Sequential replication-coupled destruction at G1/S ensures genome stability.

Area of Science:

• Cell Biology
• Molecular Biology
• Genetics

Background:

• Timely ubiquitin-mediated protein degradation is essential for cell cycle regulation.
• The precise degradation order of cell cycle proteins at phase transitions remains incompletely understood.
• CRL4(Cdt2) is a key E3 ubiquitin ligase mediating S-phase degradation of proteins like Cdt1, PR-Set7, and p21.

Purpose of the Study:

• To investigate the degradation order of CRL4(Cdt2) substrates during S-phase.
• To elucidate the role of the PIP degron in regulating substrate degradation timing.
• To understand how sequential degradation impacts S-phase progression and genome stability.

Main Methods:

• Analysis of synchronized and live single cells.
• Investigating the degradation kinetics of Cdt1 and p21.
• Utilizing genetic manipulation by fusing PIP degrons between substrates.

Main Results:

• A consistent degradation order was observed: Cdt1 is degraded before p21 during S-phase entry and DNA repair.
• The PIP degron's interaction with DNA-loaded proliferating cell nuclear antigen (PCNA(DNA)) dictates degradation timing.
• Fusing Cdt1's PIP degron to p21 accelerated p21 degradation, leading to Cdt2 recruitment and concurrent degradation with Cdt1.
• Accelerated p21 degradation resulted in stalled replication and sensitivity to replication arrest.

Conclusions:

• Sequential degradation of CRL4(Cdt2) substrates ensures orderly S-phase progression.
• This ordered degradation prevents replication stress and maintains genome stability.
• The PIP degron mechanism fine-tunes substrate degradation timing for proper cell cycle control.