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Journal articles on the topic 'Chromatin loop extrusion'
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1
Racko, Dusan, Fabrizio Benedetti, Dimos Goundaroulis, and Andrzej Stasiak. "Chromatin Loop Extrusion and Chromatin Unknotting." Polymers 10, no. 10 (October 11, 2018): 1126. http://dx.doi.org/10.3390/polym10101126.
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It has been a puzzle how decondensed interphase chromosomes remain essentially unknotted. The natural expectation is that in the presence of type II DNA topoisomerases that permit passages of double-stranded DNA regions through each other, all chromosomes should reach the state of topological equilibrium. The topological equilibrium in highly crowded interphase chromosomes forming chromosome territories would result in formation of highly knotted chromatin fibres. However, Chromosome Conformation Capture (3C) methods revealed that the decay of contact probabilities with the genomic distance in interphase chromosomes is practically the same as in the crumpled globule state that is formed when long polymers condense without formation of any knots. To remove knots from highly crowded chromatin, one would need an active process that should not only provide the energy to move the system from the state of topological equilibrium but also guide topoisomerase-mediated passages in such a way that knots would be efficiently unknotted instead of making the knots even more complex. We perform coarse-grained molecular dynamics simulations of the process of chromatin loop extrusion involving knotted and catenated chromatin fibres to check whether chromatin loop extrusion may be involved in active unknotting of chromatin fibres. Our simulations show that the process of chromatin loop extrusion is ideally suited to actively unknot, decatenate and demix chromatin fibres in interphase chromosomes.
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Matityahu, Avi, and Itay Onn. "Hit the brakes – a new perspective on the loop extrusion mechanism of cohesin and other SMC complexes." Journal of Cell Science 134, no. 1 (January 1, 2021): jcs247577. http://dx.doi.org/10.1242/jcs.247577.
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ABSTRACTThe three-dimensional structure of chromatin is determined by the action of protein complexes of the structural maintenance of chromosome (SMC) family. Eukaryotic cells contain three SMC complexes, cohesin, condensin, and a complex of Smc5 and Smc6. Initially, cohesin was linked to sister chromatid cohesion, the process that ensures the fidelity of chromosome segregation in mitosis. In recent years, a second function in the organization of interphase chromatin into topologically associated domains has been determined, and loop extrusion has emerged as the leading mechanism of this process. Interestingly, fundamental mechanistic differences exist between mitotic tethering and loop extrusion. As distinct molecular switches that aim to suppress loop extrusion in different biological contexts have been identified, we hypothesize here that loop extrusion is the default biochemical activity of cohesin and that its suppression shifts cohesin into a tethering mode. With this model, we aim to provide an explanation for how loop extrusion and tethering can coexist in a single cohesin complex and also apply it to the other eukaryotic SMC complexes, describing both similarities and differences between them. Finally, we present model-derived molecular predictions that can be tested experimentally, thus offering a new perspective on the mechanisms by which SMC complexes shape the higher-order structure of chromatin.
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Nuebler, Johannes, Geoffrey Fudenberg, Maxim Imakaev, Nezar Abdennur, and Leonid A. Mirny. "Chromatin organization by an interplay of loop extrusion and compartmental segregation." Proceedings of the National Academy of Sciences 115, no. 29 (July 2, 2018): E6697—E6706. http://dx.doi.org/10.1073/pnas.1717730115.
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Mammalian chromatin is spatially organized at many scales showing two prominent features in interphase: (i) alternating regions (1–10 Mb) of active and inactive chromatin that spatially segregate into different compartments, and (ii) domains (<1 Mb), that is, regions that preferentially interact internally [topologically associating domains (TADs)] and are central to gene regulation. There is growing evidence that TADs are formed by active extrusion of chromatin loops by cohesin, whereas compartmentalization is established according to local chromatin states. Here, we use polymer simulations to examine how loop extrusion and compartmental segregation work collectively and potentially interfere in shaping global chromosome organization. A model with differential attraction between euchromatin and heterochromatin leads to phase separation and reproduces compartmentalization as observed in Hi-C. Loop extrusion, essential for TAD formation, in turn, interferes with compartmentalization. Our integrated model faithfully reproduces Hi-C data from puzzling experimental observations where altering loop extrusion also led to changes in compartmentalization. Specifically, depletion of chromatin-associated cohesin reduced TADs and revealed finer compartments, while increased processivity of cohesin strengthened large TADs and reduced compartmentalization; and depletion of the TAD boundary protein CTCF weakened TADs while leaving compartments unaffected. We reveal that these experimental perturbations are special cases of a general polymer phenomenon of active mixing by loop extrusion. Our results suggest that chromatin organization on the megabase scale emerges from competition of nonequilibrium active loop extrusion and epigenetically defined compartment structure.
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Kabirova, Evelyn, Artem Nurislamov, Artem Shadskiy, Alexander Smirnov, Andrey Popov, Pavel Salnikov, Nariman Battulin, and Veniamin Fishman. "Function and Evolution of the Loop Extrusion Machinery in Animals." International Journal of Molecular Sciences 24, no. 5 (March 6, 2023): 5017. http://dx.doi.org/10.3390/ijms24055017.
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Structural maintenance of chromosomes (SMC) complexes are essential proteins found in genomes of all cellular organisms. Essential functions of these proteins, such as mitotic chromosome formation and sister chromatid cohesion, were discovered a long time ago. Recent advances in chromatin biology showed that SMC proteins are involved in many other genomic processes, acting as active motors extruding DNA, which leads to the formation of chromatin loops. Some loops formed by SMC proteins are highly cell type and developmental stage specific, such as SMC-mediated DNA loops required for VDJ recombination in B-cell progenitors, or dosage compensation in Caenorhabditis elegans and X-chromosome inactivation in mice. In this review, we focus on the extrusion-based mechanisms that are common for multiple cell types and species. We will first describe an anatomy of SMC complexes and their accessory proteins. Next, we provide biochemical details of the extrusion process. We follow this by the sections describing the role of SMC complexes in gene regulation, DNA repair, and chromatin topology.
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Maji, Ajoy, Ranjith Padinhateeri, and Mithun K. Mitra. "Loop Extrusion in Chromatin: A Question of Time!" Biophysical Journal 118, no. 3 (February 2020): 63a. http://dx.doi.org/10.1016/j.bpj.2019.11.522.
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Brandão, Hugo B., Payel Paul, Aafke A. van den Berg, David Z. Rudner, Xindan Wang, and Leonid A. Mirny. "RNA polymerases as moving barriers to condensin loop extrusion." Proceedings of the National Academy of Sciences 116, no. 41 (September 23, 2019): 20489–99. http://dx.doi.org/10.1073/pnas.1907009116.
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To separate replicated sister chromatids during mitosis, eukaryotes and prokaryotes have structural maintenance of chromosome (SMC) condensin complexes that were recently shown to organize chromosomes by a process known as DNA loop extrusion. In rapidly dividing bacterial cells, the process of separating sister chromatids occurs concomitantly with ongoing transcription. How transcription interferes with the condensin loop-extrusion process is largely unexplored, but recent experiments have shown that sites of high transcription may directionally affect condensin loop extrusion. We quantitatively investigate different mechanisms of interaction between condensin and elongating RNA polymerases (RNAPs) and find that RNAPs are likely steric barriers that can push and interact with condensins. Supported by chromosome conformation capture and chromatin immunoprecipitation for cells after transcription inhibition and RNAP degradation, we argue that translocating condensins must bypass transcribing RNAPs within ∼1 to 2 s of an encounter at rRNA genes and within ∼10 s at protein-coding genes. Thus, while individual RNAPs have little effect on the progress of loop extrusion, long, highly transcribed operons can significantly impede the extrusion process. Our data and quantitative models further suggest that bacterial condensin loop extrusion occurs by 2 independent, uncoupled motor activities; the motors translocate on DNA in opposing directions and function together to enlarge chromosomal loops, each independently bypassing steric barriers in their path. Our study provides a quantitative link between transcription and 3D genome organization and proposes a mechanism of interactions between SMC complexes and elongating transcription machinery relevant from bacteria to higher eukaryotes.
