A Novel Cdk2 Interactor is Phosphorylated by Cdc7 and Associates with Components of the Replication Complexes

Abstract

Initiation of DNA replication in eukaryotic cells depends on the assembly of the pre-replicationcomplexes containing two hexamers, the Origin Recognition Complex (ORC) and theMinichromosome maintenance/DNA Replication Licensing complex (MCM), and on subsequentconformational changes in the MCM complex leading to the formation of a competent DNAreplication complex, firing of the DNA polymerase and disassembly of the MCM. The dynamicsof the MCM complex is under the control of two Ser/Thr kinases, the Cell cycle-dependentkinase 2 (Cdk2) and Cell division cycle gene 7 (Cdc7). The precise substrates of the kinases atthe origins and the sequence of events leading to the origins firing are not well understood.Using the two hybrid selection in yeast, we have identified a novel gene, the Cdk2 interactingprotein, CINP. We show that CINP is a component of the active cyclin E /Cdk2 and cyclin A/Cdk2 complexes. CINP also interacts with Cdc7 and is phopshorylated by Cdc7, but not byCdk2. We further show that CINP binds to chromatin in a replication-dependent manner, andassociates with ORC2-containing complexes and MCM. We propose that CINP is part of theCdc7-dependent mechanism of origin firing and a functional and physical link between Cdk2and Cdc7 complexes at the origins.