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Updated: Jan 10, 2026
Visualizing and Quantifying Endonuclease-Based Site-Specific DNA Damage
Published on: August 21, 2021
Engineered chromatin readers track damaged chromatin dynamics in live cells and animals
Richard Cardoso da Silva1, Kristeli Eleftheriou2, Davide C Recchia2
1Genome Biology and Epigenetics, Institute of Biodynamics and Biocomplexity, Department of Biology, Utrecht University, Utrecht, The Netherlands. richicardoso.silva@gmail.com.
Abstract:
DNA damage is a constant threat to genome integrity and function. Diminished capacity for DNA repair is linked to many human diseases, therefore, understanding the molecular pathways responding to DNA damage is key for developing novel therapies. Lack of unbiased probes to report DNA damage dynamics in living cells and animals limits our current efforts to completely understand DNA repair processes. In this study, we overcome these limitations by engineering protein probes containing the tandem-BRCT domain of MCPH1, which we show to have a specific affinity for the DNA-damage-associated histone mark γH2AX. We employ these probes to track DNA damage dynamics in living cells exposed to a panel of different genotoxic insults, to visualize DNA damage targeted to heterochromatinised satellite repeats, and to map DNA double strand breaks genome-wide. Finally, we highlight the versatility of our probe to visualize programmed double strand breaks during gametogenesis in C. elegans. Taken together, we present a novel protein probe with broad application potential for DNA damage research.
Insights
Researchers developed a novel protein probe to track DNA damage dynamics in living cells and animals. This tool aids in understanding DNA repair mechanisms and developing new therapies for diseases linked to DNA damage.
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- DNA damage poses a continuous threat to genome integrity.
- Impaired DNA repair is implicated in various human diseases.
- Current methods lack unbiased probes for real-time DNA damage monitoring in vivo.
Purpose of the Study:
- To engineer and validate a novel protein probe for visualizing DNA damage dynamics.
- To overcome limitations in current DNA repair research tools.
- To enable precise tracking of DNA damage in living systems.
Main Methods:
- Engineered protein probes utilizing the tandem-BRCT domain of MCPH1.
- Assessed probe affinity for the DNA-damage marker γH2AX.
- Tracked DNA damage dynamics in living cells under genotoxic stress.
- Visualized DNA damage in heterochromatin and mapped double-strand breaks genome-wide.
Main Results:
- Developed a protein probe with specific affinity for γH2AX.
- Successfully tracked DNA damage dynamics in living cells and visualized damage in heterochromatin.
- Enabled genome-wide mapping of DNA double-strand breaks.
- Demonstrated probe utility in visualizing programmed DNA breaks during C. elegans gametogenesis.
Conclusions:
- Presented a novel protein probe for versatile DNA damage research.
- The probe offers broad application potential for studying DNA repair pathways.
- Facilitates a deeper understanding of genome integrity maintenance and disease mechanisms.
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