Dpb2 integrates the leading-strand DNA polymerase into the eukaryotic replisome

Curr Biol. 2013 Apr 8;23(7):543-52. doi: 10.1016/j.cub.2013.02.011. Epub 2013 Mar 14.


Background: The eukaryotic replisome is a critical determinant of genome integrity with a complex structure that remains poorly characterized. A central unresolved issue is how the Cdc45-MCM-GINS helicase is linked to DNA polymerase epsilon, which synthesizes the leading strand at replication forks and is an important focus of regulation.

Results: Here, we use budding yeast to show that a conserved amino-terminal domain of the Dpb2 subunit of Pol ε (Dpb2NT) interacts with the Psf1 component of GINS, via the unique "B domain" of the latter that is dispensable for assembly of the GINS complex but is essential for replication initiation. We show that Dpb2NT is required during initiation for assembly of the Cdc45-MCM-GINS helicase. Moreover, overexpressed Dpb2NT is sufficient to support assembly of the Cdc45-MCM-GINS helicase during initiation, upon depletion of endogenous Dpb2. This produces a replisome that lacks DNA polymerase epsilon, and although cells are viable, they grow extremely poorly. Finally, we use a novel in vitro assay to show that Dpb2NT is essential for Pol ε to interact with the replisome after initiation.

Conclusions: These findings indicate that the association of Dpb2 with the B domain of Psf1 plays two critical roles during chromosome replication in budding yeast. First, it is required for initiation, because it facilitates the incorporation of GINS into the Cdc45-MCM-GINS helicase at nascent forks. Second, it plays an equally important role after initiation, because it links the leading strand DNA polymerase to the Cdc45-MCM-GINS helicase within the replisome.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Cycle Proteins / metabolism
  • Chromatography, Gel
  • DNA Helicases / metabolism*
  • DNA Polymerase II / chemistry
  • DNA Polymerase II / metabolism*
  • DNA Replication / genetics*
  • DNA-Binding Proteins / metabolism*
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli
  • Models, Molecular*
  • Multiprotein Complexes / chemistry
  • Multiprotein Complexes / metabolism*
  • Nuclear Proteins / metabolism*
  • Protein Structure, Tertiary
  • Ribonucleoprotein, U4-U6 Small Nuclear / chemistry
  • Ribonucleoprotein, U4-U6 Small Nuclear / metabolism*
  • Ribonucleoprotein, U5 Small Nuclear / chemistry
  • Ribonucleoprotein, U5 Small Nuclear / metabolism*
  • Rosaniline Dyes
  • Saccharomyces cerevisiae Proteins / chemistry
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Saccharomycetales
  • Spectrophotometry, Ultraviolet
  • Two-Hybrid System Techniques


  • CDC45 protein, S cerevisiae
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • MRC1 protein, S cerevisiae
  • Multiprotein Complexes
  • Nuclear Proteins
  • PRP8 protein, S cerevisiae
  • Ribonucleoprotein, U4-U6 Small Nuclear
  • Ribonucleoprotein, U5 Small Nuclear
  • Rosaniline Dyes
  • Saccharomyces cerevisiae Proteins
  • Coomassie blue
  • Dpb2 protein, S cerevisiae
  • DNA Polymerase II
  • DNA Helicases