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Ribonucleotide incorporation by yeast DNA polymerase ζ.

Abstract
During replication in yeast, the three B family DNA replicases frequently incorporate ribonucleotides (rNMPs) into DNA, and their presence in the nuclear genome can affect genome stability. This prompted us to examine ribonucleotide incorporation by the fourth B family member, Pol ζ, the enzyme responsible for the majority of damage-induced mutagenesis in eukaryotes. We first show that Pol ζ inserts rNMPs into DNA and can extend primer termini containing 3'-ribonucleotides. We then measure rNMP incorporation by Pol ζ in the presence of its cofactors, RPA, RFC and PCNA and at normal cellular dNTP and rNTP concentrations that exist under unstressed conditions. Under these conditions, Pol ζ stably incorporates one rNMP for every 200-300 dNMPs incorporated, a frequency that is slightly higher than for the high fidelity replicative DNA polymerases. Under damage-induced conditions wherein cellular dNTP concentrations are elevated 5-fold, Pol ζ only incorporates one rNMP per 1300 dNMPs. Functional interaction of Pol ζ with the mutasome assembly factor Rev1 gives comparable rNMP incorporation frequencies. These results suggest that ribonucleotide incorporation into DNA during Pol ζ-mediated mutagenesis in vivo may be rare.
AuthorsAlena V Makarova, Stephanie A Nick McElhinny, Brian E Watts, Thomas A Kunkel, Peter M Burgers
JournalDNA repair (DNA Repair (Amst)) Vol. 18 Pg. 63-7 (Jun 2014) ISSN: 1568-7856 [Electronic] Netherlands
PMID24674899 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, N.I.H., Intramural)
CopyrightCopyright © 2014 Elsevier B.V. All rights reserved.
Chemical References
  • DNA, Fungal
  • Deoxyribonucleotides
  • REV7 protein, S cerevisiae
  • RFA1 protein, S cerevisiae
  • Replication Protein A
  • Ribonucleotides
  • Saccharomyces cerevisiae Proteins
  • DNA polymerase zeta
  • Nucleotidyltransferases
  • REV1 protein, S cerevisiae
  • DNA-Directed DNA Polymerase
  • REV3 protein, S cerevisiae
Topics
  • DNA Damage
  • DNA Replication
  • DNA, Fungal (metabolism)
  • DNA-Directed DNA Polymerase (metabolism)
  • Deoxyribonucleotides (metabolism)
  • Mutation
  • Nucleotidyltransferases (genetics, metabolism)
  • Replication Protein A (metabolism)
  • Ribonucleotides (metabolism)
  • Saccharomyces cerevisiae (enzymology)
  • Saccharomyces cerevisiae Proteins (genetics, metabolism)

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