Concerted activities of Mcm4, Sld3 and Dbf4 in control of origin activation and DNA replication fork progression

Eukaryotic chromosomes initiate DNA synthesis from multiple replication origins in a temporally specific manner during S phase. The replicative helicase Mcm2-7 functions in both initiation and fork progression and thus is an important target of regulation. Mcm4, a helicase subunit, possesses an unstructured regulatory domain that mediates control from multiple kinase signaling pathways, including the Dbf4-dependent Cdc7 kinase (DDK). Following replication stress in S phase, Dbf4 and Sld3, an initiation factor and essential target of Cyclin-Dependent Kinase (CDK), are targets of the checkpoint kinase Rad53 for inhibition of initiation from origins that have yet to be activated, so-called late origins. Here, whole genome DNA replication profile analysis is employed to access under various conditions the effect of mutations that alter the Mcm4 helicase regulatory domain and the Rad53 targets, Sld3 and Dbf4. Late origin firing occurs under genotoxic stress when the controls on Mcm4, Sld3 and Dbf4 are simultaneously eliminated. The regulatory domain of Mcm4 plays an important role in the timing of late origin firing, both in an unperturbed S phase and dNTP limitation. Furthermore, checkpoint control of Sld3 impacts fork progression ...Continue Reading