Loop stacking organizes genome folding from TADs to chromosomes

Area of Science:

• Genomics
• Cell Biology
• Molecular Biology

Background:

• Population-level studies highlight CTCF and cohesin's role in mammalian genome organization.
• Single-cell level contributions of these proteins remain poorly understood.

Purpose of the Study:

• To investigate the single-cell level effects of CTCF and cohesin removal on genome organization using super-resolution microscopy.
• To elucidate the structural contributions of CTCF and cohesin to chromatin organization at multiple scales.

Main Methods:

• Super-resolution microscopy was employed to analyze mouse embryonic stem cells.
• The impact of CTCF or cohesin depletion on single-chromosome structures was measured.

Main Results:

• Cohesin forms stacked loops at anchors, creating multi-way contact hubs that bridge TAD boundaries without intermixing intervening chromatin.
• Loop stacking provides steric insulation, separating local chromatin from ultra-long-range contacts (>4 Mb).
• Cohesin removal led to increased chromosome disorder and cell-to-cell gene expression variability.

Conclusions:

• CTCF and cohesin's genome organization roles are revised beyond a TAD-centric view.
• A multi-scale structural model explains their genome organization function at the single-cell level.
• Distinct contributions to loop stacking by CTCF and cohesin are identified.