A double-ring of human RAD52 remodels replication forks restricting fork reversal

Area of Science:

• Molecular Biology
• DNA Repair Mechanisms
• Structural Biology

Background:

• Human RAD52 is a key DNA repair protein essential for maintaining genome stability.
• It plays a critical role in protecting stalled DNA replication forks from degradation during replication stress.
• The precise structural and molecular mechanisms underlying RAD52's fork protection function were previously unclear.

Purpose of the Study:

• To elucidate the structural and molecular mechanism of RAD52-mediated replication fork protection.
• To investigate how RAD52 remodels stalled replication forks.

Main Methods:

• P1 nuclease sensitivity assays
• Biochemical analyses
• Single-molecule analyses
• Mass photometry
• Single-particle cryo-electron microscopy (cryo-EM)

Main Results:

• RAD52 dynamically remodels replication forks through its strand exchange activity.
• The single-stranded DNA binding protein RPA modulates the kinetics of RAD52 strand exchange without altering the outcome.
• Cryo-EM revealed a unique nucleoprotein structure where two undecameric RAD52 rings arrange head-to-head, forming a positively charged surface that binds all three arms of the replication fork.

Conclusions:

• The head-to-head dimeric ring structure of RAD52 is critical for its strand exchange activity at replication forks.
• This structure facilitates competition with SMARCAL1, further contributing to fork protection and genome stability.
• The findings provide new insights into the molecular basis of DNA replication fork maintenance.