Re-establishment of TAD boundary organization during DNA replication

Area of Science:

• Genomics
• Molecular Biology
• Cell Biology

Background:

• Topologically associating domains (TADs) are fundamental to higher-order genome organization.
• Cohesin and CTCF are key proteins in TAD formation and maintenance, but their duplication during replication is unclear.

Purpose of the Study:

• To identify chromatin factors involved in TAD establishment and maintenance during genome replication.
• To elucidate the mechanisms underlying TAD duplication and restoration post-replication.

Main Methods:

• High-throughput imaging-based CRISPR/Cas9 knock-out screening to identify relevant chromatin factors.
• Analysis of TAD structure dynamics during S-phase DNA replication.
• Investigating the role of the RPA complex in TAD re-folding and protein binding.

Main Results:

• TADs transiently unfold during S-phase DNA replication.
• Impeding replication progression hinders TAD structure restoration.
• RPA complex inhibition prevents cohesin/CTCF binding, delays TAD re-folding, and affects TADs in non-cycling cells.

Conclusions:

• DNA replication machinery components are crucial for TAD maintenance.
• The RPA complex is essential for proper cohesin and CTCF binding and TAD re-folding post-replication.
• Novel insights into how TAD structures are re-established during genome duplication are provided.