Cited by 70DNA Repair MechanismsBacterial Genetics and BiotechnologyDNA and Nucleic Acid Chemistry

Removal of Misincorporated Ribonucleotides from Prokaryotic Genomes: An Unexpected Role for Nucleotide Excision Repair

Open access
Vaisman, Alexandra, McDonald, John P., Huston, Donald, Kuban, Wojciech, Liu, Lili, Van Houten, Bennett, Woodgate, Roger·2013·PLoS Genetics

Abstract

Stringent steric exclusion mechanisms limit the misincorporation of ribonucleotides by high-fidelity DNA polymerases into genomic DNA. In contrast, low-fidelity Escherichia coli DNA polymerase V (pol V) has relatively poor sugar discrimination and frequently misincorporates ribonucleotides. Substitution of a steric gate tyrosine residue with alanine (umuC_Y11A) reduces sugar selectivity further and allows pol V to readily misincorporate ribonucleotides as easily as deoxynucleotides, whilst leaving its poor base-substitution fidelity essentially unchanged. However, the mutability of cells expressing the steric gate pol V mutant is very low due to efficient repair mechanisms that are triggered by the misincorporated rNMPs. Comparison of the mutation frequency between strains expressing wild-type and mutant pol V therefore allows us to identify pathways specifically directed at ribonucleotide excision repair (RER). We previously demonstrated that rNMPs incorporated by umuC_Y11A are efficiently removed from DNA in a repair pathway initiated by RNase HII. Using the same approach, we show here that mismatch repair and base excision repair play minimal back-up roles in RER in vivo. In contrast, in the absence of functional RNase HII, umuC_Y11A-dependent mutagenesis increases significantly in ΔuvrA, uvrB5 and ΔuvrC strains, suggesting that rNMPs misincorporated into DNA are actively repaired by nucleotide excision repair (NER) in vivo. Participation of NER in RER was confirmed by reconstituting ribonucleotide-dependent NER in vitro. We show that UvrABC nuclease-catalyzed incisions are readily made on DNA templates containing one, two, or five rNMPs and that the reactions are stimulated by the presence of mispaired bases. Similar to NER of DNA lesions, excision of rNMPs proceeds through dual incisions made at the 8(th) phosphodiester bond 5' and 4(th)-5(th) phosphodiester bonds 3' of the ribonucleotide. Ribonucleotides misinserted into DNA can therefore be added to the broad list of helix-distorting modifications that are substrates for NER.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1371/journal.pgen.1003878

Cites

71 of 71 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 1

  1. [59]Cone R, Duncan J, Hamilton L, Friedberg EC (1977) Partial purification and characterization of a uracil DNA N-glycosidase from Bacillus subtilis. Biochemistry 16: 3194– 3201. 407925

Cited by

Sorted by citations

Authors
Vaisman, A., McDonald, J. P., Huston, D., Kuban, W., Liu, L., Van Houten, B., & Woodgate, R.
Affiliations
  • National Institutes of Health (US)
  • UPMC Hillman Cancer Center (US)
Corresponding author
Woodgate, Roger
Venue
PLoS Genetics
Published
2013-11-07
Volume
9
Issue
11
Pages
e1003878
ISSN
  • 1553-7390 (ISSN-L)
  • 1553-7404
OpenAlex
W2144683765
Cited by
70
References
71
Citations by year
2026: 3 · 2025: 1 · 2024: 1 · 2023: 6 · 2022: 5 · 2021: 8 · 2020: 1 · 2019: 8 · 2018: 3 · 2017: 8 · 2016: 4 · 2015: 10 · 2014: 11 · 2013: 1
FWCI
3.88
Type
journal article
Language
en
Open access
Locations
  • PLoS Genetics (publishedVersion, public-domain)
  • Europe PMC (PubMed Central) (publishedVersion, public-domain)
  • PubMed Central (publishedVersion, public-domain)
  • PubMed (publishedVersion)
  • D-Scholarship@Pitt (University of Pittsburgh) (acceptedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • Figshare (submittedVersion, cc-by)
Funders
  • National Institutes of Health
  • National Institute of Environmental Health Sciences (R01ES019566)
  • National Institute of Child Health and Human Development
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
Keywords
nucleotide excision repair, ribonucleotide excision repair, DNA polymerase V, UmuC, RNase HII, mutagenesis, DNA mismatch repair, base excision repair, Escherichia coli, prokaryotic genomes
MeSH terms
DNA, DNA-Directed DNA Polymerase, DNA Repair, Endodeoxyribonucleases, Escherichia coli, Mutation, Prokaryotic Cells, Ribonucleotides, Genome, Bacterial, Ribonuclease H, Amino Acid Substitution, Escherichia coli Proteins

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review