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PreprintCited by 11DNA Repair MechanismsNuclear Structure and FunctionGenetics, Aging, and Longevity in Model Organisms

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Recovery of protein synthesis to measure transcription-coupled DNA repair in living cells and tissues

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van der Woude, Melanie, Davó-Martínez, Carlota, Thijssen, Karen L., Vermeulen, Wim, Lans, Hannes·2023·Nucleic Acids Research
DOI 10.1101/2023.01.24.525327PMC PMC10570043

Preprint — published as 10.1093/nar/gkad642 · Paper page

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Abstract

Transcription-coupled nucleotide excision repair (TC-NER) is an important DNA repair mechanism that protects against the negative effects of transcription-blocking DNA lesions. Hereditary TC-NER deficiency causes pleiotropic and often severe neurodegenerative and progeroid symptoms. Multiple assays have been developed for the clinic and for research to measure TC-NER activity, which is hampered by the relatively low abundance of repair events taking place in transcribed DNA. ‘Recovery of RNA Synthesis’ is widely used as indirect TC-NER assay based on the notion that lesion-blocked transcription only resumes after successful TC-NER. Here, we show that measuring novel synthesis of a protein that has been degraded prior to DNA damage induction is an equally effective but more versatile manner to indirectly monitor TC-NER. This ‘Recovery of Protein Synthesis’ (RPS) assay is readily adaptable for use with different degradable proteins and readouts, including fluorescence imaging and immunoblot. Moreover, with the RPS assay TC-NER activity can be measured in real-time, in various living cells types and even in differentiated tissues of living organisms. As example, we show that TC-NER capacity declines in aging muscle tissue of C. elegans . Therefore, the RPS assay constitutes an important novel clinical and research tool to investigate transcription-coupled DNA repair.

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Metadata

Authors
van der Woude, M., Davó-Martínez, C., Thijssen, K. L., Vermeulen, W., & Lans, H.
Affiliations
Erasmus MC (NL)
Corresponding author
Lans, Hannes
ORCID
Venue
Nucleic Acids Research
Publisher
openRxiv
Published
2023-01-24
Volume
51
Issue
18
Pages
e93-e93
ISSN
0305-1048 (ISSN-L)
DOI
10.1101/2023.01.24.525327
PMC
PMC10570043
OpenAlex
W4385406426
Cited by
11
References
1
Citations by year
2026: 4 · 2025: 5 · 2024: 2
FWCI
0.98
Type
journal article
Language
en
Open access
Licence
cc-by
Locations
  • Nucleic Acids Research (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • EUR Research Repository (Erasmus University Rotterdam) (publishedVersion, cc-by)
Funders
  • Nederlandse Organisatie voor Wetenschappelijk Onderzoek (711.018.007, ALWOP.494)
  • KWF Kankerbestrijding
  • Oncode Institute
  • National Institutes of Health (OD010440, P40OD010440)
  • H2020 European Research Council (340988‐ERC‐ID)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Nuclear Structure and Function
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Aging › Genetics, Aging, and Longevity in Model Organisms
Keywords
transcription-coupled repair, transcribed genes, protein degradation, Caenorhabditis elegans
MeSH terms
Aging, Animals, DNA, DNA Damage, DNA Repair, Muscles, Transcription, Genetic, Protein Biosynthesis, Caenorhabditis elegans

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