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Yamamoto, Tetsuya, Takahiro Sakaue, and Helmut Schiessel. "Slow chromatin dynamics enhances promoter accessibility to transcriptional condensates." Nucleic Acids Research 49, no. 9 (April 22, 2021): 5017–27. http://dx.doi.org/10.1093/nar/gkab275.
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Abstract Enhancers are DNA sequences at a long genomic distance from target genes. Recent experiments suggest that enhancers are anchored to the surfaces of condensates of transcription machinery and that the loop extrusion process enhances the transcription level of their target genes. Here, we theoretically study the polymer dynamics driven by the loop extrusion of the linker DNA between an enhancer and the promoter of its target gene to calculate the contact probability of the promoter to the transcription machinery in the condensate. Our theory predicts that when the loop extrusion process is active, the contact probability increases with increasing linker DNA length. This finding reflects the fact that the relaxation time, with which the promoter stays in proximity to the surface of the transcriptional condensate, increases as the length of the linker DNA increases. This contrasts the equilibrium case for which the contact probability between the promoter and the transcription machineries is smaller for longer linker DNA lengths.
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Bonato, A., C. A. Brackley, J. Johnson, D. Michieletto, and D. Marenduzzo. "Chromosome compaction and chromatin stiffness enhance diffusive loop extrusion by slip-link proteins." Soft Matter 16, no. 9 (2020): 2406–14. http://dx.doi.org/10.1039/c9sm01875a.
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Kolbin, Daniel, Benjamin L. Walker, Caitlin Hult, John Donoghue Stanton, David Adalsteinsson, M. Gregory Forest, and Kerry Bloom. "Polymer Modeling Reveals Interplay between Physical Properties of Chromosomal DNA and the Size and Distribution of Condensin-Based Chromatin Loops." Genes 14, no. 12 (December 9, 2023): 2193. http://dx.doi.org/10.3390/genes14122193.
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Transient DNA loops occur throughout the genome due to thermal fluctuations of DNA and the function of SMC complex proteins such as condensin and cohesin. Transient crosslinking within and between chromosomes and loop extrusion by SMCs have profound effects on high-order chromatin organization and exhibit specificity in cell type, cell cycle stage, and cellular environment. SMC complexes anchor one end to DNA with the other extending some distance and retracting to form a loop. How cells regulate loop sizes and how loops distribute along chromatin are emerging questions. To understand loop size regulation, we employed bead–spring polymer chain models of chromatin and the activity of an SMC complex on chromatin. Our study shows that (1) the stiffness of the chromatin polymer chain, (2) the tensile stiffness of chromatin crosslinking complexes such as condensin, and (3) the strength of the internal or external tethering of chromatin chains cooperatively dictate the loop size distribution and compaction volume of induced chromatin domains. When strong DNA tethers are invoked, loop size distributions are tuned by condensin stiffness. When DNA tethers are released, loop size distributions are tuned by chromatin stiffness. In this three-way interaction, the presence and strength of tethering unexpectedly dictates chromatin conformation within a topological domain.
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Rusková, Renáta, and Dušan Račko. "Entropic Competition between Supercoiled and Torsionally Relaxed Chromatin Fibers Drives Loop Extrusion through Pseudo-Topologically Bound Cohesin." Biology 10, no. 2 (February 7, 2021): 130. http://dx.doi.org/10.3390/biology10020130.
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We propose a model for cohesin-mediated loop extrusion, where the loop extrusion is driven entropically by the energy difference between supercoiled and torsionally relaxed chromatin fibers. Different levels of negative supercoiling are controlled by varying imposed friction between the cohesin ring and the chromatin fiber. The speed of generation of negative supercoiling by RNA polymerase associated with TOP1 is kept constant and corresponds to 10 rotations per second. The model was tested by coarse-grained molecular simulations for a wide range of frictions between 2 to 200 folds of that of generic fiber and the surrounding medium. The higher friction allowed for the accumulation of higher levels of supercoiling, while the resulting extrusion rate also increased. The obtained extrusion rates for the given range of investigated frictions were between 1 and 10 kbps, but also a saturation of the rate at high frictions was observed. The calculated contact maps indicate a qualitative improvement obtained at lower levels of supercoiling. The fits of mathematical equations qualitatively reproduce the loop sizes and levels of supercoiling obtained from simulations and support the proposed mechanism of entropically driven extrusion. The cohesin ring is bound on the fibers pseudo-topologically, and the model suggests that the topological binding is not necessary.
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Davidson, Iain F., Benedikt Bauer, Daniela Goetz, Wen Tang, Gordana Wutz, and Jan-Michael Peters. "DNA loop extrusion by human cohesin." Science 366, no. 6471 (November 21, 2019): 1338–45. http://dx.doi.org/10.1126/science.aaz3418.
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Eukaryotic genomes are folded into loops and topologically associating domains, which contribute to chromatin structure, gene regulation, and gene recombination. These structures depend on cohesin, a ring-shaped DNA-entrapping adenosine triphosphatase (ATPase) complex that has been proposed to form loops by extrusion. Such an activity has been observed for condensin, which forms loops in mitosis, but not for cohesin. Using biochemical reconstitution, we found that single human cohesin complexes form DNA loops symmetrically at rates up to 2.1 kilo–base pairs per second. Loop formation and maintenance depend on cohesin’s ATPase activity and on NIPBL-MAU2, but not on topological entrapment of DNA by cohesin. During loop formation, cohesin and NIPBL-MAU2 reside at the base of loops, which indicates that they generate loops by extrusion. Our results show that cohesin and NIPBL-MAU2 form an active holoenzyme that interacts with DNA either pseudo-topologically or non-topologically to extrude genomic interphase DNA into loops.
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Brahmachari, Sumitabha, and John F. Marko. "Chromosome disentanglement driven via optimal compaction of loop-extruded brush structures." Proceedings of the National Academy of Sciences 116, no. 50 (November 22, 2019): 24956–65. http://dx.doi.org/10.1073/pnas.1906355116.
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Eukaryote cell division features a chromosome compaction–decompaction cycle that is synchronized with their physical and topological segregation. It has been proposed that lengthwise compaction of chromatin into mitotic chromosomes via loop extrusion underlies the compaction-segregation/resolution process. We analyze this disentanglement scheme via considering the chromosome to be a succession of DNA/chromatin loops—a polymer “brush”—where active extrusion of loops controls the brush structure. Given type-II DNA topoisomerase (Topo II)-catalyzed topology fluctuations, we find that interchromosome entanglements are minimized for a certain “optimal” loop that scales with the chromosome size. The optimal loop organization is in accord with experimental data across species, suggesting an important structural role of genomic loops in maintaining a less entangled genome. Application of the model to the interphase genome indicates that active loop extrusion can maintain a level of chromosome compaction with suppressed entanglements; the transition to the metaphase state requires higher lengthwise compaction and drives complete topological segregation. Optimized genomic loops may provide a means for evolutionary propagation of gene-expression patterns while simultaneously maintaining a disentangled genome. We also find that compact metaphase chromosomes have a densely packed core along their cylindrical axes that explains their observed mechanical stiffness. Our model connects chromosome structural reorganization to topological resolution through the cell cycle and highlights a mechanism of directing Topo II-mediated strand passage via loop extrusion-driven lengthwise compaction.
