scholarly journals The cohesin ring uses its hinge to organize DNA using non-topological as well as topological mechanisms

2017 ◽  
Author(s):  
Madhusudhan Srinivasan ◽  
Johanna C. Scheinost ◽  
Naomi J. Petela ◽  
Thomas G. Gligoris ◽  
Maria Wissler ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Sister Chromatid ◽  
Atp Hydrolysis ◽  
Sister Chromatid Cohesion ◽  
Perfect Correlation ◽  
Chromatid Cohesion ◽  
Lysine Residues ◽  
Positively Charged ◽  
Hinge Domain

SummaryAs predicted by the notion that sister chromatid cohesion is mediated by entrapment of sister DNAs inside cohesin rings, there is a perfect correlation between co-entrapment of circular minichromosomes and sister chromatid cohesion in a large variety of mutants. In most cells where cohesin loads onto chromosomes but fails to form cohesion, loading is accompanied by entrapment of individual DNAs. However, cohesin with a hinge domain whose positively charged lumen has been neutralized not only loads onto and translocates along chromatin but also organizes it into chromatid-like threads, despite largely failing to entrap DNAs inside its ring. Thus, cohesin engages chromatin in a non-topological as well as a topological manner. Our finding that hinge mutations, but not fusions between Smc and kleisin subunits, abolish entrapment suggests that DNAs may enter cohesin rings through hinge opening. Lastly, mutation of three highly conserved lysine residues inside the Smc1 moiety of Smc1/3 hinges abolishes all loading without affecting cohesin’s initial recruitment to CEN loading sites or its ability to hydrolyze ATP. We suggest that loading and translocation are mediated by conformational changes in cohesin’s hinge driven by cycles of ATP hydrolysis.

scholarly journals A positively charged channel within the Smc1/Smc3 hinge required for sister chromatid cohesion

2010 ◽  
Vol 30 (2) ◽  
pp. 364-378 ◽  
Author(s):  
Alexander Kurze ◽  
Katharine A Michie ◽  
Sarah E Dixon ◽  
Ajay Mishra ◽  
Takehiko Itoh ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Sister Chromatid Cohesion ◽  
Chromatid Cohesion ◽  
Positively Charged

scholarly journals ATP dependent DNA transport within cohesin: Scc2 clamps DNA on top of engaged heads while Scc3 promotes entrapment within the SMC-kleisin ring

2020 ◽  
Author(s):  
James E Collier ◽  
Byung-Gil Lee ◽  
Maurici B Roig ◽  
Stanislav Yatskevich ◽  
Naomi J Petela ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Atp Hydrolysis ◽  
Sister Chromatid Cohesion ◽  
S Phase ◽  
Coiled Coils ◽  
Rigorous Proof ◽  
Dna Loops ◽  
Dna Transport ◽  
Chromatid Cohesion

SUMMARYIn addition to extruding DNA loops, cohesin entraps within its SMC-kleisin ring (S-K) individual DNAs during G1 and sister DNAs during S-phase. All three activities require related hook-shaped proteins called Scc2 and Scc3. Using thiol-specific crosslinking we provide rigorous proof of entrapment activity in vitro. Scc2 alone promotes entrapment of DNAs in the E-S and E-K compartments, between ATP-bound engaged heads and the SMC hinge and associated kleisin, respectively. This does not require ATP hydrolysis nor is it accompanied by entrapment within S-K rings, which is a slower process requiring Scc3. Cryo-EM reveals that DNAs transported into E-S/E-K compartments are “clamped” in a sub-compartment created by Scc2’s association with engaged heads whose coiled coils are folded around their elbow. We suggest that clamping may be a recurrent feature of cohesin complexes active in loop extrusion and that this conformation precedes the S-K entrapment required for sister chromatid cohesion.

scholarly journals A Structure-Based Mechanism for DNA Entry into the Cohesin Ring

2020 ◽  
Author(s):  
Torahiko L. Higashi ◽  
Patrik Eickhoff ◽  
Joana S. Simoes ◽  
Julia Locke ◽  
Andrea Nans ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Molecular Basis ◽  
Atp Hydrolysis ◽  
Initial Step ◽  
Sister Chromatid Cohesion ◽  
Chromosome Organization ◽  
Atp Binding ◽  
Structural Framework ◽  
Gate Opening ◽  
Chromatid Cohesion

