The conserved C terminus of Claspin interacts with Rad9 and promotes rapid activation of Chk1.

Claspin is a key mediator of the ATR-Chk1 checkpoint pathway. In response to DNA damage, Claspin interacts with Rad17 and Chk1 in a phosphorylation-dependent manner, enabling ATR to phosphorylate Chk1 efficiently. Claspin also interacts with Rad9, but how they interact and whether the interaction is functional remains unknown. Unexpectedly, our analysis of two splicing isoforms of Claspin provided an answer to these questions. The Claspin1339 isoform contains an evolutionarily conserved C terminus, but the Claspin1332 isoform does not. Although the transcripts encoding both Claspin isoforms coexist in HCT116 cells, Claspin1339 is the predominant form responsible for Chk1 activation. When expressed in cells depleted of endogenous Claspin, both Claspin1339 and Claspin1332 are able to mediate Chk1 activation. However, the activation of Chk1 is delayed in Claspin1332-expressing cells compared with Claspin1339-expressing cells. Furthermore, only Claspin1339 but not Claspin1332 interacts with Rad9 efficiently. Together, these results suggest that the conserved C terminus of Claspin interacts with Rad9 and ensures timely activation of the ATR-Chk1 pathway.