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Yamamoto, Tetsuya, and Helmut Schiessel. "Dilution of contact frequency between superenhancers by loop extrusion at interfaces." Soft Matter 15, no. 38 (2019): 7635–43. http://dx.doi.org/10.1039/c9sm01454c.
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Zhang, Xuefei, Yu Zhang, Zhaoqing Ba, Nia Kyritsis, Rafael Casellas, and Frederick W. Alt. "Fundamental roles of chromatin loop extrusion in antibody class switching." Nature 575, no. 7782 (October 30, 2019): 385–89. http://dx.doi.org/10.1038/s41586-019-1723-0.
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Nuebler, Johannes, Geoffrey Fudenberg, Maxim Imakaev, Nezar Abdennur, and Leonid Mirny. "Chromatin Organization by an Interplay of Loop Extrusion and Compartmental Segregation." Biophysical Journal 114, no. 3 (February 2018): 30a. http://dx.doi.org/10.1016/j.bpj.2017.11.211.
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Matthews, Nicholas E., and Rob White. "Chromatin Architecture in the Fly: Living without CTCF/Cohesin Loop Extrusion?" BioEssays 41, no. 9 (July 2019): 1900048. http://dx.doi.org/10.1002/bies.201900048.
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Ochs, Fena, Charlotte Green, Aleksander Tomasz Szczurek, Lior Pytowski, Sofia Kolesnikova, Jill Brown, Daniel Wolfram Gerlich, Veronica Buckle, Lothar Schermelleh, and Kim Ashley Nasmyth. "Sister chromatid cohesion is mediated by individual cohesin complexes." Science 383, no. 6687 (March 8, 2024): 1122–30. http://dx.doi.org/10.1126/science.adl4606.
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Eukaryotic genomes are organized by loop extrusion and sister chromatid cohesion, both mediated by the multimeric cohesin protein complex. Understanding how cohesin holds sister DNAs together, and how loss of cohesion causes age-related infertility in females, requires knowledge as to cohesin’s stoichiometry in vivo. Using quantitative super-resolution imaging, we identified two discrete populations of chromatin-bound cohesin in postreplicative human cells. Whereas most complexes appear dimeric, cohesin that localized to sites of sister chromatid cohesion and associated with sororin was exclusively monomeric. The monomeric stoichiometry of sororin:cohesin complexes demonstrates that sister chromatid cohesion is conferred by individual cohesin rings, a key prediction of the proposal that cohesion arises from the co-entrapment of sister DNAs.
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Koide, Hiroki, Noriyuki Kodera, Shveta Bisht, Shoji Takada, and Tsuyoshi Terakawa. "Modeling of DNA binding to the condensin hinge domain using molecular dynamics simulations guided by atomic force microscopy." PLOS Computational Biology 17, no. 7 (July 30, 2021): e1009265. http://dx.doi.org/10.1371/journal.pcbi.1009265.
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The condensin protein complex compacts chromatin during mitosis using its DNA-loop extrusion activity. Previous studies proposed scrunching and loop-capture models as molecular mechanisms for the loop extrusion process, both of which assume the binding of double-strand (ds) DNA to the hinge domain formed at the interface of the condensin subunits Smc2 and Smc4. However, how the hinge domain contacts dsDNA has remained unknown. Here, we conducted atomic force microscopy imaging of the budding yeast condensin holo-complex and used this data as basis for coarse-grained molecular dynamics simulations to model the hinge structure in a transient open conformation. We then simulated the dsDNA binding to open and closed hinge conformations, predicting that dsDNA binds to the outside surface when closed and to the outside and inside surfaces when open. Our simulations also suggested that the hinge can close around dsDNA bound to the inside surface. Based on these simulation results, we speculate that the conformational change of the hinge domain might be essential for the dsDNA binding regulation and play roles in condensin-mediated DNA-loop extrusion.
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Dekker, Bastiaan, and Job Dekker. "Regulation of the mitotic chromosome folding machines." Biochemical Journal 479, no. 20 (October 21, 2022): 2153–73. http://dx.doi.org/10.1042/bcj20210140.
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Over the last several years enormous progress has been made in identifying the molecular machines, including condensins and topoisomerases that fold mitotic chromosomes. The discovery that condensins generate chromatin loops through loop extrusion has revolutionized, and energized, the field of chromosome folding. To understand how these machines fold chromosomes with the appropriate dimensions, while disentangling sister chromatids, it needs to be determined how they are regulated and deployed. Here, we outline the current understanding of how these machines and factors are regulated through cell cycle dependent expression, chromatin localization, activation and inactivation through post-translational modifications, and through associations with each other, with other factors and with the chromatin template itself. There are still many open questions about how condensins and topoisomerases are regulated but given the pace of progress in the chromosome folding field, it seems likely that many of these will be answered in the years ahead.
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Ghosh, Surya K., and Daniel Jost. "Genome organization via loop extrusion, insights from polymer physics models." Briefings in Functional Genomics 19, no. 2 (November 8, 2019): 119–27. http://dx.doi.org/10.1093/bfgp/elz023.
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Abstract Understanding how genomes fold and organize is one of the main challenges in modern biology. Recent high-throughput techniques like Hi-C, in combination with cutting-edge polymer physics models, have provided access to precise information on 3D chromosome folding to decipher the mechanisms driving such multi-scale organization. In particular, structural maintenance of chromosome (SMC) proteins play an important role in the local structuration of chromatin, putatively via a loop extrusion process. Here, we review the different polymer physics models that investigate the role of SMCs in the formation of topologically associated domains (TADs) during interphase via the formation of dynamic loops. We describe the main physical ingredients, compare them and discuss their relevance against experimental observations.
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Cutts, Erin E., and Alessandro Vannini. "Condensin complexes: understanding loop extrusion one conformational change at a time." Biochemical Society Transactions 48, no. 5 (October 2, 2020): 2089–100. http://dx.doi.org/10.1042/bst20200241.
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Condensin and cohesin, both members of the structural maintenance of chromosome (SMC) family, contribute to the regulation and structure of chromatin. Recent work has shown both condensin and cohesin extrude DNA loops and most likely work via a conserved mechanism. This review focuses on condensin complexes, highlighting recent in vitro work characterising DNA loop formation and protein structure. We discuss similarities between condensin and cohesin complexes to derive a possible mechanistic model, as well as discuss differences that exist between the different condensin isoforms found in higher eukaryotes.
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Phipps, Jamie, and Karine Dubrana. "DNA Repair in Space and Time: Safeguarding the Genome with the Cohesin Complex." Genes 13, no. 2 (January 22, 2022): 198. http://dx.doi.org/10.3390/genes13020198.
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DNA double-strand breaks (DSBs) are a deleterious form of DNA damage, which must be robustly addressed to ensure genome stability. Defective repair can result in chromosome loss, point mutations, loss of heterozygosity or chromosomal rearrangements, which could lead to oncogenesis or cell death. We explore the requirements for the successful repair of DNA DSBs by non-homologous end joining and homology-directed repair (HDR) mechanisms in relation to genome folding and dynamics. On the occurrence of a DSB, local and global chromatin composition and dynamics, as well as 3D genome organization and break localization within the nuclear space, influence how repair proceeds. The cohesin complex is increasingly implicated as a key regulator of the genome, influencing chromatin composition and dynamics, and crucially genome organization through folding chromosomes by an active loop extrusion mechanism, and maintaining sister chromatid cohesion. Here, we consider how this complex is now emerging as a key player in the DNA damage response, influencing repair pathway choice and efficiency.
