Abstract
Replication stress is a key contributor to genomic instability and cancers. BRCA1 has well established roles in protecting stalled forks against degradation. Here we show that BRCA1 has a fork protection-independent role in driving stalled forks restoration via regulation of RAD51 in a manner that confers dependency on SCAI and REV3. BRCA1 drives SLX4-mediated DNA break formation in the absence of SCAI and REV3, which are required to complete repair synthesis and fork restart. Domain analysis revealed that the increased fork breakage seen in the absence of SCAI requires the coiled coil domain of BRCA1 and is regulated in a manner distinct from BRCA1's roles in resection control at two-ended double strand breaks. BRCA1's role also occurs independently of 53BP1 but is modulated by RIF1. Surprisingly, loss of fork reversal factors leads to increased fork breakage in the absence of SCAI/REV3 in a manner that depends on RAD51 activity. SCAI loss leads to persistent DNA breaks, genomic instability and cell death upon DNA damage. We posit that SCAI/REV3 may be required to replicate through and rescue stalled replication forks at fragile genomic regions. Failure to do so leads to increased DNA breakage and genomic instability.