DNA-replication checkpoint activation promotes domain-specific re-licensing in S-phase and extra-initiations stabilizing CFS. Brison et al.
Genome integrity requires replication to be completed before chromosome segregation. This coordination essentially relies on stress-induced activation of the DNA-replication checkpoint that inhibits CDK1, delaying mitotic onset. Under-replication of Common Fragile Sites (CFSs) however escapes surveillance, which results in mitotic chromosome breaks. Here we asked whether this leakage results from insufficient CDK1 inhibition under modest stresses used to destabilize CFSs. We found that tight CDK1 inhibition completely rescues genome stability in so-stressed human cells. Unexpectedly, molecular combing and Repli-Seq analyses showed a burst of replication initiations in mid S-phase across not yet replicated origin-poor domains shaped by transcription, including large genes hosting CFSs. Extra-initiations and CFS rescue required availability of CDC6 and CDT1 during the S-phase, showing that CDK1 inhibition promotes mistimed re-licensing. In addition to delay mitotic onset, tight checkpoint activation therefore advances replication completion of origin-poor domains at risk of under-replication, two complementary roles preserving genome stability.