The intra-S phase checkpoint directly regulates replication elongation to preserve the integrity of stalled replisomes.

Abstract

DNA replication is dramatically slowed down under replication stress. The regulation of replication speed is a conserved response in eukaryotes and, in fission yeast, requires the checkpoint kinases Rad3^ATR and Cds1^Chk2 However, the underlying mechanism of this checkpoint regulation remains unresolved. Here, we report that the Rad3^ATR-Cds1^Chk2 checkpoint directly targets the Cdc45-MCM-GINS (CMG) replicative helicase under replication stress. When replication forks stall, the Cds1^Chk2 kinase directly phosphorylates Cdc45 on the S275, S322, and S397 residues, which significantly reduces CMG helicase activity. Furthermore, in <i>cds1</i>^<i>Chk2</i> -mutated cells, the CMG helicase and DNA polymerases are physically separated, potentially disrupting replisomes and collapsing replication forks. This study demonstrates that the intra-S phase checkpoint directly regulates replication elongation, reduces CMG helicase processivity, prevents CMG helicase delinking from DNA polymerases, and therefore helps preserve the integrity of stalled replisomes and replication forks.