embj.201593552.pdf (1017.46 kB)
Phosphopeptide binding by Sld3 links Dbf4-dependent kinase to MCM replicative helicase activation
journal contributionposted on 2020-07-17, 16:33 authored by Tom D Deegan, Joseph TP Yeeles, John FX Diffley
The initiation of eukaryotic DNA replication requires the assembly of active CMG (Cdc45-MCM-GINS) helicases at replication origins by a set of conserved and essential firing factors. This process is controlled during the cell cycle by cyclin-dependent kinase (CDK) and Dbf4-dependent kinase (DDK), and in response to DNA damage by the checkpoint kinase Rad53/Chk1. Here we show that Sld3, previously shown to be an essential CDK and Rad53 substrate, is recruited to the inactive MCM double hexamer in a DDK-dependent manner. Sld3 binds specifically to DDK-phosphorylated peptides from two MCM subunits (Mcm4, 6) and then recruits Cdc45. MCM mutants that cannot bind Sld3 or Sld3 mutants that cannot bind phospho-MCM or Cdc45 do not support replication. Moreover, phosphomimicking mutants in Mcm4 and Mcm6 bind Sld3 without DDK and facilitate DDK-independent replication. Thus, Sld3 is an essential "reader" of DDK phosphorylation, integrating signals from three distinct protein kinase pathways to coordinate DNA replication during S phase.
DDKDNA replication initiationSld3Cell Cycle ProteinsDNA ReplicationDNA-Binding ProteinsMinichromosome Maintenance Complex Component 4Minichromosome Maintenance Complex Component 6Nuclear ProteinsPhosphopeptidesProtein BindingProtein-Serine-Threonine KinasesReplication OriginSaccharomyces cerevisiaeSaccharomyces cerevisiae ProteinsDiffley FC001066PC-ackFERM-ackDevelopmental Biology06 Biological Sciences08 Information and Computing Sciences11 Medical and Health Sciences