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RNA-guided transcriptional silencing in vivo with S. aureus CRISPR-Cas9 repressors

Author

Listed:
  • Pratiksha I. Thakore

    (Duke University
    Duke University)

  • Jennifer B. Kwon

    (Duke University
    Duke University Medical Center)

  • Christopher E. Nelson

    (Duke University
    Duke University)

  • Douglas C. Rouse

    (Duke University School of Medicine)

  • Matthew P. Gemberling

    (Duke University
    Duke University)

  • Matthew L. Oliver

    (Duke University)

  • Charles A. Gersbach

    (Duke University
    Duke University
    Duke University Medical Center)

Abstract

CRISPR-Cas9 transcriptional repressors have emerged as robust tools for disrupting gene regulation in vitro but have not yet been adapted for systemic delivery in adult animal models. Here we describe a Staphylococcus aureus Cas9-based repressor (dSaCas9KRAB) compatible with adeno-associated viral (AAV) delivery. To evaluate dSaCas9KRAB efficacy for gene silencing in vivo, we silenced transcription of Pcsk9, a regulator of cholesterol levels, in the liver of adult mice. Systemic administration of a dual-vector AAV8 system expressing dSaCas9KRAB and a Pcsk9-targeting guide RNA (gRNA) results in significant reductions of serum Pcsk9 and cholesterol levels. Despite a moderate host response to dSaCas9KRAB expression, Pcsk9 repression is maintained for 24 weeks after a single treatment, demonstrating the potential for long-term gene silencing in post-mitotic tissues with dSaCas9KRAB. In vivo programmable gene silencing enables studies that link gene regulation to complex phenotypes and expands the CRISPR-Cas9 perturbation toolbox for basic research and gene therapy applications.

Suggested Citation

  • Pratiksha I. Thakore & Jennifer B. Kwon & Christopher E. Nelson & Douglas C. Rouse & Matthew P. Gemberling & Matthew L. Oliver & Charles A. Gersbach, 2018. "RNA-guided transcriptional silencing in vivo with S. aureus CRISPR-Cas9 repressors," Nature Communications, Nature, vol. 9(1), pages 1-9, December.
    • Handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-018-04048-4
    DOI: 10.1038/s41467-018-04048-4
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    Cited by:

    1. Chady H. Hakim & Sandeep R. P. Kumar & Dennis O. Pérez-López & Nalinda B. Wasala & Dong Zhang & Yongping Yue & James Teixeira & Xiufang Pan & Keqing Zhang & Emily D. Million & Christopher E. Nelson & , 2021. "Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models," Nature Communications, Nature, vol. 12(1), pages 1-12, December.
    2. Boris Kantor & Bernadette O’Donovan & Joseph Rittiner & Dellila Hodgson & Nicholas Lindner & Sophia Guerrero & Wendy Dong & Austin Zhang & Ornit Chiba-Falek, 2024. "The therapeutic implications of all-in-one AAV-delivered epigenome-editing platform in neurodegenerative disorders," Nature Communications, Nature, vol. 15(1), pages 1-15, December.

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