SUMMARYDespite key roles in sister chromatid cohesion and chromosome organization, the mechanism by which cohesin rings are loaded onto DNA is still unknown. Here, we combine biophysical approaches and cryo-EM to visualize a cohesin loading intermediate in which DNA is locked between two gates that lead into the cohesin ring. Building on this structural framework, we design biochemical experiments to establish the order of events during cohesin loading. In an initial step, DNA traverses an N-terminal kleisin gate that is first opened upon ATP binding and then closed as the cohesin loader locks the DNA against a shut ATPase gate. ATP hydrolysis leads to ATPase gate opening to complete DNA entry. Whether DNA loading is successful, or rather results in loop extrusion, might be dictated by a conserved kleisin N-terminal tail that guides the DNA through the kleisin gate. Our results establish the molecular basis for cohesin loading onto DNA.

scholarly journals Multiple interactions between Scc1 and Scc2 activate cohesin’s DNA dependent ATPase and replace Pds5 during loading

2017 ◽  
Author(s):  
Naomi J Petela ◽  
Thomas G Gligoris ◽  
Jean Metson ◽  
Byung-Gil Lee ◽  
Menelaos Voulgaris ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Sister Chromatid ◽  
Atp Hydrolysis ◽  
Sister Chromatid Cohesion ◽  
Dependent Atpase ◽  
Heat Repeat ◽  
Repeat Proteins ◽  
Chromatid Cohesion ◽  
Multiple Interactions

SummaryIn addition to sharing with condensin an ability to organize DNA into chromatids, cohesin regulates enhancer-promoter interactions and confers sister chromatid cohesion. Association with chromosomes is regulated by hook-shaped HEAT repeat proteins that Associate With its Kleisin (Scc1) subunit (HAWKs), namely Scc3, Pds5, and Scc2. Unlike Pds5, Scc2 is not a stable cohesin constituent but, as shown here, transiently displaces Pds5 during loading. Scc1 mutations that compromise its interaction with Scc2 adversely affect cohesin’s ATPase activity, loading, and translocation while Scc2 mutations that alter how the ATPase responds to DNA abolish loading despite cohesin’s initial association with loading sites. Lastly, Scc2 mutations that permit loading in the absence of Scc4 increase Scc2’s association with chromosomal cohesin and reduce that of Pds5. We suggest that cohesin switches between two states, one with Pds5 bound to Scc1 that is not able to hydrolyse ATP efficiently but is capable of release from chromosomes and another in which Scc2, transiently replacing Pds5, stimulates the ATP hydrolysis necessary for loading and translocation away from loading sites.

scholarly journals Transport of DNA within cohesin involves clamping on top of engaged heads by Scc2 and entrapment within the ring by Scc3

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
James E Collier ◽  
Byung-Gil Lee ◽  
Maurici Brunet Roig ◽  
Stanislav Yatskevich ◽  
Naomi J Petela ◽  
...  
Keyword(s):  
Sister Chromatid ◽  
Atp Hydrolysis ◽  
Sister Chromatid Cohesion ◽  
S Phase ◽  
Coiled Coils ◽  
Rigorous Proof ◽  
Dna Loops ◽  
Chromatid Cohesion

In addition to extruding DNA loops, cohesin entraps within its SMC-kleisin ring (S-K) individual DNAs during G1 and sister DNAs during S-phase. All three activities require related hook-shaped proteins called Scc2 and Scc3. Using thiol-specific crosslinking we provide rigorous proof of entrapment activity in vitro. Scc2 alone promotes entrapment of DNAs in the E-S and E-K compartments, between ATP-bound engaged heads and the SMC hinge and associated kleisin, respectively. This does not require ATP hydrolysis nor is it accompanied by entrapment within S-K rings, which is a slower process requiring Scc3. Cryo-EM reveals that DNAs transported into E-S/E-K compartments are ‘clamped’ in a sub-compartment created by Scc2’s association with engaged heads whose coiled coils are folded around their elbow. We suggest that clamping may be a recurrent feature of cohesin complexes active in loop extrusion and that this conformation precedes the S-K entrapment required for sister chromatid cohesion.

