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Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage

Author

Listed:
  • Nicole M. Gaudelli

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard)

  • Alexis C. Komor

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard
    University of California, San Diego)

  • Holly A. Rees

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard)

  • Michael S. Packer

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard
    Beam Therapeutics)

  • Ahmed H. Badran

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard)

  • David I. Bryson

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard
    Beam Therapeutics)

  • David R. Liu

    (Harvard University
    Howard Hughes Medical Institute, Harvard University
    Broad Institute of MIT and Harvard)

Abstract

The spontaneous deamination of cytosine is a major source of transitions from C•G to T•A base pairs, which account for half of known pathogenic point mutations in humans. The ability to efficiently convert targeted A•T base pairs to G•C could therefore advance the study and treatment of genetic diseases. The deamination of adenine yields inosine, which is treated as guanine by polymerases, but no enzymes are known to deaminate adenine in DNA. Here we describe adenine base editors (ABEs) that mediate the conversion of A•T to G•C in genomic DNA. We evolved a transfer RNA adenosine deaminase to operate on DNA when fused to a catalytically impaired CRISPR–Cas9 mutant. Extensive directed evolution and protein engineering resulted in seventh-generation ABEs that convert targeted A•T base pairs efficiently to G•C (approximately 50% efficiency in human cells) with high product purity (typically at least 99.9%) and low rates of indels (typically no more than 0.1%). ABEs introduce point mutations more efficiently and cleanly, and with less off-target genome modification, than a current Cas9 nuclease-based method, and can install disease-correcting or disease-suppressing mutations in human cells. Together with previous base editors, ABEs enable the direct, programmable introduction of all four transition mutations without double-stranded DNA cleavage.

Suggested Citation

  • Nicole M. Gaudelli & Alexis C. Komor & Holly A. Rees & Michael S. Packer & Ahmed H. Badran & David I. Bryson & David R. Liu, 2017. "Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage," Nature, Nature, vol. 551(7681), pages 464-471, November.
  • Handle: RePEc:nat:nature:v:551:y:2017:i:7681:d:10.1038_nature24644
    DOI: 10.1038/nature24644
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    Citations

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    Cited by:

