RRC ID 80898
Author van der Woude M, Davó-Martínez C, Thijssen KL, Vermeulen W, Lans H.
Title Recovery of protein synthesis to assay DNA repair activity in transcribed genes in living cells and tissues.
Journal Nucleic Acids Res
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 deficiencies cause pleiotropic and often severe neurodegenerative and progeroid symptoms. While multiple assays have been developed to determine TC-NER activity for clinical and research purposes, monitoring TC-NER is hampered by the low frequency of repair events occurring 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 after its compound-induced degradation prior to DNA damage induction is an equally effective but more versatile manner to indirectly monitor DNA repair activity in transcribed genes. This 'Recovery of Protein Synthesis' (RPS) assay can be adapted to various degradable proteins and readouts, including imaging and immunoblotting. Moreover, RPS allows real-time monitoring of TC-NER activity in various living cells types and even in differentiated tissues of living organisms. To illustrate its utility, we show that DNA repair in transcribed genes 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.
Volume 51(18)
Pages e93
Published 2023-10-13
DOI 10.1093/nar/gkad642
PII 7233916
PMID 37522336
PMC PMC10570043
MeSH Aging / metabolism Animals Caenorhabditis elegans* / physiology DNA / metabolism DNA Damage DNA Repair* Muscles / metabolism Protein Biosynthesis* Transcription, Genetic*
C.elegans tm3886