Dominant negative rat polymerase β mutants interfere with base excision repair in saccharomyces cerevisiae

Caroline A. Clairmont, Joann B. Sweasy

Research output: Contribution to journalArticlepeer-review

Abstract

DNA polymerase β (Pol β) is one of the smallest known eukaryotic DNA polymerases. Mutations in DNA polymerase β have been detected in both colon and prostatic tumors. Therefore, investigating the in vivo role of Pol β may provide important insights into a cells' ability to maintain its DNA integrity as well the prevention of tumorigenesis. This polymerase has been very well characterized in vitro, and has been shown to be involved in gap filling during base excision repair. In mammalian cells Pol β is also thought to play a role in base excision repair in vivo however, nothing is known about its in vivo role in Saccharomyces cerevisiae. Previous attempts to disrupt DNA polymerase IV, a putative Pol β homologue in Saccharomyces cerevisiae, in vivo by gene disruption techniques have not been informative, most likely due to redundancy in this system, whereby other polymerases are able to substitute for DNA polymerase IV. To overcome this problem we have isolated two dominant negative mutants of Pol β, using a novel competition assay we developed in E. coli. When we overexpressed the dominant negative Pol β mutant proteins in Saccharomyces cerevisiae, the cells became sensitive to methyl methanesulfonate. Interestingly, overexpression of the same polymerase β mutant proteins did not confer sensitivity to UV damage, strongly suggesting that the mutant proteins interfere with the process of base excision repair, but not nucleotide excision repair, in S. cerevisiae. Our data implicate a role for polymerase IV, the S. cerevisiae polymerase β homologue, in base excision repair in yeast.

Original languageEnglish (US)
Pages (from-to)A960
JournalFASEB Journal
Volume10
Issue number6
StatePublished - 1996
Externally publishedYes

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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