Background

The genome is normally organized into A/B compartments, TADs, and chromatin loops, but the mechanisms that actually drive compartment formation and separation are still not fully understood. This is especially unclear because, in interphase, many factors act at the same time, including transcription, chromatin-binding proteins, nuclear lamina attachment, and cohesin-mediated loop extrusion. During mitosis, most of these structures disappear and condensin becomes dominant, making mitosis a useful system to reveal more fundamental folding forces. 

In mitosis, condensin is a key structural protein complex that organizes chromosomes to ensure their proper segregation. Its primary functions include chromosome condensation (core function), chromosome individualization and segregation, suppression of alternative chromatin folding, and disassembly of interphase chromosome architecture. Notably, condensin suppresses compartmentalization (A/B-like segregation) during mitosis. When condensin is removed, chromosomes unexpectedly regain compartment-like organization, even in the absence of transcription or cohesin.

Question

what genome-folding principles become visible when mitotic chromosomes are stripped of condensin-driven loop extrusion? More specifically, whether compartments and other chromatin contacts can re-emerge in mitosis without condensin, and what this reveals about the forces underlying chromatin compartmentalization.

Main findings

When both condensin I and condensin II were acutely removed, mitotic chromosomes progressively regained strong compartmentalization patterns similar to interphase, even though TADs and canonical structural loops did not reappear. This showed that condensin normally suppresses a latent compartmentalization program in mitosis.

The study further found that condensin loss uncovered two distinct active/euchromatic compartments, called mA1 and mA2, which are hard to distinguish in interphase but behave very differently in mitosis. It also showed that constitutive heterochromatin self-aggregates and can even cocompartmentalize with facultative heterochromatin, unlike in interphase.

Another important result was that some cis-regulatory element contacts remained detectable even without canonical CTCF/cohesin-based architecture, and that HP1 proteins were dispensable for constitutive heterochromatin compartment formation and re-establishment after mitosis. Overall, the paper argues that condensin-deficient mitotic chromosomes expose genome-folding forces that are masked in interphase cells.

Notes: Euchromatin is a lightly packed form of chromatin that is genetically active, meaning it contains genes that are frequently transcribed. 

Facultative heterochromatin is a type of heterochromatin that is reversibly condensed and can switch between a silent, tightly packed state (heterochromatin) and an active, loose state (euchromatin), depending on the cell type, developmental stage, or environmental signals.

Key methods used

The core strategy was acute protein depletion using an auxin-inducible degron system combined with CRISPR-based tagging of condensin subunits, especially SMC2, in mouse erythroblast cells. The authors synchronized cells in mitosis, depleted condensin over time, and then profiled chromosome architecture. Their main analytical method was in situ Hi-C, including time-course Hi-C after condensin depletion, followed by eigenvector decomposition, saddle plots, and compartment analysis to quantify changes in chromatin interactions. They also compared these results with cohesin depletion, incorporated histone-mark information such as H3K27ac, H3K27me3, and H3K9me3, and used HP1 triple depletion (HP1α, HP1β, and HP1γ) to test whether HP1 is required for compartment re-formation.

My question: How to distinct euchromatin and heterchromatin, constitutive heterochromatin and facultative heterchromatin?

Euchromatin is characterized by active histone modifications, including H3K27ac (enhancers), H3K4me3 (promoters), and H3K36me3 (gene bodies).Constitutive heterochromatin is marked by H3K9me3 and enrichment of HP1 proteins, reflecting its stable and transcriptionally silent nature.Facultative heterochromatin is defined by H3K27me3, indicative of reversible and context-dependent gene repression.

Research article link: https://doi.org/10.1038/s41588-024-01759-x