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Updated: Feb 19, 2026
Proofreading and DNA Repair Assay Using Single Nucleotide Extension and MALDI-TOF Mass Spectrometry Analysis
Published on: June 19, 2018
DNA polymerase β: A missing link of the base excision repair machinery in mammalian mitochondria
Rajendra Prasad1, Melike Çağlayan1, Da-Peng Dai1
1Genome Integrity and Structural Biology Laboratory, National Institutes of Health, NIEHS, 111 T.W. Alexander Drive, P.O. Box 12233, Research Triangle Park, NC 27709, USA.
Abstract:
Mitochondrial genome integrity is fundamental to mammalian cell viability. Since mitochondrial DNA is constantly under attack from oxygen radicals released during ATP production, DNA repair is vital in removing oxidatively generated lesions in mitochondrial DNA, but the presence of a strong base excision repair system has not been demonstrated. Here, we addressed the presence of such a system in mammalian mitochondria involving the primary base lesion repair enzyme DNA polymerase (pol) β. Pol β was localized to mammalian mitochondria by electron microscopic-immunogold staining, immunofluorescence co-localization and biochemical experiments. Extracts from purified mitochondria exhibited base excision repair activity that was dependent on pol β. Mitochondria from pol β-deficient mouse fibroblasts had compromised DNA repair and showed elevated levels of superoxide radicals after hydrogen peroxide treatment. Mitochondria in pol β-deficient fibroblasts displayed altered morphology by electron microscopy. These results indicate that mammalian mitochondria contain an efficient base lesion repair system mediated in part by pol β and thus pol β plays a role in preserving mitochondrial genome stability.
Insights
Mammalian mitochondria possess a base excision repair system, partly mediated by DNA polymerase beta (pol β), crucial for maintaining mitochondrial genome stability and cell viability.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- Mitochondrial genome integrity is essential for cell survival.
- Mitochondrial DNA (mtDNA) is susceptible to oxidative damage from ATP production.
- A robust base excision repair (BER) system in mitochondria has not been definitively established.
Purpose of the Study:
- To investigate the presence and role of a base excision repair system in mammalian mitochondria.
- To determine if DNA polymerase beta (pol β) is involved in mitochondrial DNA repair.
Main Methods:
- Localization of pol β in mitochondria using electron microscopy and immunofluorescence.
- Biochemical assays to assess base excision repair activity in mitochondrial extracts.
- Analysis of DNA repair, oxidative stress, and mitochondrial morphology in pol β-deficient fibroblasts.
Main Results:
- DNA polymerase beta (pol β) was successfully localized to mammalian mitochondria.
- Mitochondrial extracts demonstrated pol β-dependent base excision repair activity.
- Pol β-deficient mitochondria exhibited impaired DNA repair, increased superoxide radicals post-H2O2 treatment, and altered morphology.
Conclusions:
- Mammalian mitochondria possess an effective base lesion repair system.
- DNA polymerase beta (pol β) plays a significant role in this mitochondrial repair system.
- Pol β is vital for preserving mitochondrial genome stability and overall cell viability.
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