The Dbf4-Cdc7 kinase promotes S phase by alleviating an inhibitory activity in Mcm4.
Eukaryotic DNA replication uses kinase regulatory pathways to facilitate coordination with other processes during cell division cycles and response to environmental cues. At least two cell cycle-regulated protein kinase systems, the S-phase-specific cyclin-dependent protein kinases (S-CDKs) and the Dbf4-Cdc7 kinase (DDK, Dbf4-dependent protein kinase) are essential activators for initiation of DNA replication. Although the essential mechanism of CDK activation of DNA replication in Saccharomyces cerevisiae has been established, exactly how DDK acts has been unclear. Here we show that the amino terminal serine/threonine-rich domain (NSD) of Mcm4 has both inhibitory and facilitating roles in DNA replication control and that the sole essential function of DDK is to relieve an inhibitory activity residing within the NSD. By combining an mcm4 mutant lacking the inhibitory activity with mutations that bypass the requirement for CDKs for initiation of DNA replication, we show that DNA synthesis can occur in G1 phase when CDKs and DDK are limited. However, DDK is still required for efficient S phase progression. In the absence of DDK, CDK phosphorylation at the distal part of the Mcm4 NSD becomes crucial. Moreover, DDK-null cells fail to activate the intra-S-phase checkpoint in the presence of hydroxyurea-induced DNA damage and are unable to survive this challenge. Our studies establish that the eukaryote-specific NSD of Mcm4 has evolved to integrate several protein kinase regulatory signals for progression through S phase.
Sheu
Yi-Jun
YJ
Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, New York 11724, USA.
Stillman
Bruce
B
D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016193:G1 Phase / Q000187:drug effects; D020043:Genes, Essential; D006918:Hydroxyurea / Q000494:pharmacology; D050296:Microbial Viability / Q000187:drug effects; D064148:Minichromosome Maintenance Complex Component 4; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D016196:S Phase / Q000187:drug effects / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D013379:Substrate Specificity
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C088096:Dbf4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C489404:CDC7 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C092802:MCM4 protein, S cerevisiae; D064148:Minichromosome Maintenance Complex Component 4; D006918:Hydroxyurea