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Zhang, Yu, Xuefei Zhang, Zhaoqing Ba, Zhuoyi Liang, Edward W. Dring, Hongli Hu, Jiangman Lou, et al. "The fundamental role of chromatin loop extrusion in physiological V(D)J recombination." Nature 573, no. 7775 (September 11, 2019): 600–604. http://dx.doi.org/10.1038/s41586-019-1547-y.
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Thomas, Naiju, Timothy E. Reznicek, Erez Lieberman Aiden, M. Jordan Rowley, Eric Wagner, and Guy Nir. "Abstract 1699: Defining the impact of aberrant transcription on the chromatin structure." Cancer Research 84, no. 6_Supplement (March 22, 2024): 1699. http://dx.doi.org/10.1158/1538-7445.am2024-1699.
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Abstract Genome organization in humans is governed by two major mechanisms: loop extrusion by cohesin and CTCF and the spatial compartmentalization of the chromosomes. These folding mechanisms have been shown to regulate several genomic functions, including gene expression. However, how transcription might regulate the chromatin structure remains a subject of many ongoing investigations. In this study, we use the Integrator protein complex as a tool to understand the impact of aberrant transcription on the 3D structure of the genome. During transcription initiation, RNA Polymerase-II (Pol-II) often pauses proximally to the promoter before further elongating transcripts. The Integrator protein complex regulates this checkpoint by its endonuclease and phosphatase activity. This protein complex is essential in premature transcription termination in hundreds of protein-coding genes. It also facilitates the synthesis of non-coding (ncRNA), such as enhancer RNAs (eRNAs) and long non-coding RNAs (lncRNAs), which have been shown to support loop extrusion domains, and their absence may disrupt these domains. Using HiC sequencing, we found that depleting Integrator proteins can interfere with forming and maintaining loop domains. This study aims to be the first to decipher the relationship between chromosome structure and Integrator protein activity and address our very little knowledge of how misregulation of transcription termination may influence the folding of chromosomes. Citation Format: Naiju Thomas, Timothy E. Reznicek, Erez Lieberman Aiden, M. Jordan Rowley, Eric Wagner, Guy Nir. Defining the impact of aberrant transcription on the chromatin structure [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1699.
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Korsak, Sevastianos, and Dariusz Plewczynski. "LoopSage: An energy-based Monte Carlo approach for the loop extrusion modeling of chromatin." Methods 223 (March 2024): 106–17. http://dx.doi.org/10.1016/j.ymeth.2024.01.015.
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Zhang, Xuefei, Hye Suk Yoon, Aimee M. Chapdelaine-Williams, Nia Kyritsis, and Frederick W. Alt. "Physiological role of the 3′IgH CBEs super-anchor in antibody class switching." Proceedings of the National Academy of Sciences 118, no. 3 (January 13, 2021): e2024392118. http://dx.doi.org/10.1073/pnas.2024392118.
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IgH class switch recombination (CSR) replaces Cμ constant region (CH) exons with one of six downstream CHs by joining transcription-targeted double-strand breaks (DSBs) in the Cμ switch (S) region to DSBs in a downstream S region. Chromatin loop extrusion underlies fundamental CSR mechanisms including 3′IgH regulatory region (3′IgHRR)-mediated S region transcription, CSR center formation, and deletional CSR joining. There are 10 consecutive CTCF-binding elements (CBEs) downstream of the 3′IgHRR, termed the “3′IgH CBEs.” Prior studies showed that deletion of eight 3′IgH CBEs did not detectably affect CSR. Here, we report that deletion of all 3′IgH CBEs impacts, to varying degrees, germline transcription and CSR of upstream S regions, except that of Sγ1. Moreover, deletion of all 3′IgH CBEs rendered the 6-kb region just downstream highly transcribed and caused sequences within to be aligned with Sμ, broken, and joined to form aberrant CSR rearrangements. These findings implicate the 3′IgH CBEs as critical insulators for focusing loop extrusion-mediated 3′IgHRR transcriptional and CSR activities on upstream CH locus targets.
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Conte, Mattia, Andrea M. Chiariello, Alex Abraham, Simona Bianco, Andrea Esposito, Mario Nicodemi, Tommaso Matteuzzi, and Francesca Vercellone. "Polymer Models of Chromatin Imaging Data in Single Cells." Algorithms 15, no. 9 (September 16, 2022): 330. http://dx.doi.org/10.3390/a15090330.
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Recent super-resolution imaging technologies enable tracing chromatin conformation with nanometer-scale precision at the single-cell level. They revealed, for example, that human chromosomes fold into a complex three-dimensional structure within the cell nucleus that is essential to establish biological activities, such as the regulation of the genes. Yet, to decode from imaging data the molecular mechanisms that shape the structure of the genome, quantitative methods are required. In this review, we consider models of polymer physics of chromosome folding that we benchmark against multiplexed FISH data available in human loci in IMR90 fibroblast cells. By combining polymer theory, numerical simulations and machine learning strategies, the predictions of the models are validated at the single-cell level, showing that chromosome structure is controlled by the interplay of distinct physical processes, such as active loop-extrusion and thermodynamic phase-separation.
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Brandão, Hugo B., Johanna Gassler, Maxim Imakaev, Ilya M. Flyamer, Sabrina Ladstätter, Wendy A. Bickmore, Jan-Michael Peters, Kikuë Tachibana-Konwalski, and Leonid A. Mirny. "A Mechanism of Cohesin-Dependent Loop Extrusion Organizes Mammalian Chromatin Structure in the Developing Embryo." Biophysical Journal 114, no. 3 (February 2018): 255a. http://dx.doi.org/10.1016/j.bpj.2017.11.1417.
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Sanborn, Adrian L., Suhas S. P. Rao, Su-Chen Huang, Neva C. Durand, Miriam H. Huntley, Andrew I. Jewett, Ivan D. Bochkov, et al. "Chromatin extrusion explains key features of loop and domain formation in wild-type and engineered genomes." Proceedings of the National Academy of Sciences 112, no. 47 (October 23, 2015): E6456—E6465. http://dx.doi.org/10.1073/pnas.1518552112.
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We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these maps with new Hi-C, microscopy, and genome-editing experiments to study the physical structure of chromatin fibers, domains, and loops. We find that the observed contact domains are inconsistent with the equilibrium state for an ordinary condensed polymer. Combining Hi-C data and novel mathematical theorems, we show that contact domains are also not consistent with a fractal globule. Instead, we use physical simulations to study two models of genome folding. In one, intermonomer attraction during polymer condensation leads to formation of an anisotropic “tension globule.” In the other, CCCTC-binding factor (CTCF) and cohesin act together to extrude unknotted loops during interphase. Both models are consistent with the observed contact domains and with the observation that contact domains tend to form inside loops. However, the extrusion model explains a far wider array of observations, such as why loops tend not to overlap and why the CTCF-binding motifs at pairs of loop anchors lie in the convergent orientation. Finally, we perform 13 genome-editing experiments examining the effect of altering CTCF-binding sites on chromatin folding. The convergent rule correctly predicts the affected loops in every case. Moreover, the extrusion model accurately predicts in silico the 3D maps resulting from each experiment using only the location of CTCF-binding sites in the WT. Thus, we show that it is possible to disrupt, restore, and move loops and domains using targeted mutations as small as a single base pair.
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Conte, Mattia, Andrea Esposito, Francesca Vercellone, Alex Abraham, and Simona Bianco. "Unveiling the Machinery behind Chromosome Folding by Polymer Physics Modeling." International Journal of Molecular Sciences 24, no. 4 (February 11, 2023): 3660. http://dx.doi.org/10.3390/ijms24043660.
