title: Rum1 and Cdc18 link inhibition of cyclin-dependent kinase to the initiation of DNA replication in Schizosaccharomyces pombe.
abstract: Eukaryotic cells have evolved regulatory mechanisms to ensure the strict alternation of DNA replication and mitosis. Recent work has suggested that the mitotic form of cyclin-dependent kinase (Cdc2/cyclin B) has a role in preventing re-replication of the genome before mitosis, but the relevant targets of this inhibition are unknown. In this report we present evidence that the mitotic cyclin-dependent kinase affects DNA replication by inhibiting the accumulation and function of Cdc18, a critical regulator of S-phase entry. We found that the ruml+ gene efficiently suppresses the lethality of a conditional cdc18 mutant. Conversely, deletion of ruml+ increases the severity of the cdc18 mutant phenotype, resulting in inappropriate cell division and a rapid loss of viability. Biochemical experiments indicate that Ruml potently inhibits Cdc2 phosphorylation of histone H1 or a Cdc18 fusion protein by directly interacting with the Cdc2/cyclin B complex. Overexpression of Ruml under conditions that promote re-replication of the genome induces a striking accumulation of Cdc18 protein by a largely post-transcriptional mechanism. Overexpression of SIC1, an unrelated cyclin-dependent kinase inhibitor from budding yeast, causes a similar accumulation of Cdc18 and also leads to re-replication. Our data link a potent inhibitor of Cdc2 kinase to a key protein required for the initiation of DNA replication and strongly suggest that inhibition of Cdc18 by cyclin-dependent kinases has an important role in ensuring that the genome is duplicated precisely once each cell cycle.
authors.affiliation: Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: P V
authors.initials: PV
authors.lastname: Jallepalli
authors.forename: T J
authors.initials: TJ
authors.lastname: Kelly
mesh_terms: D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors*; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002455:Cell Division / Q000235:genetics; D016213:Cyclins / Q000235:genetics / Q000378:metabolism; D004261:DNA Replication; D004791:Enzyme Inhibitors; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D015870:Gene Expression; D005800:Genes, Fungal; D005982:Glutathione Transferase / Q000235:genetics; D006657:Histones / Q000235:genetics / Q000378:metabolism; D008938:Mitosis / Q000235:genetics; D008957:Models, Genetic; D011401:Promoter Regions, Genetic; D012323:RNA Processing, Post-Transcriptional; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012568:Schizosaccharomyces / Q000235:genetics*; D029702:Schizosaccharomyces pombe Proteins; D013489:Suppression, Genetic; D013831:Thiamine / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D016213:Cyclins; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D006657:Histones; C093953:NMT1 protein, S pombe; D011993:Recombinant Fusion Proteins; C086077:Rum1 protein, S pombe; D029702:Schizosaccharomyces pombe Proteins; C082462:cdc18 protein, S pombe; D005982:Glutathione Transferase; D016203:CDC2 Protein Kinase; D013831:Thiamine