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CRISPR/Cas9-mediated base editors and their prospects for mitochondrial genome engineering

Gene Therapy (2024)Cite this article

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Abstract

Base editors are a type of double-stranded break (DSB)-free gene editing technology that has opened up new possibilities for precise manipulation of mitochondrial DNA (mtDNA). This includes cytosine and adenosine base editors and more recently guanosine base editors. Because of having low off-target and indel rates, there is a growing interest in developing and evolving this research field. Here, we provide a detailed update on DNA base editors. While base editing has widely been used for nuclear genome engineering, the growing interest in applying this technology to mitochondrial DNA has been faced with several challenges. While Cas9 protein has been shown to enter mitochondria, use of smaller Cas proteins, such as Cas12a, has higher import efficiency. However, sgRNA transfer into mitochondria is the most challenging step. sgRNA structure and ratio of Cas protein to sgRNA are both important factors for efficient sgRNA entry into mitochondria. In conclusion, while there are still several challenges to be addressed, ongoing research in this field holds the potential for new treatments and therapies for mitochondrial disorders.

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Fig. 1: Schematic presentation of base editors.
Fig. 2: Mitochondrial genome engineering in zygotes.

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Funding

This project was partly funded by the grant number: Ref 3.4 - IRN - 1191261 - GF-E from the Alexander von Humboldt Foundation, Germany. The authors confirm that their research is supported by an institution that is primarily involved in education or research.

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Authors and Affiliations

  1. School of BioSciences, The University of Melbourne, Parkville, VIC, Australia

    Shahin Eghbalsaied, Clancy Lawler & Stephen Frankenberg

  2. Department of Animal Science, Isfahan Branch, Islamic Azad University (IAU), Isfahan, Iran

    Shahin Eghbalsaied

  3. Department of Cardiology and Pneumology, University Medical Center Göttingen, Göttingen, Germany

    Shahin Eghbalsaied

  4. Department of Biotechnology, Institute of Farm Animal Genetics, Friedrich-Loeffler-Institute (FLI), Mariensee, Germany

    Björn Petersen

  5. eGenesis, 2706 HWY E, 53572, Mount Horeb, WI, USA

    Björn Petersen

  6. Department of Genome Engineering, NovoHelix, Miami, FL, USA

    Raul A. Hajiyev & Steve R. Bischoff

  7. Department of Computer Science, Kent State University, Kent, OH, USA

    Raul A. Hajiyev

  8. Foundry for Genome Engineering & Reproductive Medicine (FGERM), Miami, FL, USA

    Steve R. Bischoff

Authors
  1. Shahin Eghbalsaied
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  2. Clancy Lawler
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  3. Björn Petersen
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  4. Raul A. Hajiyev
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  5. Steve R. Bischoff
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  6. Stephen Frankenberg
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Contributions

Conceptualization and writing the original draft: SE. Manuscript content enrichment, overview, and editing: CL, SRB, SF, and BP. Tables, Figures, and graphical design: RAH, SRB, and SE.

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Correspondence to Shahin Eghbalsaied or Stephen Frankenberg.

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Eghbalsaied, S., Lawler, C., Petersen, B. et al. CRISPR/Cas9-mediated base editors and their prospects for mitochondrial genome engineering. Gene Ther (2024). https://doi.org/10.1038/s41434-023-00434-w

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  • DOI: https://doi.org/10.1038/s41434-023-00434-w

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