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Understanding the mechanisms underlying the complex 3D architecture of mammalian genomes poses, at a more fundamental level, the problem of how two or multiple genomic sites can establish physical contacts in the nucleus of the cells. Beyond stochastic and fleeting encounters related to the polymeric nature of chromatin, experiments have revealed specific, privileged patterns of interactions that suggest the existence of basic organizing principles of folding. In this review, we focus on two major and recently proposed physical processes of chromatin organization: loop-extrusion and polymer phase-separation, both supported by increasing experimental evidence. We discuss their implementation into polymer physics models, which we test against available single-cell super-resolution imaging data, showing that both mechanisms can cooperate to shape chromatin structure at the single-molecule level. Next, by exploiting the comprehension of the underlying molecular mechanisms, we illustrate how such polymer models can be used as powerful tools to make predictions in silico that can complement experiments in understanding genome folding. To this aim, we focus on recent key applications, such as the prediction of chromatin structure rearrangements upon disease-associated mutations and the identification of the putative chromatin organizing factors that orchestrate the specificity of DNA regulatory contacts genome-wide.
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Zhang, Yu, Xuefei Zhang, Zhuoyi Liang, Zhaoqing Ba, Eddie Dring, Jeffrey Zurita, Aviva Presser Aiden, Erez Lieberman Aiden, and Frederick W. Alt. "Physiological V(D)J Recombination is Mediated by RAG Scanning of Loop-extruded Chromatin." Journal of Immunology 202, no. 1_Supplement (May 1, 2019): 123.18. http://dx.doi.org/10.4049/jimmunol.202.supp.123.18.
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Abstract RAG endonuclease initiates V(D)J recombination by cleaving paired V, D, and J gene segments flanked by complementary recombination signal sequences (RSSs). We address physiological relevance of RAG scanning and the underlying mechanisms through studies of the ~300 kb 3′-proximal IgH locus CBE-anchored loop domain. This domain contains nine Ds and four JHs within a 57-kb region between the upstream IGCR1 loop anchor and the V(D)J recombination center (RC). IgH D to JH-rearrangements occur in deletional orientation, mediated by convergent downstream D-RSSs and JH-RSSs, despite Ds having upstream RSSs in the same orientation as JH-RSSs that could promote inversion. Extra-chromosomal V(D)J recombination substrate studies attributed this deletional orientation-specific joining bias to D-RSS sequence differences. However, we find that deletional joining of 8 of 9 chromosomal IgH Ds, results from RAG linear scanning of chromatin upstream of the RAG-bound RC, a process that detects convergent D-RSSs but to which D-RSSs in the same orientation are invisible. The exceptional D employs an RSS-based mechanism to counteract proximal location to JHs in the RC. By deleting JHs, we reveal that RAG scans directionally downstream from 3′D-RSSs until reaching the IgH 3′CBE loop anchor. Targeting catalytic-dead Cas9 (dCas9) binding in this scanning path impedes RAG scanning in association with cohesin accumulation and loop formation between the block site and RC. Strong transcription through a repetitive downstream switch region blocks RAG scanning similarly to the dCas9 block. We demonstrate a critical role for RAG scanning in physiological V(D)J recombination and implicate cohesin-mediated loop extrusion as a driving force.
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Khabarova, A. A., A. S. Ryzhkova, and N. R. Battulin. "Reorganisation of chromatin during erythroid differentiation." Vavilov Journal of Genetics and Breeding 23, no. 1 (February 26, 2019): 95–99. http://dx.doi.org/10.18699/vj19.467.
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A totipotent zygote has unlimited potential for differentiation into all cell types found in an adult organism. During ontogenesis proliferating and maturing cells gradually lose their differentiation potential, limiting the spectrum of possible developmental transitions to a specific cell type. Following the initiation of the developmental program cells acquire specific morphological and functional properties. Deciphering the mechanisms that coordinate shifts in gene expression revealed a critical role of three-dimensional chromatin structure in the regulation of gene activity during lineage commitment. Several levels of DNA packaging have been recently identified using chromosome conformation capture based techniques such a Hi-C. It is now clear that chromatin regions with high transcriptional activity assemble into Mb-scale compartments in the nuclear space, distinct from transcriptionally silent regions. More locally chromatin is organized into topological domains, serving as functionally insulated units with cell type – specific regulatory loop interactions. However, molecular mechanisms establishing and maintaining such 3D organization are yet to be investigated. Recent focus on studying chromatin reorganization accompanying cell cycle progression and cellular differentiation partially explained some aspects of 3D genome folding. Throughout erythropoiesis cells undergo a dramatic reorganization of the chromatin landscape leading to global nuclear condensation and transcriptional silencing, followed by nuclear extrusion at the final stage of mammalian erythropoiesis. Drastic changes of genome architecture and function accompanying erythroid differentiation seem to be an informative model for studying the ways of how genome organization and dynamic gene activity are connected. Here we summarize current views on the role of global rearrangement of 3D chromatin structure in erythroid differentiation.
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Jeong, Mira, Xiangfan Huang, Xiaotian Zhang, Jianzhong Su, Muhammad S. Shamim, Ivan D. Bochkov, Jaime M. Reyes, et al. "Large DNA Methylation Canyons Anchor Chromatin Loops Maintaining Hematopoietic Stem Cell Identity." Blood 132, Supplement 1 (November 29, 2018): 534. http://dx.doi.org/10.1182/blood-2018-99-119485.
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Abstract Higher order chromatin structure and DNA methylation are implicated in multiple developmental processes, but their relationship to cell state is unknown. In order to understand how the DNA methylation is connected with nuclear architecture and can vary between cell types and during cell differentiation, we began to explore the 3D architecture of human hematopoietic stem and progenitor cells (HSPCs) by performing in situ Hi-C experiments at 5kb resolution. We found that large (~10kb) DNA methylation canyons can form long loops connecting anchor loci that may be dozens of megabases apart. These canyons also can form interchromosomal links (Fig.1a and 1b). We further confirmed these long-range interactions by performing 3D-FISH using two color fluorescent labeled probes that spanned the HOXA locus loop anchor (green) and the SP8 locus loop anchor (red), which are ~7MB apart (Fig. 1c). In order to begin to investigate mechanisms that may regulate these long loops and how they relate to commonly studied loops that are mediated by CTCF-extrusion, we examined their properties systematically. Interestingly, the anchors of long loops exhibited minimal enrichment for CTCF (1.04-fold), and, even when CTCF was bound, they did not obey the convergent rule. The data suggest these loops are formed by phase separation of the interacting loci to form a genomic subcompartment, rather than by CTCF-mediated extrusion. Next, we sought to determine whether other features correlated with these long loops. By aligning DNA methylation profiles with the Hi-C data, we observed that anchors often corresponded to regions of very low DNA methylation, and thus sought to analyze the relationship in detail. We found that the anchor position of the long loops had lower average DNA methylation levels than standard loop anchors and very often overlapped with DNA methylation canyons. Canyons are typically decorated with either active or repressive histone marks. We considered whether a particular group of canyons was associated with the long loops. Our findings further indicate that repressed regions marked by the polycomb-mediated histone modification H3K27me3 at DNA methylation canyons generally mediate the formation of canyon loops. Next, we considered whether the long loops associated with repressive grand canyons that we had annotated in HSPCs were present in other cell types. Using Aggregate Peak Analysis (APA), a computational strategy in which the Hi-C submatrices from the vicinity of multiple putative loops are superimposed, we examined 19 human cell types and 10 murine cell types in which loop-resolution Hi-C maps are available. Interestingly, unlike previously characterized genomic subcompartments, these long-range loops are only present in stem and progenitor cells, but not in differentiated cell types, such as T cells and erythroid progenitors (Fig. 1d). Further, we identified one particular loop anchor that lay at the anchor of a long loop and contained no apparent genes ("geneless" canyon, or "GLS"). The GLS harboring this anchor is 17 kb long, lies 1.4 Mb upstream of the HOXA1 gene, and forms long loops with a 28 kb grand canyon in the HOXA region. In order to understand the role of the GLS region in hematopoietic stem cells (HSCs), we deleted the GLS in HSPCs using Cas9-mediated editing and assayed the edited cells for their ability to form colonies. Strikingly, after deleting the GLS, the number of colonies and their size was greatly reduced in edited cells compared to control experiments using either random guide RNAs or electroporation only (Fig. 1e). After ex vivo culture, the overwhelming majority of GLS-knock out HSPCs acquired the marker CD38, indicating that they were differentiating. Similarly, HOXA gene expression, an indicator of HSPC function, was greatly diminished after GLS deletion compared to control cells. These data indicate that the GLS identified in our study is functionally associated with maintenance of the HSC state. Overall, our work reveals long-range interactions between H3K27me3-marked DNA methylation canyons comprising a novel microcompartment associated with cellular identity. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.
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Racko, Dusan, Fabrizio Benedetti, Julien Dorier, and Andrzej Stasiak. "Transcription-induced supercoiling as the driving force of chromatin loop extrusion during formation of TADs in interphase chromosomes." Nucleic Acids Research 46, no. 4 (November 13, 2017): 1648–60. http://dx.doi.org/10.1093/nar/gkx1123.
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Yin, Zihang, Shuang Cui, Song Xue, Yufan Xie, Yefan Wang, Chengling Zhao, Zhiyu Zhang, et al. "Identification of Two Subsets of Subcompartment A1 Associated with High Transcriptional Activity and Frequent Loop Extrusion." Biology 12, no. 8 (July 27, 2023): 1058. http://dx.doi.org/10.3390/biology12081058.
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Three-dimensional genome organization has been increasingly recognized as an important determinant of the precise regulation of gene expression in mammalian cells, yet the relationship between gene transcriptional activity and spatial subcompartment positioning is still not fully comprehended. Here, we first utilized genome-wide Hi-C data to infer eight types of subcompartment (labeled A1, A2, A3, A4, B1, B2, B3, and B4) in mouse embryonic stem cells and four primary differentiated cell types, including thymocytes, macrophages, neural progenitor cells, and cortical neurons. Transitions of subcompartments may confer gene expression changes in different cell types. Intriguingly, we identified two subsets of subcompartments defined by higher gene density and characterized by strongly looped contact domains, named common A1 and variable A1, respectively. We revealed that common A1, which includes highly expressed genes and abundant housekeeping genes, shows a ~2-fold higher gene density than the variable A1, where cell type-specific genes are significantly enriched. Thus, our study supports a model in which both types of genomic loci with constitutive and regulatory high transcriptional activity can drive the subcompartment A1 formation. Special chromatin subcompartment arrangement and intradomain interactions may, in turn, contribute to maintaining proper levels of gene expression, especially for regulatory non-housekeeping genes.
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Luppino, Jennifer M., Andrew Field, Son C. Nguyen, Daniel S. Park, Parisha P. Shah, Richard J. Abdill, Yemin Lan, et al. "Co-depletion of NIPBL and WAPL balance cohesin activity to correct gene misexpression." PLOS Genetics 18, no. 11 (November 30, 2022): e1010528. http://dx.doi.org/10.1371/journal.pgen.1010528.
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The relationship between cohesin-mediated chromatin looping and gene expression remains unclear. NIPBL and WAPL are two opposing regulators of cohesin activity; depletion of either is associated with changes in both chromatin folding and transcription across a wide range of cell types. However, a direct comparison of their individual and combined effects on gene expression in the same cell type is lacking. We find that NIPBL or WAPL depletion in human HCT116 cells each alter the expression of ~2,000 genes, with only ~30% of the genes shared between the conditions. We find that clusters of differentially expressed genes within the same topologically associated domain (TAD) show coordinated misexpression, suggesting some genomic domains are especially sensitive to both more or less cohesin. Finally, co-depletion of NIPBL and WAPL restores the majority of gene misexpression as compared to either knockdown alone. A similar set of NIPBL-sensitive genes are rescued following CTCF co-depletion. Together, this indicates that altered transcription due to reduced cohesin activity can be functionally offset by removal of either its negative regulator (WAPL) or the physical barriers (CTCF) that restrict loop-extrusion events.
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Vitriolo, Alessandro, Michele Gabriele, and Giuseppe Testa. "From enhanceropathies to the epigenetic manifold underlying human cognition." Human Molecular Genetics 28, R2 (August 14, 2019): R226—R234. http://dx.doi.org/10.1093/hmg/ddz196.
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Abstract A vast portion of intellectual disability and autism spectrum disorders is genetically caused by mutations in chromatin modulators. These proteins play key roles in development and are also highly expressed in the adult brain. Specifically, the pivotal role of chromatin regulation in transcription has placed enhancers at the core of neurodevelopmental disorders (NDDs) studies, ushering in the coining of the term enhanceropathies. The convergence of these disorders is multilayered, spanning from molecular causes to pathophysiological traits, including extensive overlaps between enhanceropathies and neurocristopathies. The reconstruction of epigenetic circuitries wiring development and underlying cognitive functions has gone hand in hand with the development of tools that increase the sensitivity of identifying regulatory regions and linking enhancers to their target genes. The available models, including loop extrusion and phase separation, have been bringing into relief complementary aspects to interpret gene regulation datasets, reinforcing the idea that enhancers are not all the same and that regulatory regions possess shades of enhancer-ness and promoter-ness. The current limits in enhancer definition, within the emerging broader understanding of chromatin dynamics in time and space, are now on the verge of being transformed by the possibility to interrogate developmentally relevant three-dimensional cellular models at single-cell resolution. Here we discuss the contours of how these technological advances, as well as the epistemic limitations they are set to overcome, may well usher in a change of paradigm for NDDs, moving the quest for convergence from enhancers to the four-dimensional (4D) genome.
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Orlandini, Enzo, Davide Marenduzzo, and Davide Michieletto. "Synergy of topoisomerase and structural-maintenance-of-chromosomes proteins creates a universal pathway to simplify genome topology." Proceedings of the National Academy of Sciences 116, no. 17 (April 8, 2019): 8149–54. http://dx.doi.org/10.1073/pnas.1815394116.
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Topological entanglements severely interfere with important biological processes. For this reason, genomes must be kept unknotted and unlinked during most of a cell cycle. Type II topoisomerase (TopoII) enzymes play an important role in this process but the precise mechanisms yielding systematic disentanglement of DNA in vivo are not clear. Here we report computational evidence that structural-maintenance-of-chromosomes (SMC) proteins—such as cohesins and condensins—can cooperate with TopoII to establish a synergistic mechanism to resolve topological entanglements. SMC-driven loop extrusion (or diffusion) induces the spatial localization of essential crossings, in turn catalyzing the simplification of knots and links by TopoII enzymes even in crowded and confined conditions. The mechanism we uncover is universal in that it does not qualitatively depend on the specific substrate, whether DNA or chromatin, or on SMC processivity; we thus argue that this synergy may be at work across organisms and throughout the cell cycle.
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Mao, Albert, Carrie Chen, Stephanie Portillo-Ledesma, and Tamar Schlick. "Effect of Single-Residue Mutations on CTCF Binding to DNA: Insights from Molecular Dynamics Simulations." International Journal of Molecular Sciences 24, no. 7 (March 29, 2023): 6395. http://dx.doi.org/10.3390/ijms24076395.
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In humans and other eukaryotes, DNA is condensed into chromatin fibers that are further wound into chromosomes. This organization allows regulatory elements in the genome, often distant from each other in the linear DNA, to interact and facilitate gene expression through regions known as topologically associating domains (TADs). CCCTC–binding factor (CTCF) is one of the major components of TAD formation and is responsible for recruiting a partner protein, cohesin, to perform loop extrusion and facilitate proper gene expression within TADs. Because single-residue CTCF mutations have been linked to the development of a variety of cancers in humans, we aim to better understand how these mutations affect the CTCF structure and its interaction with DNA. To this end, we compare all-atom molecular dynamics simulations of a wildtype CTCF–DNA complex to those of eight different cancer-linked CTCF mutant sequences. We find that most mutants have lower binding energies compared to the wildtype protein, leading to the formation of less stable complexes. Depending on the type and position of the mutation, this loss of stability can be attributed to major changes in the electrostatic potential, loss of hydrogen bonds between the CTCF and DNA, and/or destabilization of specific zinc fingers. Interestingly, certain mutations in specific fingers can affect the interaction with the DNA of other fingers, explaining why mere single mutations can impair CTCF function. Overall, these results shed mechanistic insights into experimental observations and further underscore CTCF’s importance in the regulation of chromatin architecture and gene expression.
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Aiden, Erez Lieberman. "Three-D Codes in the Human Genome." Blood 134, Supplement_1 (November 13, 2019): SCI—50—SCI—50. http://dx.doi.org/10.1182/blood-2019-121474.
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Stretched out from end-to-end, the human genome -a sequence of 3 billion chemical letters inscribed in a molecule called DNA -is over 2 meters long. Famously, short stretches of DNA fold into a double helix, which wind around histone proteins to form the 10nm fiber. But what about longer pieces? Does the genome's fold influence function? How does the information contained in such an ultra-dense packing even remain accessible?In this talk, I describe our work developing 'Hi-C' (Lieberman-Aiden et al., Science, 2009; Aiden, Science, 2011) and more recently 'in-situHi-C' (Rao & Huntley et al., Cell, 2014), which use proximity ligation to transform pairs of physically adjacent DNA loci into chimeric DNA sequences. Sequencing a library of such chimeras makes it possible to create genome-wide maps of physical contacts between pairs of loci, revealing features ofgenome folding in 3D. Next, I will describe recent work using in situ Hi-C to construct haploid and diploid maps of nine cell types. The densest, in human lymphoblastoid cells, contains 4.9 billion contacts, achieving 1 kb resolution. We find that genomes are partitioned into contact domains (median length, 185 kb), which are associated with distinct patterns of histone marks and segregate into six subcompartments. We identify ∼10,000 loops. These loops frequently link promoters and enhancers, correlate with gene activation, and show conservation across cell types and species. Loop anchors typically occur at domain boundaries and bind the protein CTCF. The CTCF motifs at loop anchors occur predominantly (>90%) in a convergent orientation, with the asymmetric motifs "facing" one another. Next, I will discuss the biophysical mechanism that underlies chromatin looping. Specifically, our data is consistent with the formation of loops by extrusion (Sanborn & Rao et al., PNAS, 2015). In fact, in many cases, the local structure of Hi-C maps may be predicted in silicobased on patterns of CTCF binding and an extrusion-based model. Finally, I will show that by modifying CTCF motifs using CRISPR, we can reliably add, move, and delete loops and domains. Thus, it possible not only to "read" the genome's 3D architecture, but also to write it. Disclosures No relevant conflicts of interest to declare.
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Subramanian, Vijayalakshmi V. "Preprint Highlight: Cohesin mediates DNA loop extrusion and sister chromatid cohesion by distinct mechanisms." Molecular Biology of the Cell 34, no. 5 (May 1, 2023). http://dx.doi.org/10.1091/mbc.p23-03-0010.
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Cohesins organize chromatin within the nucleus by promoting loop extrusion via cis contacts on chromatin and sister chromatid cohesion via trans contacts between chromatids. Whether cohesins employ a distinct mechanism for these two functions is not known. The authors use a carefully designed conditional expression of cohesin, mutated at its hinge domain, and assess its function with a variety of in vitro and in vivo experiments including live TIRF imaging, loop extrusion assay, Hi-C, and calibrated ChIP sequencing. A mutation in the hinge domain separates the two functions of cohesin, as the mutant supports loop extrusion but not sister chromatid cohesion. A unique mechanism of cohesins in promoting loop extrusion has implications for their distinct role in genome organization and for transcriptional regulation and cell fate determination. This study will be of broad interest to genome biologists with respect to these phenomena. This preprint has been assigned the following badges: New Hypothesis, New Materials. Read the preprint on bioRxiv ( Nagasaka et al., 2022 ): https://doi.org/10.1101/2022.09.23.509019 .
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Bailey, Mary Lou P., Ivan Surovtsev, Jessica F. Williams, Hao Yan, Tianyu Yuan, Kevin Li, Katherine Duseau, Simon G. J. Mochrie, and Megan C. King. "Loops and the activity of loop extrusion factors constrain chromatin dynamics." Molecular Biology of the Cell, April 26, 2023. http://dx.doi.org/10.1091/mbc.e23-04-0119.
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The chromosomes - DNA polymers and their binding proteins - are compacted into a spatially organized, yet dynamic, three-dimensional structure. Recent genome-wide chromatin conformation capture experiments reveal a hierarchical organization of the DNA structure that is imposed, at least in part, by looping interactions arising from the activity of loop extrusion factors. The dynamics of chromatin reflects the response of the polymer to a combination of thermal fluctuations and active processes. However, how chromosome structure and enzymes acting on chromatin together define its dynamics remains poorly understood. To gain insight into the structure-dynamics relationship of chromatin, we combine high-precision microscopy in living Schizosaccharomyces pombe cells with systematic genetic perturbations and Rouse-model polymer simulations. We first investigated how the activity of two loop extrusion factors, the cohesin and condensin complexes, influences chromatin dynamics. We observed that deactivating cohesin, or to a lesser extent condensin, increased chromatin mobility, suggesting that loop extrusion constrains rather than agitates chromatin motion. Our corresponding simulations reveal that the introduction of loops is sufficient to explain the constraining activity of loop extrusion factors, highlighting that the conformation adopted by the polymer plays a key role in defining its dynamics. Moreover, we find that the number loops or residence times of loop extrusion factors influences the dynamic behavior of the chromatin polymer. Last, we observe that the activity of the INO80 chromatin remodeler, but not the SWI/SNF or RSC complexes, is critical for ATP-dependent chromatin mobility in fission yeast. Taken together we suggest that thermal and INO80-dependent activities exert forces that drive chromatin fluctuations, which are constrained by the organization of the chromosome into loops.
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Yan, Hao, Ivan Surovtsev, Jessica F. Williams, Mary Lou P. Bailey, Megan C. King, and Simon G. J. Mochrie. "Extrusion of chromatin loops by a composite loop extrusion factor." Physical Review E 104, no. 2 (August 23, 2021). http://dx.doi.org/10.1103/physreve.104.024414.
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Matityahu, Avi, and Itay Onn. "Hit the brakes – a new perspective on the loop extrusion mechanism of cohesin and other SMC complexes." Journal of Cell Science 134, no. 1 (January 1, 2021). http://dx.doi.org/10.1242/jcs.247577.
