Phosphorylation inhibits intramolecular interactions, DNA-binding and protein interactions of Claspin through disordered/ structured conformation transition

Claspin, known to be highly disordered, plays important roles in replication fork progression, initiation and cellular responses to replication stress. However, regulation of its structure and molecular interactions is not completely understood. We show here, through Proximity-Ligation-Assays, the evidence for intramolecular interaction between the N- and C-terminal segments of Claspin, which depends on the Acidic-Patch [AP] segment near its C-terminus. Interaction of Claspin with DNA and replication factors is highly stimulated in ∆AP mutant and by prior dephosphorylation. The wild-type Claspin inhibits the helicase activity of MCM in an AP-dependent manner. ∆AP and dephosphorylated Claspin exhibit resistance to trypsin digestion compared to wild-type, suggesting the presence of structural domains in the formers. We propose that Claspin is converted from disordered (closed) to structured (open) conformation at initiation, which stimulates its DNA binding and interaction with replication factors and counteracts its helicase inhibitory activity to trigger initiation of DNA replication.