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Inhibition of CRISPR-Cas9 ribonucleoprotein complex assembly by anti-CRISPR AcrIIC2

Author

Listed:
  • Annoj Thavalingam

    (University of Toronto)

  • Zhi Cheng

    (Chinese Academy of Sciences
    University of Chinese Academy of Sciences)

  • Bianca Garcia

    (University of Toronto)

  • Xue Huang

    (Chinese Academy of Sciences
    University of Science and Technology of China)

  • Megha Shah

    (University of Toronto)

  • Wei Sun

    (Chinese Academy of Sciences)

  • Min Wang

    (Chinese Academy of Sciences)

  • Lucas Harrington

    (University of California, Berkeley)

  • Sungwon Hwang

    (University of Toronto)

  • Yurima Hidalgo-Reyes

    (University of Toronto)

  • Erik J. Sontheimer

    (University of Massachusetts Medical School
    University of Massachusetts Medical School)

  • Jennifer Doudna

    (University of California, Berkeley
    Lawrence Berkeley National Laboratory
    University of California
    University of California)

  • Alan R. Davidson

    (University of Toronto
    University of Toronto)

  • Trevor F. Moraes

    (University of Toronto)

  • Yanli Wang

    (Chinese Academy of Sciences
    University of Chinese Academy of Sciences
    Institute of Biophysics, Chinese Academy of Sciences)

  • Karen L. Maxwell

    (University of Toronto)

Abstract

CRISPR-Cas adaptive immune systems function to protect bacteria from invasion by foreign genetic elements. The CRISPR-Cas9 system has been widely adopted as a powerful genome-editing tool, and phage-encoded inhibitors, known as anti-CRISPRs, offer a means of regulating its activity. Here, we report the crystal structures of anti-CRISPR protein AcrIIC2Nme alone and in complex with Nme1Cas9. We demonstrate that AcrIIC2Nme inhibits Cas9 through interactions with the positively charged bridge helix, thereby preventing sgRNA loading. In vivo phage plaque assays and in vitro DNA cleavage assays show that AcrIIC2Nme mediates its activity through a large electronegative surface. This work shows that anti-CRISPR activity can be mediated through the inhibition of Cas9 complex assembly.

Suggested Citation

  • Annoj Thavalingam & Zhi Cheng & Bianca Garcia & Xue Huang & Megha Shah & Wei Sun & Min Wang & Lucas Harrington & Sungwon Hwang & Yurima Hidalgo-Reyes & Erik J. Sontheimer & Jennifer Doudna & Alan R. D, 2019. "Inhibition of CRISPR-Cas9 ribonucleoprotein complex assembly by anti-CRISPR AcrIIC2," Nature Communications, Nature, vol. 10(1), pages 1-11, December.
    • Handle: RePEc:nat:natcom:v:10:y:2019:i:1:d:10.1038_s41467-019-10577-3
    DOI: 10.1038/s41467-019-10577-3
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    Cited by:

    1. Yanan Zhao & Jiaojiao Hu & Shan-Shan Yang & Jing Zhong & Jianping Liu & Shuo Wang & Yuzhuo Jiao & Fang Jiang & Ruiyang Zhai & Bingnan Ren & Hua Cong & Yuwei Zhu & Fengtong Han & Jixian Zhang & Yue Xu , 2022. "A redox switch regulates the assembly and anti-CRISPR activity of AcrIIC1," Nature Communications, Nature, vol. 13(1), pages 1-13, December.
    2. Xieshuting Deng & Wei Sun & Xueyan Li & Jiuyu Wang & Zhi Cheng & Gang Sheng & Yanli Wang, 2024. "An anti-CRISPR that represses its own transcription while blocking Cas9-target DNA binding," Nature Communications, Nature, vol. 15(1), pages 1-13, December.

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