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Chromatin extrusion explains key features of loop and domain formation in wild-type and engineered genomes

ABSTRACT: We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these 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, CTCF and cohesin act together to extrude 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 loop 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.

ORGANISM(S): Homo sapiens

PROVIDER: GSE74072 | GEO | 2015/10/23

SECONDARY ACCESSION(S): PRJNA299007

REPOSITORIES: GEO

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Similar Datasets

Chromatin extrusion explains key features of loop and domain formation in wild-type and engineered genomes

Project description:We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian genomes. Here, we combine these 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, CTCF and cohesin act together to extrude 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 loop 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. in situ Hi-C and HYbrid Capture Hi-C (Hi-C2) were used to probe the three-dimensional structure of the genome in two different human cell types before and after genome editing.

2015-10-23 | E-GEOD-74072 | biostudies-arrayexpress

A three-dimensional map of the human genome at kilobase resolution reveals prinicples of chromatin looping

Project description:We use in situ Hi-C to probe the three-dimensional architecture of genomes, constructing haploid and diploid maps of nine cell types. The densest, in human lymphoblastoid cells, contains 4.9 billion contacts, achieving 1-kilobase resolution. We find that genomes are partitioned into local domains, 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 CTCF. CTCF sites at loop anchors occur predominantly (>90%) in a convergent orientation, with the asymmetric motifs ‘facing’ one another. The inactive X-chromosome splits into two massive domains and contains large loops anchored at CTCF-binding repeats. in situ Hi-C and dilution Hi-C were used to probe the three-dimensional structure of the genome in eight diverse human cell types and one mouse cell type

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Single-allele chromatin interactions identify regulatory hubs in dynamic compartmentalized domains [Hi-C]

Project description:The genome is organized into self-interacting chromatin domains containing genes and the cis-regulatory elements controlling their expression. How these domains form and how elements within them interact is unknown. We have developed Tri-C, a sensitive, high-resolution Chromosome Conformation Capture (3C) approach to identify concurrent chromatin interactions in single cells. Combining Tri-C with conventional 3C and polymer modeling, we show that, rather than folding into stable pre-formed loops, self-interacting domains form dynamic compartmentalized chromatin structures, delimited by CTCF/Cohesin boundaries. Within these tissue-specific domains, all regions of chromatin contact each other, but preferential structures are formed in which multiple enhancers and promoters interact simultaneously. Flanking CTCF/Cohesin-bound elements are excluded from these interactions and form distinct structures. These observations are best explained by a dynamic loop extrusion mechanism and subsequent stabilization of enhancer-promoter interactions, rather than the current view of long-range interactions occurring via the formation of discrete pre-formed chromatin loops.

2018-08-17 | GSE107939 | GEO

Single-allele chromatin interactions identify regulatory hubs in dynamic compartmentalized domains [Tri-C]

Project description:The genome is organized into self-interacting chromatin domains containing genes and the cis-regulatory elements controlling their expression. How these domains form and how elements within them interact is not fully understood. We have developed Tri-C, a sensitive, high-resolution Chromosome Conformation Capture (3C) approach to identify concurrent chromatin interactions in single cells. Combining Tri-C with conventional 3C and polymer modeling, we show that, rather than folding into stable pre-formed loops, self-interacting domains form dynamic compartmentalized chromatin structures, delimited by CTCF/Cohesin boundaries. Within these tissue-specific domains, all regions of chromatin contact each other, but preferential structures are formed in which multiple enhancers and promoters interact simultaneously. Flanking CTCF/Cohesin-bound elements are excluded from these interactions and form distinct structures. These observations are best explained by a dynamic loop extrusion mechanism and subsequent stabilization of enhancer-promoter interactions, rather than the current view of long-range interactions occurring via the formation of discrete pre-formed chromatin loops.

2018-08-17 | GSE107755 | GEO

Single-allele chromatin interactions identify regulatory hubs in dynamic compartmentalized domains [Capture-C]

Project description:The genome is organized into self-interacting chromatin domains containing genes and the cis-regulatory elements controlling their expression. How these domains form and how elements within them interact is not fully understood. We have developed Tri-C, a sensitive, high-resolution Chromosome Conformation Capture (3C) approach to identify concurrent chromatin interactions in single cells. Combining Tri-C with conventional 3C and polymer modeling, we show that, rather than folding into stable pre-formed loops, self-interacting domains form dynamic compartmentalized chromatin structures, delimited by CTCF/Cohesin boundaries. Within these tissue-specific domains, all regions of chromatin contact each other, but preferential structures are formed in which multiple enhancers and promoters interact simultaneously. Flanking CTCF/Cohesin-bound elements are excluded from these interactions and form distinct structures. These observations are best explained by a dynamic loop extrusion mechanism and subsequent stabilization of enhancer-promoter interactions, rather than the current view of long-range interactions occurring via the formation of discrete pre-formed chromatin loops.

