How gene editing can address mitochondrial DNA?
Image credit: http://www.cellimagelibrary.org/images/11397
Sun N, Youle RJ, Finkel T. The Mitochondrial Basis of Aging. Mol Cell. 2016;61(5):654-666. doi:10.1016/j.molcel.2016.01.028
Heidi Chial, Ph.D. (Write Science Right) & Joanna Craig, Ph.D. (Write Science Right) © 2008 Nature Education Citation: Chial, H. & Craig, J. (2008) mtDNA and mitochondrial diseases. Nature Education 1(1):217
A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity BY MARTIN JINEK, KRZYSZTOF CHYLINSKI, INES FONFARA, MICHAEL HAUER, JENNIFER A. DOUDNA, EMMANUELLE CHARPENTIER SCIENCE17 AUG 2012 : 816-821
Hirakawa MP, Krishnakumar R, Timlin JA, Carney JP, Butler KS. Gene editing and CRISPR in the clinic: current and future perspectives. Biosci Rep. 2020;40(4):BSR20200127. doi:10.1042/BSR20200127
Gammage PA, Moraes CT, Minczuk M. Mitochondrial Genome Engineering: The Revolution May Not Be CRISPR-Ized. Trends Genet. 2018;34(2):101-110. doi:10.1016/j.tig.2017.11.001
Using TALEs and Base editing to edit mtDNA
Delivering the gRNA expression cassette as a self-replicating plasmid
- Can we apply this in vitro delivery method to human cells? Or at least can we find a similar approach that allows us to deliver self-replicating plasmid DNA to our mitochondria? No such thing has been published so far, but I'm sure this issue will be addressed in the future, maybe with adaptations of technologies such as MITO-Porter
- Is expression of Cas9 going to be constant in transformed mitochondria? Could that lead to off-target effects that could deplete mitochondrial DNA? In that case, maybe it would be interesting to deliver Cas9 as a protein, to avoid its long-term expression, while delivering the gRNA and potential donor DNA as a plasmid. However, there are yet no reports of such a strategy being effective for mitochondrial gene editing.
- Is this going to be a better approach than just using TALENs of Base Editing?
Bonnefoy N, Fox TD. Directed alteration of Saccharomyces cerevisiae mitochondrial DNA by biolistic transformation and homologous recombination. Methods Mol Biol. 2007;372:153-166. doi:10.1007/978-1-59745-365-3_11
Yoo BC, Yadav NS, Orozco EM Jr, Sakai H. Cas9/gRNA-mediated genome editing of yeast mitochondria and Chlamydomonas chloroplasts. PeerJ. 2020;8:e8362. Published 2020 Jan 6. doi:10.7717/peerj.8362
Yamada Y, Akita H, Kamiya H, et al. MITO-Porter: A liposome-based carrier system for delivery of macromolecules into mitochondria via membrane fusion. Biochim Biophys Acta. 2008;1778(2):423-432. doi:10.1016/j.bbamem.2007.11.002