Unscheduled origin building in S-phase upon tight CDK1 inhibition suppresses CFS instability

Abstract

Genome integrity requires replication to be completed before chromosome segregation. This coordination essentially relies on replication-dependent activation of a dedicated checkpoint that inhibits CDK1, delaying mitotic onset. Under-replication of Common Fragile Sites (CFSs) however escapes surveillance, which triggers chromosome breakage. Using human cells, we asked here whether such leakage results from insufficient CDK1 inhibition under modest stresses used to destabilize CFSs. We found that tight CDK1 inhibition suppresses CFS instability. Repli-Seq and molecular combing analyses consistently showed a burst of replication initiations in mid S phase across large origin-poor domains shaped by transcription, including CFSs. Strikingly, CDC6 or CDT1 depletion or CDC7-DBF4 inhibition during the S phase prevented both extra-initiations and CFS rescue, showing that CDK1 inhibition promotes targeted and mistimed building of functional extra-origins. In addition to delay mitotic onset, checkpoint activation therefore advances replication completion of chromosome domains at risk of under-replication, two complementary roles preserving genome stability.