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Updated: Jun 21, 2026
Visualization of DNA Repair Proteins Interaction by Immunofluorescence
Published on: June 26, 2020
MRE11-RAD50-NBS1 is a critical regulator of FANCD2 stability and function during DNA double-strand break repair
Céline Roques1, Yan Coulombe, Mathieu Delannoy
1Genome Stability Laboratory, Laval University Cancer Research Center, Hôtel-Dieu de Québec, Québec, Canada.
Abstract:
Monoubiquitination of the Fanconi anaemia protein FANCD2 is a key event leading to repair of interstrand cross-links. It was reported earlier that FANCD2 co-localizes with NBS1. However, the functional connection between FANCD2 and MRE11 is poorly understood. In this study, we show that inhibition of MRE11, NBS1 or RAD50 leads to a destabilization of FANCD2. FANCD2 accumulated from mid-S to G2 phase within sites containing single-stranded DNA (ssDNA) intermediates, or at sites of DNA damage, such as those created by restriction endonucleases and laser irradiation. Purified FANCD2, a ring-like particle by electron microscopy, preferentially bound ssDNA over various DNA substrates. Inhibition of MRE11 nuclease activity by Mirin decreased the number of FANCD2 foci formed in vivo. We propose that FANCD2 binds to ssDNA arising from MRE11-processed DNA double-strand breaks. Our data establish MRN as a crucial regulator of FANCD2 stability and function in the DNA damage response.
Insights
The MRN complex is essential for stabilizing FANCD2, a key protein in DNA repair. This study reveals MRN regulates FANCD2
Area of Science:
- Molecular Biology
- DNA Repair Mechanisms
- Cellular Response to DNA Damage
Background:
- Monoubiquitination of FANCD2 is critical for repairing interstrand DNA cross-links.
- FANCD2 is known to co-localize with NBS1, but its functional link with MRE11 remains unclear.
Purpose of the Study:
- To investigate the functional connection between FANCD2 and the MRE11-NBS1-RAD50 (MRN) complex in DNA damage response.
- To elucidate the role of MRN in FANCD2 stability and localization.
Main Methods:
- Inhibition of MRN complex components (MRE11, NBS1, RAD50) to assess FANCD2 stability.
- Induction of DNA damage using restriction endonucleases and laser irradiation.
- Analysis of FANCD2 foci formation and ssDNA binding properties.
- Utilizing Mirin to inhibit MRE11 nuclease activity.
Main Results:
- Inhibition of MRE11, NBS1, or RAD50 led to FANCD2 destabilization.
- FANCD2 accumulated at sites of single-stranded DNA (ssDNA) and DNA damage during S to G2 phase.
- Purified FANCD2 showed preferential binding to ssDNA.
- MRE11 nuclease inhibition by Mirin reduced FANCD2 foci formation in vivo.
Conclusions:
- The MRN complex is a critical regulator of FANCD2 stability and function.
- FANCD2 likely binds to ssDNA generated by MRE11-processed DNA double-strand breaks.
- These findings establish MRN's crucial role in the DNA damage response pathway involving FANCD2.
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