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Base editing in human cells with monomeric DddA-TALE fusion deaminases

Author

Listed:
  • Young Geun Mok

    (Institute for Basic Science)

  • Ji Min Lee

    (Institute for Basic Science
    Seoul National University)

  • Eugene Chung

    (Institute for Basic Science
    Seoul National University)

  • Jaesuk Lee

    (Institute for Basic Science
    Seoul National University)

  • Kayeong Lim

    (Institute for Basic Science)

  • Sung-Ik Cho

    (Institute for Basic Science
    Seoul National University)

  • Jin-Soo Kim

    (Institute for Basic Science)

Abstract

Inter-bacterial toxin DddA-derived cytosine base editors (DdCBEs) enable targeted C-to-T conversions in nuclear and organellar DNA. DddAtox, the deaminase catalytic domain derived from Burkholderia cenocepacia, is split into two inactive halves to avoid its cytotoxicity in eukaryotic cells, when fused to transcription activator-like effector (TALE) DNA-binding proteins to make DdCBEs. As a result, DdCBEs function as pairs, which hampers gene delivery via viral vectors with a small cargo size. Here, we present non-toxic, full-length DddAtox variants to make monomeric DdCBEs (mDdCBEs), enabling mitochondrial DNA editing with high efficiencies of up to 50%, when transiently expressed in human cells. We demonstrate that mDdCBEs expressed via AAV in cultured human cells can achieve nearly homoplasmic C-to-T editing in mitochondrial DNA. Interestingly, mDdCBEs often produce mutation patterns different from those obtained with conventional dimeric DdCBEs. Furthermore, mDdCBEs allow base editing at sites for which only one TALE protein can be designed. We also show that transfection of mDdCBE-encoding mRNA, rather than plasmid, can reduce off-target editing in human mitochondrial DNA.

Suggested Citation

  • Young Geun Mok & Ji Min Lee & Eugene Chung & Jaesuk Lee & Kayeong Lim & Sung-Ik Cho & Jin-Soo Kim, 2022. "Base editing in human cells with monomeric DddA-TALE fusion deaminases," Nature Communications, Nature, vol. 13(1), pages 1-10, December.
  • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-31745-y
    DOI: 10.1038/s41467-022-31745-y
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    References listed on IDEAS

    as
    1. Hyunji Lee & Seonghyun Lee & Gayoung Baek & Annie Kim & Beum-Chang Kang & Huiyun Seo & Jin-Soo Kim, 2021. "Mitochondrial DNA editing in mice with DddA-TALE fusion deaminases," Nature Communications, Nature, vol. 12(1), pages 1-6, December.
    2. Kayeong Lim & Sung-Ik Cho & Jin-Soo Kim, 2022. "Nuclear and mitochondrial DNA editing in human cells with zinc finger deaminases," Nature Communications, Nature, vol. 13(1), pages 1-10, December.
    3. Beverly Y. Mok & Marcos H. de Moraes & Jun Zeng & Dustin E. Bosch & Anna V. Kotrys & Aditya Raguram & FoSheng Hsu & Matthew C. Radey & S. Brook Peterson & Vamsi K. Mootha & Joseph D. Mougous & David R, 2020. "A bacterial cytidine deaminase toxin enables CRISPR-free mitochondrial base editing," Nature, Nature, vol. 583(7817), pages 631-637, July.
    4. Alexis C. Komor & Yongjoo B. Kim & Michael S. Packer & John A. Zuris & David R. Liu, 2016. "Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage," Nature, Nature, vol. 533(7603), pages 420-424, May.
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    Cited by:

    1. Friedrich Fauser & Bhakti N. Kadam & Sebastian Arangundy-Franklin & Jessica E. Davis & Vishvesha Vaidya & Nicola J. Schmidt & Garrett Lew & Danny F. Xia & Rakshaa Mureli & Colman Ng & Yuanyue Zhou & N, 2024. "Compact zinc finger architecture utilizing toxin-derived cytidine deaminases for highly efficient base editing in human cells," Nature Communications, Nature, vol. 15(1), pages 1-11, December.

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