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Inducible and multiplex gene regulation using CRISPR–Cpf1-based transcription factors

Nature Methods volume 14, pages 11631166 (2017) | Download Citation

Abstract

Targeted and inducible regulation of mammalian gene expression is a broadly important capability. We engineered drug-inducible catalytically inactive Cpf1 nuclease fused to transcriptional activation domains to tune the expression of endogenous genes in human cells. Leveraging the multiplex capability of the Cpf1 platform, we demonstrate both synergistic and combinatorial gene expression in human cells. Our work should enable the development of multiplex gene perturbation library screens for understanding complex cellular phenotypes.

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Acknowledgements

We thank M. Welch and A. Sousa for technical assistance with constructing certain plasmids. B.P.K. acknowledges support from Banting (Natural Sciences and Engineering Research Council of Canada) and Charles A. King Trust Postdoctoral Fellowships. This work was supported by the National Institutes of Health R35 GM118158 (J.K.J.), NIH R01 GM107427 (J.K.J.), R01 DA036858 (J.S.W.) and U01 CA168370 (J.S.W.), the Howard Hughes Medical Institute (J.S.W.), and the Desmond and Ann Heathwood Massachusetts General Hospital Research Scholar Award (J.K.J.).

Author information

Affiliations

  1. Molecular Pathology Unit, Center for Cancer Research, and Center for Computational and Integrative Biology, Massachusetts General Hospital, Charlestown, Massachusetts, USA.

    • Y Esther Tak
    • , Benjamin P Kleinstiver
    • , Jonathan Y Hsu
    • , Joy E Horng
    • , Jingyi Gong
    •  & J Keith Joung
  2. Department of Pathology, Harvard Medical School, Boston, Massachusetts, USA.

    • Y Esther Tak
    • , Benjamin P Kleinstiver
    •  & J Keith Joung
  3. Department of Cellular & Molecular Pharmacology University of California, San Francisco, San Francisco, California, USA.

    • James K Nuñez
    •  & Jonathan S Weissman
  4. Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Jonathan Y Hsu
  5. Howard Hughes Medical Institute, San Francisco, California, USA.

    • Jonathan S Weissman

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Contributions

Y.E.T., B.P.K., J.K.N., J.Y.H., J.S.W., and J.K.J. conceived of and designed experiments. Y.E.T., B.P.K., J.K.N., J.Y.H., J.E.H., and J.G. performed experiments. Y.E.T., B.P.K., J.K.N., J.Y.H., J.S.W., and J.K.J. wrote the manuscript.

Competing interests

B.P.K. is a consultant for Avectas. J.S.W. is a founder and scientific advisory board member of KSQ Therapeutics. J.K.J. has financial interests in Beam Therapeutics, Editas Medicine, Monitor Biotechnologies, Pairwise Plants, Poseida Therapeutics, and Transposagen Biopharmaceuticals. J.K.J.'s interests were reviewed and are managed by Massachusetts General Hospital and Partners HealthCare in accordance with their conflict of interest policies.

Corresponding author

Correspondence to J Keith Joung.

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DOI

https://doi.org/10.1038/nmeth.4483

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