title: Robust Replication Control Is Generated by Temporal Gaps between Licensing and Firing Phases and Depends on Degradation of Firing Factor Sld2.
abstract: Temporal separation of DNA replication initiation into licensing and firing phases ensures the precise duplication of the genome during each cell cycle. Cyclin-dependent kinase (CDK) is known to generate this separation by activating firing factors and at the same time inhibiting licensing factors but may not be sufficient to ensure robust separation at transitions between both phases. Here, we show that a temporal gap separates the inactivation of firing factors from the re-activation of licensing factors during mitosis in budding yeast. We find that gap size critically depends on phosphorylation-dependent degradation of the firing factor Sld2 mediated by CDK, DDK, Mck1, and Cdc5 kinases and the ubiquitin-ligases Dma1/2. Stable mutants of Sld2 minimize the gap and cause increased genome instability in an origin-dependent manner when combined with deregulation of other replication regulators or checkpoint mechanisms. Robust separation of licensing and firing phases therefore appears indispensable to safeguard genome stability.
authors.affiliation: Max Planck Institute of Biochemistry, DNA Replication and Genome Integrity, 82152 Martinsried, Germany.
authors.forename: Karl-Uwe
authors.initials: KU
authors.lastname: Reusswig
authors.affiliation: Max Planck Institute of Biochemistry, DNA Replication and Genome Integrity, 82152 Martinsried, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Zimmermann
authors.affiliation: Max Planck Institute of Biochemistry, DNA Replication and Genome Integrity, 82152 Martinsried, Germany.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Galanti
authors.affiliation: Max Planck Institute of Biochemistry, DNA Replication and Genome Integrity, 82152 Martinsried, Germany. Electronic address: bpfander@biochem.mpg.de.
authors.forename: Boris
authors.initials: B
authors.lastname: Pfander
mesh_terms: D018797:Cell Cycle Proteins / Q000235:genetics* / Q000378:metabolism; D004261:DNA Replication / Q000235:genetics*; D016681:Genome, Fungal; D042822:Genomic Instability / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000235:genetics; D059748:Proteolysis; D018741:Replication Origin / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; C493793:DMA2 protein, S cerevisiae; C493792:Dma1 protein, S cerevisiae; C115043:SLD2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D046988:Proteasome Endopeptidase Complex
keywords: DNA replication; cell cycle; genome stability; post-translational modification