Sequence-specific modification of mitochondrial DNA using a chimeric zinc finger methylase
Open accessAbstract
We used engineered zinc finger peptides (ZFPs) to bind selectively to predetermined sequences in human mtDNA. Surprisingly, we found that engineered ZFPs cannot be reliably routed to mitochondria by using only conventional mitochondrial targeting sequences. We here show that addition of a nuclear export signal allows zinc finger chimeric enzymes to be imported into human mitochondria. The selective binding of mitochondria-specific ZFPs to mtDNA was exemplified by targeting the T8993G mutation, which causes two mitochondrial diseases, neurogenic muscle weakness, ataxia, and retinitis pigmentosa (NARP) and also maternally inherited Leigh's syndrome. To develop a system that allows the monitoring of site-specific alteration of mtDNA we combined a ZFP with the easily assayed DNA-modifying activity of hDNMT3a methylase. Expression of the mutation-specific chimeric methylase resulted in the selective methylation of cytosines adjacent to the mutation site. This is a proof of principle that it is possible to target and alter mtDNA in a sequence-specific manner by using zinc finger technology.
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Targeted chimaeric zinc finger DNA methylase can bind and modify mutant genomes in cells with m.8993T N G NARP mutation.
“Selective targeting of mutated mtDNA has also been successfully attempted with a targeted chimaeric zinc finger DNA methylase, which can bind and modify, in a sequence-specific manner, mutant genomes in cells harbouring the m.8993T N G NARP mutation [237].”
From Mitochondrial DNA mutations and human disease (2010) · cites this paper as [237]
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Mitochondrial DNA mutations and human disease
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- Authors
- Minczuk, M., Papworth, M. A., Kolasinska, P., Murphy, M. P., & Klug, A.
- Affiliations
- MRC Laboratory of Molecular Biology (GB)
- *Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, United Kingdom; and
- Medical Research Council (GB)
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 2006-12-26
- Volume
- 103
- Issue
- 52
- Pages
- 19689-19694
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC1750892
- OpenAlex
- W1995146753
- Cited by
- 166
- References
- 34
- Citations by year
- 2026: 4 · 2025: 8 · 2024: 5 · 2023: 11 · 2022: 14 · 2021: 7 · 2020: 10 · 2019: 9 · 2018: 13 · 2017: 9 · 2016: 11 · 2015: 7 · 2014: 12 · 2013: 2 · 2012: 9
- FWCI
- 2.31
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Medical Research Council
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
- Keywords
- mitochondrial DNA, mitochondrial targeting, nuclear export signal, NARP, mitochondrial disease, cytosine methylation
- MeSH terms
- Amino Acid Sequence, Animals, Base Sequence, Cell Line, Chlorocebus aethiops, DNA, Mitochondrial, Mitochondria, Models, Molecular, Mutation, Protein Binding, Recombinant Proteins, Substrate Specificity, DNA Modification Methylases, Zinc Fingers, Protein Structure, Tertiary, DNA Methylation, Protein Transport
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