Abstract
Mutations in mitochondrial DNA (mtDNA) are responsible for several severe diseases that have no available cures. The multicopy nature of the mitochondrial genome means that mutations often exist in a state known as heteroplasmy, where both mutant and wild-type mtDNA are present in the same cell. The wild-type mtDNA can functionally compensate for the mutant mtDNA until a mutation threshold is reached, beyond which disease symptoms begin to manifest. Despite the interest mitochondrial genetics has generated, the double mitochondrial membrane proved to be a formidable barrier to genetic manipulation. However, in the past two decades, scientists have discovered that mtDNA could be modified by importing gene editing proteins to target specific DNA sequences. Mitochondria-targeted nucleases specifically cleave and eliminate mutant mtDNA in heteroplasmic cells and in animal models. More recently, base editors have been adapted to modify mtDNA via precise C>T or A>G transitions. Therefore, tools to modify mtDNA are, finally, a reality with the promise to revolutionize the mitochondrial genetics field. This Primer delves into mitochondrial gene editing, providing details on the selection of mitochondrial gene editing tools, best practices for designing experiments, relevant types of analyses and specific applications and limitations pertaining to the different technologies and the field.
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References
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Acknowledgements
The authors thank C. L. Gorsuch for her suggestions related to this work. The work in the Moraes laboratory was funded by National Institute of Health (NIH) Grants 5R01EY010804 and 1R01NS079965, the Florida Biomedical Foundation (21K05), the Muscular Dystrophy Association (MDA 964119) and the Army Research Office (W911NF-21-1-0248).
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Contributions
Introduction (W.K.S. and C.T.M.); Experimentation (W.K.S., S.R.B., J.D.B.-P. and C.T.M.); Results (W.K.S., S.R.B., J.D.B.-P. and C.T.M.); Applications (W.K.S., S.R.B., J.D.B.-P. and C.T.M.); Reproducibility and data deposition (C.T.M.); Limitations and optimizations (W.K.S., S.R.B., J.D.B.-P. and C.T.M.); Outlook (W.K.S. and C.T.M.); Overview of the Primer (W.K.S., S.R.B., J.D.B.-P. and C.T.M.).
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W.K.S. is employed by Precision BioSciences. All other authors declare no competing interests.
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Nature Reviews Methods Primers thanks Gino Cortopassi, Giovanni Manfredi, Chengqi Yi and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
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Glossary
- Cytoplasmic hybrid
-
A cell derived from the fusion of an enucleated patient cell with a cell lacking mitochondrial DNA.
- Heteroduplex
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A double-stranded DNA sequence that contains a mismatch.
- Heteroplasmy
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A state in which a cell contains more than one mitochondrial DNA haplotype.
- Homoplasmic
-
Describes a state in which all of the mitochondrial DNA molecules are identical.
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Shoop, W.K., Bacman, S.R., Barrera-Paez, J.D. et al. Mitochondrial gene editing. Nat Rev Methods Primers 3, 19 (2023). https://doi.org/10.1038/s43586-023-00200-7
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DOI: https://doi.org/10.1038/s43586-023-00200-7
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