2018-08-17 | GSE107753 | GEO

A Mechanism of Cohesin-Dependent Loop Extrusion Organizes Zygotic Genome Architecture

Project description:Fertilization triggers assembly of higher-order chromatin structure from a nucleosomal state to generate a totipotent embryo. Chromatin loops and domains are detected in mouse zygotes by single-nucleus Hi-C (snHi-C) but not bulk Hi-C; both are absent in Drosophila embryos. We investigated whether zygotic chromatin structures are generated by cohesin-dependent loop extrusion. Using snHi-C of mouse knockout embryos, we demonstrate the zygotic genome folds into loops and domains that critically depend on Scc1-cohesin and are regulated in size by Wapl. Remarkably, we discovered distinct effects on maternal and paternal chromatin loop sizes, likely reflecting loop extrusion dynamics and reprogramming states. Polymer simulations based on snHi-C are consistent with a model where cohesin locally compacts chromatin and restricts inter-chromosomal interactions by active loop extrusion, whose processivity is controlled by Wapl. Our simulations and experimental data provide evidence that cohesin-dependent loop extrusion organizes mammalian genomes over multiple scales from the one-cell embryo onwards.

2017-08-25 | GSE100569 | GEO

MCM complexes are barriers that restrict cohesin-mediated loop extrusion [snHi-C]

Project description:Eukaryotic genomes are compacted into loops and topologically associating domains (TADs), which contribute to transcription, recombination and genomic stability. Cohesin extrudes DNA into loops that are thought to lengthen until CTCF boundaries are encountered. Little is known about whether loop extrusion is impeded by DNA-bound macromolecular machines. We demonstrate that the replicative helicase MCM is a barrier that restricts loop extrusion in G1 phase. Single-nucleus Hi-C of one-cell embryos revealed that MCM loading reduces CTCF-anchored loops and decreases TAD boundary insulation, suggesting loop extrusion is impeded before reaching CTCF. Single-molecule imaging shows that MCMs are physical barriers that constrain cohesin translocation in vitro. Simulations are consistent with MCMs as abundant, random barriers. We conclude that distinct loop extrusion barriers contribute to shaping 3D genomes.

2022-03-08 | GSE196300 | GEO

MCM complexes are barriers that restrict loops and TADs

Project description:Eukaryotic genomes are compacted into loops and topologically associating domains (TADs), which contribute to transcription, recombination and genomic stability. Cohesin extrudes DNA into loops that are thought to lengthen until it encounters CTCF boundaries. Little is known whether loop extrusion is impeded by macromolecular machines. We demonstrate that the replicative helicase MCM is a barrier that restricts loops and TADs in G1 phase. Single-nucleus Hi-C of one-cell embryos revealed that MCM loading reduces CTCF-anchored loops and increases TAD boundary insulation, suggesting loop extrusion is impeded before reaching CTCF. Single-molecule imaging provides evidence that MCM are physical barriers that constrain cohesin translocation in vitro. Simulations are consistent with MCM as abundant, random barriers with low permeability. We conclude that distinct loop extrusion barriers contribute to shaping 3D genomes.

2022-03-07 | GSE155971 | GEO

Chromatin jets define the properties of cohesin-driven in vivo loop extrusion

Project description:Complex genomes show intricate organization in three-dimensional (3D) nuclear space. Current models posit that cohesin extrudes loops to form self-interacting domains delimited by the DNA binding protein CTCF. Here, we describe and quantitatively characterize cohesin-propelled, jet-like chromatin contacts as landmarks of loop extrusion in quiescent mammalian lymphocytes. Experimental observations and polymer simulations indicate that narrow origins of loop extrusion favor jet formation. Unless constrained by CTCF, jets propagate symmetrically for 1-2 Mb, providing an estimate for the range of in vivo loop extrusion. Asymmetric CTCF binding deflects the angle of jet propagation as experimental evidence that cohesin-mediated loop extrusion can switch from bi- to unidirectional and is controlled independently in both directions. These data offer new insights into the physiological behavior of in vivo cohesin-mediated loop extrusion and further our understanding of the principles that underlie genome organization.

2022-08-15 | GSE199059 | GEO

Cohesin and CTCF control chromosomal contact dynamics in living cells [integration sites]

Project description:Mammalian chromosomes are partitioned into topologically associating domains (TADs) by the loop-extrusion activity of cohesin that is blocked at specific DNA sites bound by CTCF. Chromosome structure inside TADs is highly variable in single cells, yet little is known about its temporal dynamics, how it influences the rates and durations of chromosomal contacts, and how it depends on CTCF and cohesin. To address these questions we combine two quantitative live-cell imaging strategies that minimize locus-specific confounding effects. We show that loop extrusion by cohesin globally reduces the mobility of the chromatin fiber in living cells, while also increasing the rates of formation and durations of contacts between sequences inside the same TAD. Quantitative analysis of high-resolution microscopy data reveals that contacts assemble and disassemble frequently in the course of the cell cycle, and become substantially more frequent and longer in the presence of convergent CTCF sites. Comparison with polymer modeling additionally reveals that cohesin-mediated CTCF loops last around 10 minutes on average. Our data support the notion that chromosome structure within TADs is highly dynamic and provide a quantitative framework for understanding the principles that link chromosome structure to biological function.

2022-02-27 | GSE197237 | GEO

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