Genome-wide in vivo dynamics of cohesin-mediated loop extrusion and its role in transcription activation [SLAM-seq]
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1164730 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30786100_1.fastq.gz | Fastqsanger.gz | |||
| SRR30786100_2.fastq.gz | Fastqsanger.gz | |||
| SRR30786101_1.fastq.gz | Fastqsanger.gz | |||
| SRR30786101_2.fastq.gz | Fastqsanger.gz | |||
| SRR30786102_1.fastq.gz | Fastqsanger.gz |
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Project description:Most animal genomes are partitioned into Topologically Associating Domains (TADs), created by cohesin-mediated loop extrusion and defined by convergently oriented CTCF sites. The dynamics of loop extrusion and its regulation remains poorly characterized in vivo. Here, we tracked TAD anchors in living human cells to visualize and quantify cohesin-dependent loop extrusion across multiple endogenous genomic regions. We show that TADs are dynamic structures whose anchors are brought in proximity about once per hour and for 6-19 min (~16% of the time). TADs are continuously subjected to extrusion by multiple cohesin complexes, extruding loops at ~0.1 kb/s. Remarkably, despite strong differences of Hi-C patterns between the chromatin regions, their dynamics is consistent with the same density, residence time and speed of cohesin. Our results suggest that TAD dynamics is governed primarily by CTCF site location and affinity, which allows genome-wide predictive models of cohesin-dependent interactions.
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Project description:Universal dynamics of cohesin-mediated loop extrusion
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Project description:Cohesin extrudes genomic DNA into loops that promote chromatin assembly, gene regulation and recombination. Loop extrusion depends on large-scale conformational changes in cohesin, but how these translocate DNA is poorly understood. Here we provide evidence that cohesin negatively supercoils DNA loops during extrusion. Supercoiling requires engagement of cohesin’s ATPase heads, DNA clamping by these heads, and a DNA binding site on cohesin’s hinge, indicating that cohesin twists DNA when constraining it between the hinge and the clamp. A cohesin mutant defective in negative supercoiling forms shorter loops in cells, and a similar, although weaker, phenotype is observed after depletion of topoisomerase I. These results suggest that supercoiling is an integral part of the loop extrusion mechanism and that relaxation of supercoiled DNA is required for cohesin-mediated loop extrusion and genome architecture.
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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.
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Project description:Cohesin supercoils DNA during loop extrusion
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Project description:The organization of the genome in three-dimensional space is highly dynamic, yet how these dynamics are regulated and the role they play in genome function is poorly understood. Here, we utilized acute depletion of NIPBL to characterize the role of cohesin-mediated loop extrusion in vivo. Using this approach, we find that many chromatin loops are rapidly diminished upon loss of NIBPL, consistent with recent single locus imaging studies showing that chromatin loops are transient. However, we also identified a subset of cohesin-dependent chromatin loops that are anchored in repressive chromatin and may be “long-lived”, given that they require NIPBL for their establishment upon mitotic exit, but are persistent when NIPBL is depleted from interphase cells. In addition to the reformation of 3D genome structures, mitotic exit coincides with widespread transcriptional activation. We found that NIPBL is essential for establishing the expression of lineage-defining genes during the M-G1 transition but has diminished impact in the steady-state maintenance of their expression. At genes sensitive to its depletion, NIPBL supports a unique local genome organization defined by greater spatial proximity to nearby enhancers and weaker transcription start site insulation of genomic contacts. Overall, we show that NIPBL-mediated loop extrusion is critical to genome organization and transcriptional regulation in vivo.
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Project description:The cohesin complex shapes chromosomes by DNA loop extrusion, but individual extrusion trajectories were so far unappreciable in vivo. Here, we developed and validated TArgeted Cohesin Loader (TACL), a system enabling the strong activation of anchored loop extrusion from dozens of defined genomic sites in living cells. Studying their individual loop extrusion trajectories revealed that extruding cohesin-STAG2 stops not only at domain boundaries but at all flanking CTCF sites, engaging them in a complex transient looping network that supports intradomain contacts. Cohesin-STAG1 cannot associate with weak CTCF binding sites and fails to similarly support intradomain interactions. NIPBL-MAU2 remains associated with cohesin when stalled at looping CTCF sites, suggesting these factors may also be required for loop stabilization. TACL induces cohesin traffic jams and illegal loops with divergent CTCF sites, demonstrating that stalled cohesin can block extruding cohesin in vivo. Genes exposed to TACL-induced loop extrusion were collectively hindered in transcription and the underlying chromatin altered its accessibility and reduced its H3K27ac marks. Thus, by enabling the study of individual loop extrusion trajectories in vivo, we could assign new functions to players, identify new looping networks and uncover an interplay between loop extrusion, gene transcription and chromatin composition.