scholarly journals A role for the Smc3 hinge domain in the maintenance of sister chromatid cohesion

2017 ◽  
Author(s):  
Brett Robison ◽  
Vincent Guacci ◽  
Douglas Koshland
Keyword(s):  
Sister Chromatid ◽  
Sister Chromatid Cohesion ◽  
Insertion Mutagenesis ◽  
Regulation Of Transcription ◽  
Dna Duplexes ◽  
Damage Repair ◽  
Chromatid Cohesion ◽  
Mutant Phenotypes ◽  
Random Insertion ◽  
Hinge Domain

AbstractCohesin is a conserved protein complex required for sister chromatid cohesion, chromosome condensation, DNA damage repair, and regulation of transcription. Although cohesin functions to tether DNA duplexes, the contribution of its individual domains to this activity remains poorly understood. We interrogated the Smc3p subunit of cohesin by random insertion mutagenesis. Analysis of a mutant in the Smc3p hinge revealed an unexpected role for this domain in cohesion maintenance and condensation. Further investigation revealed that the Smc3p hinge functions at a step following cohesin’s stable binding to chromosomes and independently of Smc3p’s regulation by the Eco1p acetyltransferase. Hinge mutant phenotypes resemble loss of Pds5p, which binds opposite the hinge near Smc3p’s head domain. We propose that a specific conformation of the Smc3p hinge and Pds5p cooperate to promote cohesion maintenance and condensation.

scholarly journals The acetyltransferase Eco1 elicits cohesin dimerization during S phase

2020 ◽  
Vol 295 (22) ◽  
pp. 7554-7565 ◽  
Author(s):  
Di Shi ◽  
Shuaijun Zhao ◽  
Mei-Qing Zuo ◽  
Jingjing Zhang ◽  
Wenya Hou ◽  
...  
Keyword(s):  
Dna Replication ◽  
Sister Chromatid ◽  
Sister Chromatid Cohesion ◽  
S Phase ◽  
Cross Linking ◽  
Temperature Sensitive ◽  
Exact Nature ◽  
Chromatid Cohesion ◽  
Lysine Residues

Cohesin is a DNA-associated protein complex that forms a tripartite ring controlling sister chromatid cohesion, chromosome segregation and organization, DNA replication, and gene expression. Sister chromatid cohesion is established by the protein acetyltransferase Eco1, which acetylates two conserved lysine residues on the cohesin subunit Smc3 and thereby ensures correct chromatid separation in yeast (Saccharomyces cerevisiae) and other eukaryotes. However, the consequence of Eco1-catalyzed cohesin acetylation is unknown, and the exact nature of the cohesive state of chromatids remains controversial. Here, we show that self-interactions of the cohesin subunits Scc1/Rad21 and Scc3 occur in a DNA replication–coupled manner in both yeast and human cells. Using cross-linking MS-based and in vivo disulfide cross-linking analyses of purified cohesin, we show that a subpopulation of cohesin may exist as dimers. Importantly, upon temperature-sensitive and auxin-induced degron-mediated Eco1 depletion, the cohesin-cohesin interactions became significantly compromised, whereas deleting either the deacetylase Hos1 or the Eco1 antagonist Wpl1/Rad61 increased cohesin dimer levels by ∼20%. These results indicate that cohesin dimerizes in the S phase and monomerizes in mitosis, processes that are controlled by Eco1, Wpl1, and Hos1 in the sister chromatid cohesion-dissolution cycle. These findings suggest that cohesin dimerization is controlled by the cohesion cycle and support the notion that a double-ring cohesin model operates in sister chromatid cohesion.

scholarly journals A role for the Smc3 hinge domain in the maintenance of sister chromatid cohesion

2018 ◽  
Vol 29 (3) ◽  
pp. 339-355 ◽  
Author(s):  
Brett Robison ◽  
Vincent Guacci ◽  
Douglas Koshland
Keyword(s):  
Sister Chromatid ◽  
Sister Chromatid Cohesion ◽  
Chromatid Cohesion ◽  
Cohesin Subunit ◽  
Hinge Domain

A screen of cohesin subunit Smc3 reveals that its hinge is a nexus controlling the maintenance of sister chromatid cohesion and condensation.

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