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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 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.

SUBMITTER: Sanborn AL

PROVIDER: S-EPMC4664323 | biostudies-literature | 2015 Nov

REPOSITORIES: biostudies-literature

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

Sanborn Adrian L AL Rao Suhas S P SS Huang Su-Chen SC Durand Neva C NC Huntley Miriam H MH Jewett Andrew I AI Bochkov Ivan D ID Chinnappan Dharmaraj D Cutkosky Ashok A Li Jian J Geeting Kristopher P KP Gnirke Andreas A Melnikov Alexandre A McKenna Doug D Stamenova Elena K EK Lander Eric S ES Aiden Erez Lieberman EL

Proceedings of the National Academy of Sciences of the United States of America 20151023 47

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. ...[more]

PMID: 26499245

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Project description:Antibody class switch recombination (CSR) in B lymphocytes replaces immunoglobulin heavy chain locus (Igh) C? constant region exons (CHs) with one of six CHs lying 100-200 kb downstream1. Each CH is flanked upstream by an I promoter and long repetitive switch (S) region1. Cytokines and activators induce activation-induced cytidine deaminase (AID)2 and I-promoter transcription, with 3' IgH regulatory region (3' IgHRR) enhancers controlling the latter via I-promoter competition for long-range 3' IgHRR interactions3-8. Transcription through donor S? and an activated downstream acceptor S-region targets AID-generated deamination lesions at, potentially, any of hundreds of individual S-region deamination motifs9-11. General DNA repair pathways convert these lesions to double-stranded breaks (DSBs) and join an S?-upstream DSB-end to an acceptor S-region-downstream DSB-end for deletional CSR12. AID-initiated DSBs at targets spread across activated S regions routinely participate in such deletional CSR joining11. Here we report that chromatin loop extrusion underlies the mechanism11 by which IgH organization in cis promotes deletional CSR. In naive B cells, loop extrusion dynamically juxtaposes 3' IgHRR enhancers with the 200-kb upstream S? to generate a CSR centre (CSRC). In CSR-activated primary B cells, I-promoter transcription activates cohesin loading, leading to generation of dynamic subdomains that directionally align a downstream S region with S? for deletional CSR. During constitutive S? CSR in CH12F3 B lymphoma cells, inversional CSR can be activated by insertion of a CTCF-binding element (CBE)-based impediment in the extrusion path. CBE insertion also inactivates upstream S-region CSR and converts adjacent downstream sequences into an ectopic S region by inhibiting and promoting their dynamic alignment with S? in the CSRC, respectively. Our findings suggest that, in a CSRC, dynamically impeded cohesin-mediated loop extrusion juxtaposes proper ends of AID-initiated donor and acceptor S-region DSBs for deletional CSR. Such a mechanism might also contribute to pathogenic DSB joining genome-wide.

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Dataset Information

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

Publications

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

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