XRCC3 and Rad51 modulate replication fork progression on damaged vertebrate chromosomes

Area of Science:

• Molecular Biology
• Genetics
• DNA Repair

Background:

• The precise mechanisms controlling eukaryotic replication fork progression following DNA damage remain largely unknown.
• Understanding these mechanisms is crucial for comprehending genome stability and cellular response to genotoxic stress.

Purpose of the Study:

• To elucidate the role of homologous recombination proteins in regulating replication fork progression after DNA damage.
• To identify specific proteins involved in the active process of replication fork slowing.

Main Methods:

• Utilized cisplatin and UV treatment in intact vertebrate cells and in vitro replication assays.
• Examined replication fork progression in irs1SF CHO cells and XRCC3(-/-) chicken DT40 cells.
• Assessed the effect of purified human Rad51C-XRCC3 complex and human Rad51 addition on fork slowing.

Main Results:

• Replication fork progression was significantly slowed by cisplatin or UV treatment in wild-type cells.
• Fork slowing was reduced or absent in irs1SF CHO and XRCC3(-/-) chicken DT40 cells, indicating an active, XRCC3-dependent process.
• The addition of Rad51C-XRCC3 complex restored fork slowing in XRCC3(-/-) cells, and human Rad51 could bypass the XRCC3 requirement.

Conclusions:

• Homologous recombination protein XRCC3 is essential for the active process of replication fork slowing after DNA damage.
• XRCC3 and Rad51 function cooperatively to modulate replication fork progression on damaged vertebrate chromosomes.
• These findings reveal a critical role for recombination proteins in maintaining genome integrity during DNA replication stress.