The Replication Checkpoint Prevents Two Types of Fork Collapse without Regulating Replisome Stability

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• The ATR replication checkpoint is crucial for maintaining genome stability by regulating stalled replication forks.
• Understanding replisome dynamics during replication stress is essential for comprehending DNA repair mechanisms.

Purpose of the Study:

• To quantitatively characterize human replisome dynamics and the replication stress response proteome.
• To elucidate the role of the ATR checkpoint in preventing replication fork collapse.
• To identify novel proteins associated with replication forks and their functions.

Main Methods:

• Utilized an improved iPOND protocol combined with SILAC mass spectrometry.
• Analyzed protein abundance on nascent DNA across 32 experimental conditions.
• Employed unsupervised hierarchical clustering for protein complex identification and pathway analysis.

Main Results:

• The ATR checkpoint was found to prevent, rather than stabilize, two distinct types of replication fork collapse.
• Quantitative proteomic data revealed the dynamics of the replisome and replication stress response.
• Identified ZNF644 as a novel protein complexing with G9a/GLP methyltransferase at replication forks, crucial for preventing DNA damage.

Conclusions:

• The ATR replication checkpoint actively prevents replication fork collapse to preserve genome integrity.
• The study provides insights into the mechanism of action for ATR inhibitors.
• The findings offer a valuable resource for researchers in replication, DNA repair, and chromatin biology.