Helicase Subunit Cdc45 Targets the Checkpoint Kinase Rad53 to Both Replication Initiation and Elongation Complexes after Fork Stalling

Abstract

Across eukaryotes, disruption of DNA replication causes an S phase checkpoint response, which regulates multiple processes, including inhibition of replication initiation and fork stabilization. How these events are coordinated remains poorly understood. Here, we show that the replicative helicase component Cdc45 targets the checkpoint kinase Rad53 to distinct replication complexes in the budding yeast Saccharomyces cerevisiae. Rad53 binds to forkhead-associated (FHA) interaction motifs in an unstructured loop region of Cdc45, which is phosphorylated by Rad53 itself, and this interaction is necessary for the inhibition of origin firing through Sld3. Cdc45 also recruits Rad53 to stalled replication forks, which we demonstrate is important for the response to replication stress. Finally, we show that a Cdc45 mutation found in patients with Meier-Gorlin syndrome disrupts the functional interaction with Rad53 in yeast. Together, we present a single mechanism by which a checkpoint kinase targets replication initiation and elongation complexes, which may be relevant to human disease.