    1. Jaesuk Lee & Kayeong Lim & Annie Kim & Young Geun Mok & Eugene Chung & Sung-Ik Cho & Ji Min Lee & Jin-Soo Kim, 2023. "Prime editing with genuine Cas9 nickases minimizes unwanted indels," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    2. Jeonghun Kwon & Minyoung Kim & Seungmin Bae & Anna Jo & Youngho Kim & Jungjoon K. Lee, 2022. "TAPE-seq is a cell-based method for predicting genome-wide off-target effects of prime editor," Nature Communications, Nature, vol. 13(1), pages 1-13, December.
    3. Jiajia Lin & Ming Jin & Dong Yang & Zhifang Li & Yu Zhang & Qingquan Xiao & Yin Wang & Yuyang Yu & Xiumei Zhang & Zhurui Shao & Linyu Shi & Shu Zhang & Wan-jin Chen & Ning Wang & Shiwen Wu & Hui Yang , 2024. "Adenine base editing-mediated exon skipping restores dystrophin in humanized Duchenne mouse model," Nature Communications, Nature, vol. 15(1), pages 1-12, December.
    4. 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.
    5. Chengdong Zhang & Yuan Yang & Tao Qi & Yuening Zhang & Linghui Hou & Jingjing Wei & Jingcheng Yang & Leming Shi & Sang-Ging Ong & Hongyan Wang & Hui Wang & Bo Yu & Yongming Wang, 2023. "Prediction of base editor off-targets by deep learning," Nature Communications, Nature, vol. 14(1), pages 1-11, December.
    6. Lin Zhao & Sabrina R. T. Koseki & Rachel A. Silverstein & Nadia Amrani & Christina Peng & Christian Kramme & Natasha Savic & Martin Pacesa & Tomás C. Rodríguez & Teodora Stan & Emma Tysinger & Lauren , 2023. "PAM-flexible genome editing with an engineered chimeric Cas9," Nature Communications, Nature, vol. 14(1), pages 1-8, December.
    7. Dominique L. Brooks & Manuel J. Carrasco & Ping Qu & William H. Peranteau & Rebecca C. Ahrens-Nicklas & Kiran Musunuru & Mohamad-Gabriel Alameh & Xiao Wang, 2023. "Rapid and definitive treatment of phenylketonuria in variant-humanized mice with corrective editing," Nature Communications, Nature, vol. 14(1), pages 1-9, December.
    8. Daniel Whisenant & Kayeong Lim & Gwladys Revêchon & Haidong Yao & Martin O. Bergo & Piotr Machtel & Jin-Soo Kim & Maria Eriksson, 2022. "Transient expression of an adenine base editor corrects the Hutchinson-Gilford progeria syndrome mutation and improves the skin phenotype in mice," Nature Communications, Nature, vol. 13(1), pages 1-13, December.
    9. Zeyu Lu & Lingtian Zhang & Qing Mu & Junyang Liu & Yu Chen & Haoyuan Wang & Yanjun Zhang & Rui Su & Ruijun Wang & Zhiying Wang & Qi Lv & Zhihong Liu & Jiasen Liu & Yunhua Li & Yanhong Zhao, 2024. "Progress in Research and Prospects for Application of Precision Gene-Editing Technology Based on CRISPR–Cas9 in the Genetic Improvement of Sheep and Goats," Agriculture, MDPI, vol. 14(3), pages 1-17, March.
    10. Qian Wang & Jie Yang & Zhicheng Zhong & Jeffrey A. Vanegas & Xue Gao & Anatoly B. Kolomeisky, 2021. "A general theoretical framework to design base editors with reduced bystander effects," Nature Communications, Nature, vol. 12(1), pages 1-10, December.
    11. Yakun Wang & Shengjia Tang & Naihui Guo & Ruihu An & Zongliang Ren & Shikai Hu & Xiangjin Wei & Guiai Jiao & Lihong Xie & Ling Wang & Ying Chen & Fengli Zhao & Peisong Hu & Zhonghua Sheng & Shaoqing T, 2023. "Base Editing of EUI1 Improves the Elongation of the Uppermost Internode in Two-Line Male Sterile Rice Lines," Agriculture, MDPI, vol. 13(3), pages 1-13, March.
    12. Daphne Collias & Elena Vialetto & Jiaqi Yu & Khoa Co & Éva d. H. Almási & Ann-Sophie Rüttiger & Tatjana Achmedov & Till Strowig & Chase L. Beisel, 2023. "Systematically attenuating DNA targeting enables CRISPR-driven editing in bacteria," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    13. Chao Yang & Zhenzhen Ma & Keshan Wang & Xingxiao Dong & Meiyu Huang & Yaqiu Li & Xiagu Zhu & Ju Li & Zhihui Cheng & Changhao Bi & Xueli Zhang, 2023. "HMGN1 enhances CRISPR-directed dual-function A-to-G and C-to-G base editing," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    14. Luke Hoberecht & Pirunthan Perampalam & Aaron Lun & Jean-Philippe Fortin, 2022. "A comprehensive Bioconductor ecosystem for the design of CRISPR guide RNAs across nucleases and technologies," Nature Communications, Nature, vol. 13(1), pages 1-20, December.
    15. Yidong Wu & Xiaoling Wan & Dongdong Zhao & Xuxu Chen & Yujie Wang & Xinxin Tang & Ju Li & Siwei Li & Xiaodong Sun & Changhao Bi & Xueli Zhang, 2023. "AAV-mediated base-editing therapy ameliorates the disease phenotypes in a mouse model of retinitis pigmentosa," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    16. Daniel C. Volke & Román A. Martino & Ekaterina Kozaeva & Andrea M. Smania & Pablo I. Nikel, 2022. "Modular (de)construction of complex bacterial phenotypes by CRISPR/nCas9-assisted, multiplex cytidine base-editing," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    17. Alice Rovai & BoMee Chung & Qingluan Hu & Sebastian Hook & Qinggong Yuan & Tibor Kempf & Florian Schmidt & Dirk Grimm & Steven R. Talbot & Lars Steinbrück & Jasper Götting & Jens Bohne & Simon A. Kroo, 2022. "In vivo adenine base editing reverts C282Y and improves iron metabolism in hemochromatosis mice," Nature Communications, Nature, vol. 13(1), pages 1-10, December.

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