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Project description:The cohesin complex shapes chromosomes by DNA loop extrusion, but individual extrusion trajectories were so far unappreciable in vivo. Here, we developed and validated TArgeted Cohesin Loader (TACL), a system enabling the strong activation of anchored loop extrusion from dozens of defined genomic sites in living cells. Studying their individual loop extrusion trajectories revealed that extruding cohesin-STAG2 stops not only at domain boundaries but at all flanking CTCF sites, engaging them in a complex transient looping network that supports intradomain contacts. Cohesin-STAG1 cannot associate with weak CTCF binding sites and fails to similarly support intradomain interactions. NIPBL-MAU2 remains associated with cohesin when stalled at looping CTCF sites, suggesting these factors may also be required for loop stabilization. TACL induces cohesin traffic jams and illegal loops with divergent CTCF sites, demonstrating that stalled cohesin can block extruding cohesin in vivo. Genes exposed to TACL-induced loop extrusion were collectively hindered in transcription and the underlying chromatin altered its accessibility and reduced its H3K27ac marks. Thus, by enabling the study of individual loop extrusion trajectories in vivo, we could assign new functions to players, identify new looping networks and uncover an interplay between loop extrusion, gene transcription and chromatin composition.
2025-06-26 | GSE278982 | GEO
Project description:The cohesin complex shapes chromosomes by DNA loop extrusion, but individual extrusion trajectories were so far unappreciable in vivo. Here, we developed and validated TArgeted Cohesin Loader (TACL), a system enabling the strong activation of anchored loop extrusion from dozens of defined genomic sites in living cells. Studying their individual loop extrusion trajectories revealed that extruding cohesin-STAG2 stops not only at domain boundaries but at all flanking CTCF sites, engaging them in a complex transient looping network that supports intradomain contacts. Cohesin-STAG1 cannot associate with weak CTCF binding sites and fails to similarly support intradomain interactions. NIPBL-MAU2 remains associated with cohesin when stalled at looping CTCF sites, suggesting these factors may also be required for loop stabilization. TACL induces cohesin traffic jams and illegal loops with divergent CTCF sites, demonstrating that stalled cohesin can block extruding cohesin in vivo. Genes exposed to TACL-induced loop extrusion were collectively hindered in transcription and the underlying chromatin altered its accessibility and reduced its H3K27ac marks. Thus, by enabling the study of individual loop extrusion trajectories in vivo, we could assign new functions to players, identify new looping networks and uncover an interplay between loop extrusion, gene transcription and chromatin composition.
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Project description:The cohesin complex shapes chromosomes by DNA loop extrusion, but individual extrusion trajectories were so far unappreciable in vivo. Here, we developed and validated TArgeted Cohesin Loader (TACL), a system enabling the strong activation of anchored loop extrusion from dozens of defined genomic sites in living cells. Studying their individual loop extrusion trajectories revealed that extruding cohesin-STAG2 stops not only at domain boundaries but at all flanking CTCF sites, engaging them in a complex transient looping network that supports intradomain contacts. Cohesin-STAG1 cannot associate with weak CTCF binding sites and fails to similarly support intradomain interactions. NIPBL-MAU2 remains associated with cohesin when stalled at looping CTCF sites, suggesting these factors may also be required for loop stabilization. TACL induces cohesin traffic jams and illegal loops with divergent CTCF sites, demonstrating that stalled cohesin can block extruding cohesin in vivo. Genes exposed to TACL-induced loop extrusion were collectively hindered in transcription and the underlying chromatin altered its accessibility and reduced its H3K27ac marks. Thus, by enabling the study of individual loop extrusion trajectories in vivo, we could assign new functions to players, identify new looping networks and uncover an interplay between loop extrusion, gene transcription and chromatin composition.
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