Ubc4 and Not4 regulate steady-state levels of DNA polymerase-α to promote efficient and accurate DNA replication

Mol Biol Cell. 2010 Sep 15;21(18):3205-19. doi: 10.1091/mbc.E09-06-0452. Epub 2010 Jul 21.

Abstract

The accurate duplication of chromosomal DNA is required to maintain genomic integrity. However, from an evolutionary point of view, a low mutation rate during DNA replication is desirable. One way to strike the right balance between accuracy and limited mutagenesis is to use a DNA polymerase that lacks proofreading activity but contributes to DNA replication in a very restricted manner. DNA polymerase-α fits this purpose exactly, but little is known about its regulation at the replication fork. Minichromosome maintenance protein (Mcm) 10 regulates the stability of the catalytic subunit of pol-α in budding yeast and human cells. Cdc17, the catalytic subunit of pol-α in yeast, is rapidly degraded after depletion of Mcm10. Here we show that Ubc4 and Not4 are required for Cdc17 destabilization. Disruption of Cdc17 turnover resulted in sensitivity to hydroxyurea, suggesting that this pathway is important for DNA replication. Furthermore, overexpression of Cdc17 in ubc4 and not4 mutants caused slow growth and synthetic dosage lethality, respectively. Our data suggest that Cdc17 levels are very tightly regulated through the opposing forces of Ubc4 and Not4 (destabilization) and Mcm10 (stabilization). We conclude that regular turnover of Cdc17 via Ubc4 and Not4 is required for proper cell proliferation.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • Cell Nucleus / metabolism
  • Cell Proliferation
  • Chromosomal Proteins, Non-Histone / genetics
  • Chromosomal Proteins, Non-Histone / metabolism
  • DNA Polymerase I / genetics
  • DNA Polymerase I / metabolism*
  • DNA Replication*
  • Genomic Instability
  • Humans
  • Minichromosome Maintenance Proteins
  • Mutation
  • Proteasome Endopeptidase Complex / metabolism
  • Repressor Proteins
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / physiology
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Ubiquitin-Conjugating Enzymes / genetics
  • Ubiquitin-Conjugating Enzymes / metabolism*
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*

Substances

  • Cell Cycle Proteins
  • Chromosomal Proteins, Non-Histone
  • MCM10 protein, S cerevisiae
  • MCM10 protein, human
  • Repressor Proteins
  • Saccharomyces cerevisiae Proteins
  • Ubc4 protein, S cerevisiae
  • Ubiquitin-Conjugating Enzymes
  • MOT2 protein, S cerevisiae
  • Ubiquitin-Protein Ligases
  • DNA Polymerase I
  • POL1 protein, S cerevisiae
  • Proteasome Endopeptidase Complex
  • Minichromosome Maintenance Proteins