title: Genetics of lagging strand DNA synthesis and maturation in fission yeast: suppression analysis links the Dna2-Cdc24 complex to DNA polymerase delta.
abstract: The Cdc24 protein is essential for the completion of chromosomal DNA replication in fission yeast. Although its precise role in this process is unclear, Cdc24 forms a complex with Dna2, a conserved endonuclease-helicase implicated in the removal of the RNA-DNA primer during Okazaki fragment processing. To gain further insights into Cdc24-Dna2 function, we screened for chromosomal suppressors of the temperature-sensitive cdc24-M38 allele and mapped the suppressing mutations into six complementation groups. Two of these mutations defined genes encoding the Pol3 and Cdc27 subunits of DNA polymerase delta. Sequence analysis revealed that all the suppressing mutations in Cdc27 resulted in truncation of the protein and loss of sequences that included the conserved C-terminal PCNA binding motif, previously shown to play an important role in maximizing enzyme processivity in vitro. Deletion of this motif is shown to be sufficient for suppression of both cdc24-M38 and dna2-C2, a temperature-sensitive allele of dna2(+), suggesting that disruption of the interaction between Cdc27 and PCNA renders the activity of the Cdc24-Dna2 complex dispensable.
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, Michael Swann Building, King's Buildings, Mayfield Road, Edinburgh EH9 3JR, UK.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Tanaka
authors.forename: Gi-Hyuck
authors.initials: GH
authors.lastname: Ryu
authors.forename: Yeon-Soo
authors.initials: YS
authors.lastname: Seo
authors.forename: Stuart A
authors.initials: SA
authors.lastname: MacNeill
mesh_terms: D000483:Alleles; D001483:Base Sequence; D018797:Cell Cycle Proteins / Q000235:genetics* / Q000502:physiology*; D004258:DNA Polymerase III / Q000235:genetics* / Q000378:metabolism; D004261:DNA Replication; D004271:DNA, Fungal / Q000096:biosynthesis; D045585:Flap Endonucleases / Q000235:genetics* / Q000378:metabolism; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D012568:Schizosaccharomyces / Q000235:genetics* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D013489:Suppression, Genetic
chemical_list: C468288:Cdc24 protein, S pombe; D018797:Cell Cycle Proteins; D004271:DNA, Fungal; D018809:Proliferating Cell Nuclear Antigen; D029702:Schizosaccharomyces pombe Proteins; C513024:cdc27 protein, S pombe; D004258:DNA Polymerase III; C493531:Dna2 protein, S pombe; D045585:Flap Endonucleases