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Programmable A-to-Y base editing by fusing an adenine base editor with an N-methylpurine DNA glycosylase

Nature Biotechnology volume 41, pages 1080–1084 (2023)Cite this article

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Abstract

Here we developed an adenine transversion base editor, AYBE, for A-to-C and A-to-T transversion editing in mammalian cells by fusing an adenine base editor (ABE) with hypoxanthine excision protein N-methylpurine DNA glycosylase (MPG). We also engineered AYBE variants enabling targeted editing at genomic loci with higher transversion editing activity (up to 72% for A-to-C or A-to-T editing).

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Fig. 1: Engineering and optimization of AYBE.
Fig. 2: Characterization of editing profiles for AYBE via high-throughput target sequencing.

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Data availability

Expression plasmids used in this study have been deposited at Addgene and will be available at https://www.addgene.org/Huawei_Tong/ (Addgene plasmid nos. 193966–193968). All data supporting the findings of this study are available in the paper (and in its Supplementary Information files). Targeted amplicon sequencing data have been deposited at the Sequence Read Archive and can be accessed at https://www.ncbi.nlm.nih.gov/bioproject/PRJNA874457 (ref. 21). All relevant original data are available from the corresponding authors upon reasonable request.

Code availability

Custom scripts for CRISPResso analyses supporting the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgements

We acknowledge technical support from the FACS facility of HuiGene Therapeutics Co., Ltd. This work was supported by HuiGene Therapeutics Co., Ltd. (H.T.).

Author information

Author notes

  1. These authors contributed equally: Huawei Tong, Xuchen Wang, Yuanhua Liu, Nana Liu.

Authors and Affiliations

  1. HuiGene Therapeutics Co., Ltd., Shanghai, China

    Huawei Tong, Nana Liu, Yun Li, Jiamin Luo, Qian Ma, Danni Wu, Jiyong Li & Hui Yang

  2. Institute of Neuroscience, State Key Laboratory of Neuroscience, Key Laboratory of Primate Neurobiology, Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China

    Xuchen Wang, Yuanhua Liu & Hui Yang

  3. College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China

    Xuchen Wang

  4. Lingang Laboratory, Shanghai, China

    Chunlong Xu

  5. Shanghai Research Center for Brain Science and Brain-Inspired Intelligence, Shanghai, China

    Hui Yang

  6. HuiEdit Therapeutics Co., Ltd., Shanghai, China

    Hui Yang

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  1. Huawei Tong
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Contributions

H.Y. and H.T. jointly conceived the project. H.T. designed and conducted experiments. X.W. and N.L. performed experiments. Y.L. performed target sequencing data analysis. Y.L. and J.L assisted with cell experiments. Q.M., D.W. and J.L. participated in vector construction and FACS. H.Y., H.T. and C.X. supervised the whole project. H.Y., C.X. and H.T. wrote the manuscript draft, and all authors contributed to the editing of the manuscript.

Corresponding authors

Correspondence to Huawei Tong, Chunlong Xu or Hui Yang.

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Competing interests

H.T. discloses a patent application related to the proteins described in this manuscript. H.T., N.L., Y.L., J.L, Q.M., D.W. and J.L. are employees of HuiGene Therapeutics Co., Ltd. H.Y. is a founder of HuiGene Therapeutics Co., Ltd. and HuiEdit Therapeutics Co., Ltd. The remaining authors declare no competing interests.

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Nature Biotechnology thanks the anonymous reviewers for their contribution to the peer review of this work.

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Supplementary information

Supplementary Information

Supplementary Figs. 1–16 and Supplementary Tables 1–4

Reporting Summary

Supplementary Table 5

gRNA spacers, barcoded primers and target sequences used in this study

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Tong, H., Wang, X., Liu, Y. et al. Programmable A-to-Y base editing by fusing an adenine base editor with an N-methylpurine DNA glycosylase. Nat Biotechnol 41, 1080–1084 (2023). https://doi.org/10.1038/s41587-022-01595-6

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