Dataset Information

The absence of the catalytic domains of Saccharomyces cerevisiae DNA polymerase ? strongly reduces DNA replication fidelity.

ABSTRACT: The four B-family DNA polymerases ?, ?, ? and ? cooperate to accurately replicate the eukaryotic nuclear genome. Here, we report that a Saccharomyces cerevisiae strain encoding the pol2-16 mutation that lacks Pol ?'s polymerase and exonuclease activities has increased dNTP concentrations and an increased mutation rate at the CAN1 locus compared to wild type yeast. About half of this mutagenesis disappears upon deleting the REV3 gene encoding the catalytic subunit of Pol ?. The remaining, still strong, mutator phenotype is synergistically elevated in an msh6? strain and has a mutation spectrum characteristic of mistakes made by Pol ?. The results support a model wherein slow-moving replication forks caused by the lack of Pol ?'s catalytic domains result in greater involvement of mutagenic DNA synthesis by Pol ? as well as diminished proofreading by Pol ? during replication.

SUBMITTER: Garbacz MA

PROVIDER: S-EPMC6486559 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The absence of the catalytic domains of Saccharomyces cerevisiae DNA polymerase ϵ strongly reduces DNA replication fidelity.

Garbacz Marta A MA Cox Phillip B PB Sharma Sushma S Lujan Scott A SA Chabes Andrei A Kunkel Thomas A TA

Nucleic acids research 20190501 8

The four B-family DNA polymerases α, δ, ϵ and ζ cooperate to accurately replicate the eukaryotic nuclear genome. Here, we report that a Saccharomyces cerevisiae strain encoding the pol2-16 mutation that lacks Pol ϵ's polymerase and exonuclease activities has increased dNTP concentrations and an increased mutation rate at the CAN1 locus compared to wild type yeast. About half of this mutagenesis disappears upon deleting the REV3 gene encoding the catalytic subunit of Pol ζ. The remaining, still s ...[more]

PMID: 30698744

Similar Datasets

Project description:Humans are exposed to N-nitroso compounds (NOCs) both endogenously and exogenously from a number of environmental sources, and NOCs are both mutagenic and carcinogenic. After metabolic activation, some NOCs can induce carboxymethylation of nucleobases through a diazoacetate intermediate, which could give rise to p53 mutations similar to those seen in human gastrointestinal cancers. It was previously found that the growth of polymerase ?-deficient human cells was inhibited by treatment with azaserine, a DNA carboxymethylation agent, suggesting the importance of this polymerase in bypassing the azaserine-induced carboxymethylated DNA lesions. In this study, we examined how carboxymethylated DNA lesions, which included N(6)-carboxymethyl-2'-deoxyadenosine (N(6)-CMdA), N(4)-carboxymethyl-2'-deoxycytidine (N(4)-CMdC), N3-carboxymethylthymidine (N3-CMdT), and O(4)-carboxymethylthymidine (O(4)-CMdT), perturbed the efficiency and fidelity of DNA replication mediated by Saccharomyces cerevisiae polymerase ? (pol ?). Our results from steady-state kinetic assay showed that pol ? could readily bypass and extend past N(6)-CMdA and incorporated the correct nucleotides opposite the lesion and its neighboring 5'-nucleoside with high efficiency. By contrast, the polymerase could bypass N(4)-CMdC inefficiently, with substantial misincorporation of dCMP followed by dAMP, though pol ? could extend past the lesion with high fidelity and efficiency when dGMP was incorporated opposite the lesion. On the other hand, yeast pol ? experienced great difficulty in bypassing O(4)-CMdT and N3-CMdT, and the polymerase inserted preferentially the incorrect dGMP opposite these two DNA lesions; the extension step, nevertheless, occurred with high fidelity and efficiency when the correct dAMP was opposite the lesion, as opposed to the preferentially incorporated incorrect dGMP. These results suggest that these lesions may contribute significantly to diazoacetate-induced mutations and those in the p53 gene observed in human gastrointestinal tumors.

Dataset Information

The absence of the catalytic domains of Saccharomyces cerevisiae DNA polymerase ? strongly reduces DNA replication fidelity.

Publications

The absence of the catalytic domains of Saccharomyces cerevisiae DNA polymerase ϵ strongly reduces DNA replication fidelity.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets