Background

Higher-order chromatin structures (A/B compartments, TADs, and loops) are largely disrupted during mitosis and must be re-established after cell division. However, the temporal order, mechanisms, and dependencies of this reorganization remain unclear, especially how different architectural features (compartments vs loops vs domains) are rebuilt and coordinated with transcription.

Question

How is genome architecture re-established after mitosis, and what are the kinetics and mechanisms governing the formation of compartments, domains, and loops?

Main findings

The main finding of this study is that chromatin architecture is re-established after mitosis through a stepwise and mechanistically distinct process. A/B compartments reappear very rapidly in ana/telophase and then progressively strengthen and expand as cells enter G1. By contrast, contact domains are rebuilt hierarchically in a bottom-up manner, with small subTADs forming first and later converging into larger TAD structures. The study further shows that CTCF rapidly reassociates with chromatin after mitosis, whereas cohesin reloads more slowly, and this delayed cohesin accumulation determines the timing of structural loop formation. Consistent with this, CTCF/cohesin-anchored structural loops emerge gradually and display features of loop extrusion, whereas enhancer–promoter contacts can form more rapidly, often independently of canonical CTCF/cohesin-mediated looping. The authors also identify a subset of transient regulatory contacts that appear early after mitosis and are later dissolved as stable chromatin boundaries and nearby structural loops become established. Overall, these results show that post-mitotic genome reorganization is driven by multiple distinct but interacting forces, rather than by a single unified mechanism.

Key methods used

The authors used a highly synchronized mouse erythroid cell system to follow chromatin reorganization during the mitosis-to-G1 transition with precise temporal resolution. They first arrested cells in prometaphase and then released them, using fluorescence-activated cell sorting to purify distinct post-mitotic stages with high purity. To map 3D genome architecture over time, they performed time-course in situ Hi-C, which allowed them to track the reappearance of A/B compartments, contact domains, and chromatin loops. They complemented this with ChIP-seq for key architectural and transcriptional factors, including CTCF, the cohesin subunit Rad21, and RNA polymerase II, to relate structural changes to factor binding and transcriptional reactivation. Capture-C was used to validate selected chromatin interactions at higher resolution, and live-cell imaging was performed to monitor the dynamic reassociation of CTCF and cohesin with chromosomes after mitosis. These datasets were then analyzed computationally using eigenvector decomposition for compartments, domain-calling algorithms for TAD and subTAD detection, and loop-calling and clustering approaches to define the kinetics of structural and regulatory contacts.

Summary

Genome architecture is rebuilt after mitosis through a hierarchical and kinetically distinct process, where compartments form first, domains emerge bottom-up, and cohesin-dependent loop extrusion gradually establishes structural loops.

My question: What are the key drivers of the hierarchical steps in the genome architecture rebuilding process?

The hierarchical rebuilding of genome architecture after mitosis is driven by sequential, kinetically distinct contributions: chromatin state-dependent compartmentalization, rapid cis-regulatory interactions, and slower cohesin-mediated loop extrusion. Among these, cohesin loading serves as the rate-limiting step for domain and loop formation. However, the specific key drivers and regulators remain largely unknown.

Research article link: https://doi.org/10.1038/s41586-019-1778-y