MukB hinge might stimulate the MukBF ATPase in trans . This proved to be the case.…”

Section: Resultsmentioning

confidence: 81%

“…Mapping of MukB head and neck mutations to the MukBEF structures of Burmann et al (23). A, overall view of the head, neck and larynx regions of the MukBF dimer in the core MukBEF cryo-EM structure.…”

Section: Discussionmentioning

confidence: 99%

“…The mutations in MukB DNA (R187E R189E), were made based on comparison to residues in the head region of the B. subtilis SMC protein that were required for DNA binding (27). The cryo-EM structure of Burmann et al (23) shows R187 and R189 to be just adjacent to the DNA binding site occupied by the short duplex DNA. Given their position, these residues could interact with DNA in the absence of MukF.…”

Section: A Push By the Hinge Domain At The Head And Neck Of Mukb Is R...mentioning

confidence: 99%

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Inter- and intra-Subunit interactions driving the MukBEF ATPase

Bahng

Kumar

Marians

2022

Preprint

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MukBEF, an SMC-like protein complex, is the bacterial condensin. It is likely that it compacts DNA via an ATP hydrolysis-dependent, DNA loop-extrusion reaction as has been demonstrated for the yeast SMC proteins condensin and cohesin. MukB also interacts with the ParC subunit of the cellular chromosomal decatenase topoisomerase IV, an interaction that is required for proper chromosome condensation and segregation in E. coli, although it suppresses the MukB ATPase activity. We have investigated the MukBEF ATPase activity, identifying inter- and intra-subunit interactions by protein-protein crosslinking and site-specific mutagenesis. We show that interactions between the hinge of MukB and its neck region are essential for the ATPase activity, that the ParC subunit of Topo IV inhibits the MukB ATPase by preventing this interaction, that MukE interaction with DNA is likely essential for viability, and that interactions between MukF and the MukB neck region are necessary for ATPase activity and viability.

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“…The latter establishes the overall ring structure of the SMCC, and formation of the ATPase-ATP-ATPase "bridge" (shown in blue in Fig. 1B) can divide the ring into two compartments (25,28,61,62).…”

Section: Model Geometrymentioning

confidence: 93%

DNA tension-modulated translocation and loop extrusion by SMC complexes revealed by molecular dynamics simulations

Nomidis

Carlon

Gruber

et al. 2021

Preprint

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Structural Maintenance of Chromosomes (SMC) protein complexes play essential roles in genome folding and organization across all domains of life. In order to determine how the activities of these large (about 50 nm) complexes are controlled by ATP binding and hydrolysis, we have developed a molecular dynamics (MD) model that realistically accounts for thermal conformational motions of SMC and DNA. The model SMCs make use of DNA flexibility and looping, together with an ATP-induced "power stroke", to capture and transport DNA segments, so as to robustly translocate along DNA. This process is sensitive to DNA tension: at low tension (about 0.1 pN), the model performs steps of roughly 60 nm size, while, at higher tension, a distinct inchworm-like translocation mode appears, with steps that depend on SMC arm flexibility. By permanently tethering DNA to an experimentally-observed additional binding site ("safety belt"), the same model performs loop extrusion. We find that the dependence of loop extrusion on DNA tension is remarkably different when DNA tension is fixed vs when DNA end points are fixed: Loop extrusion reversal occurs above 0.5 pN for fixed tension, while loop extrusion stalling without reversal occurs at about 2 pN for fixed end points. Our model quantitatively matches recent experimental results on condensin and cohesin, and makes a number of clear predictions. Finally we investigate how specific structural changes affect the SMC function, which is testable in experiments on varied or mutant SMCs.

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Cryo-EM structure of MukBEF reveals DNA loop entrapment at chromosomal unloading sites

Cited by 15 publications

References 91 publications

SMC complexes can traverse physical roadblocks bigger than their ring size

SMC complexes can traverse physical roadblocks bigger than their ring size

Inter- and intra-Subunit interactions driving the MukBEF ATPase

DNA tension-modulated translocation and loop extrusion by SMC complexes revealed by molecular dynamics simulations

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