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ABSTRACT The three-dimensional structure of chromatin is determined by the action of protein complexes of the structural maintenance of chromosome (SMC) family. Eukaryotic cells contain three SMC complexes, cohesin, condensin, and a complex of Smc5 and Smc6. Initially, cohesin was linked to sister chromatid cohesion, the process that ensures the fidelity of chromosome segregation in mitosis. In recent years, a second function in the organization of interphase chromatin into topologically associated domains has been determined, and loop extrusion has emerged as the leading mechanism of this process. Interestingly, fundamental mechanistic differences exist between mitotic tethering and loop extrusion. As distinct molecular switches that aim to suppress loop extrusion in different biological contexts have been identified, we hypothesize here that loop extrusion is the default biochemical activity of cohesin and that its suppression shifts cohesin into a tethering mode. With this model, we aim to provide an explanation for how loop extrusion and tethering can coexist in a single cohesin complex and also apply it to the other eukaryotic SMC complexes, describing both similarities and differences between them. Finally, we present model-derived molecular predictions that can be tested experimentally, thus offering a new perspective on the mechanisms by which SMC complexes shape the higher-order structure of chromatin.
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"Chromatin Loop Extrusion Regulates Neutrophil Differentiation." Cancer Discovery, 2024. http://dx.doi.org/10.1158/2159-8290.cd-rw2024-032.
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Banigan, Edward J., Aafke A. van den Berg, Hugo B. Brandão, John F. Marko, and Leonid A. Mirny. "Chromosome organization by one-sided and two-sided loop extrusion." eLife 9 (April 6, 2020). http://dx.doi.org/10.7554/elife.53558.
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SMC complexes, such as condensin or cohesin, organize chromatin throughout the cell cycle by a process known as loop extrusion. SMC complexes reel in DNA, extruding and progressively growing DNA loops. Modeling assuming two-sided loop extrusion reproduces key features of chromatin organization across different organisms. In vitro single-molecule experiments confirmed that yeast condensins extrude loops, however, they remain anchored to their loading sites and extrude loops in a ‘one-sided’ manner. We therefore simulate one-sided loop extrusion to investigate whether ‘one-sided’ complexes can compact mitotic chromosomes, organize interphase domains, and juxtapose bacterial chromosomal arms, as can be done by ‘two-sided’ loop extruders. While one-sided loop extrusion cannot reproduce these phenomena, variants can recapitulate in vivo observations. We predict that SMC complexes in vivo constitute effectively two-sided motors or exhibit biased loading and propose relevant experiments. Our work suggests that loop extrusion is a viable general mechanism of chromatin organization.
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Golov, Arkadiy K., Anastasia V. Golova, Alexey A. Gavrilov, and Sergey V. Razin. "Sensitivity of cohesin–chromatin association to high-salt treatment corroborates non-topological mode of loop extrusion." Epigenetics & Chromatin 14, no. 1 (July 28, 2021). http://dx.doi.org/10.1186/s13072-021-00411-w.
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AbstractCohesin is a key organizer of chromatin folding in eukaryotic cells. The two main activities of this ring-shaped protein complex are the maintenance of sister chromatid cohesion and the establishment of long-range DNA–DNA interactions through the process of loop extrusion. Although the basic principles of both cohesion and loop extrusion have been described, we still do not understand several crucial mechanistic details. One of such unresolved issues is the question of whether a cohesin ring topologically embraces DNA string(s) during loop extrusion. Here, we show that cohesin complexes residing on CTCF-occupied genomic sites in mammalian cells do not interact with DNA topologically. We assessed the stability of cohesin-dependent loops and cohesin association with chromatin in high-ionic-strength conditions in G1-synchronized HeLa cells. We found that increased salt concentration completely displaces cohesin from those genomic regions that correspond to CTCF-defined loop anchors. Unsurprisingly, CTCF-anchored cohesin loops also dissipate in these conditions. Because topologically engaged cohesin is considered to be salt resistant, our data corroborate a non-topological model of loop extrusion. We also propose a model of cohesin activity throughout the interphase, which essentially equates the termination of non-topological loop extrusion with topological loading of cohesin. This theoretical framework enables a parsimonious explanation of various seemingly contradictory experimental findings.
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Golfier, Stefan, Thomas Quail, Hiroshi Kimura, and Jan Brugués. "Cohesin and condensin extrude DNA loops in a cell cycle-dependent manner." eLife 9 (May 12, 2020). http://dx.doi.org/10.7554/elife.53885.
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Loop extrusion by structural maintenance of chromosomes (SMC) complexes has been proposed as a mechanism to organize chromatin in interphase and metaphase. However, the requirements for chromatin organization in these cell cycle phases are different, and it is unknown whether loop extrusion dynamics and the complexes that extrude DNA also differ. Here, we used Xenopus egg extracts to reconstitute and image loop extrusion of single DNA molecules during the cell cycle. We show that loops form in both metaphase and interphase, but with distinct dynamic properties. Condensin extrudes DNA loops non-symmetrically in metaphase, whereas cohesin extrudes loops symmetrically in interphase. Our data show that loop extrusion is a general mechanism underlying DNA organization, with dynamic and structural properties that are biochemically regulated during the cell cycle.
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Higashi, Torahiko L., Georgii Pobegalov, Minzhe Tang, Maxim I. Molodtsov, and Frank Uhlmann. "A Brownian ratchet model for DNA loop extrusion by the cohesin complex." eLife 10 (July 26, 2021). http://dx.doi.org/10.7554/elife.67530.
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The cohesin complex topologically encircles DNA to promote sister chromatid cohesion. Alternatively, cohesin extrudes DNA loops, thought to reflect chromatin domain formation. Here, we propose a structure-based model explaining both activities. ATP and DNA binding promote cohesin conformational changes that guide DNA through a kleisin N-gate into a DNA gripping state. Two HEAT-repeat DNA binding modules, associated with cohesin’s heads and hinge, are now juxtaposed. Gripping state disassembly, following ATP hydrolysis, triggers unidirectional hinge module movement, which completes topological DNA entry by directing DNA through the ATPase head gate. If head gate passage fails, hinge module motion creates a Brownian ratchet that, instead, drives loop extrusion. Molecular-mechanical simulations of gripping state formation and resolution cycles recapitulate experimentally observed DNA loop extrusion characteristics. Our model extends to asymmetric and symmetric loop extrusion, as well as z-loop formation. Loop extrusion by biased Brownian motion has important implications for chromosomal cohesin function.
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Chan, Brian, and Michael Rubinstein. "Activity-driven chromatin organization during interphase: Compaction, segregation, and entanglement suppression." Proceedings of the National Academy of Sciences 121, no. 21 (May 16, 2024). http://dx.doi.org/10.1073/pnas.2401494121.
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In mammalian cells, the cohesin protein complex is believed to translocate along chromatin during interphase to form dynamic loops through a process called active loop extrusion. Chromosome conformation capture and imaging experiments have suggested that chromatin adopts a compact structure with limited interpenetration between chromosomes and between chromosomal sections. We developed a theory demonstrating that active loop extrusion causes the apparent fractal dimension of chromatin to cross-over between two and four at contour lengths on the order of 30 kilo-base pairs. The anomalously high fractal dimension D = 4 is due to the inability of extruded loops to fully relax during active extrusion. Compaction on longer contour length scales extends within topologically associated domains (TADs), facilitating gene regulation by distal elements. Extrusion-induced compaction segregates TADs such that overlaps between TADs are reduced to less than 35% and increases the entanglement strand of chromatin by up to a factor of 50 to several Mega-base pairs. Furthermore, active loop extrusion couples cohesin motion to chromatin conformations formed by previously extruding cohesins and causes the mean square displacement of chromatin loci during lag times ( Δ t ) longer than tens of minutes to be proportional to Δ t 1 / 3 . We validate our results with hybrid molecular dynamics—Monte Carlo simulations and show that our theory is consistent with experimental data. This work provides a theoretical basis for the compact organization of interphase chromatin, explaining the physical reason for TAD segregation and suppression of chromatin entanglements which contribute to efficient gene regulation.