Article | Published:

Epigenome editing by a CRISPR-Cas9-based acetyltransferase activates genes from promoters and enhancers

Nature Biotechnology volume 33, pages 510517 (2015) | Download Citation

Abstract

Technologies that enable targeted manipulation of epigenetic marks could be used to precisely control cell phenotype or interrogate the relationship between the epigenome and transcriptional control. Here we describe a programmable, CRISPR-Cas9-based acetyltransferase consisting of the nuclease-null dCas9 protein fused to the catalytic core of the human acetyltransferase p300. The fusion protein catalyzes acetylation of histone H3 lysine 27 at its target sites, leading to robust transcriptional activation of target genes from promoters and both proximal and distal enhancers. Gene activation by the targeted acetyltransferase was highly specific across the genome. In contrast to previous dCas9-based activators, the acetyltransferase activates genes from enhancer regions and with an individual guide RNA. We also show that the core p300 domain can be fused to other programmable DNA-binding proteins. These results support targeted acetylation as a causal mechanism of transactivation and provide a robust tool for manipulating gene regulation.

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Acknowledgements

P. Perez-Pinera, D. Kocak, D. Ousterout and D. Lim provided assistance with gRNA design, plasmid cloning, PCR primer validation, and/or RNA isolations. The gene encoding the ICAM1-targeted zinc finger protein was provided by C. Barbas, III. This work was supported by a US National Institutes of Health (NIH) grants to G.E.C., C.A.G. and T.E.R. (R01DA036865 and U01HG007900), a NIH Director's New Innovator Award (DP2OD008586) and National Science Foundation (NSF) Faculty Early Career Development (CAREER) Award (CBET-1151035) to C.A.G. and NIH grant P30AR066527.

Author information

Affiliations

  1. Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.

    • Isaac B Hilton
    • , Pratiksha I Thakore
    •  & Charles A Gersbach
  2. Center for Genomic & Computational Biology, Duke University, Durham, North Carolina, USA.

    • Isaac B Hilton
    • , Anthony M D'Ippolito
    • , Christopher M Vockley
    • , Pratiksha I Thakore
    • , Gregory E Crawford
    • , Timothy E Reddy
    •  & Charles A Gersbach
  3. University Program in Genetics and Genomics, Duke University Medical Center, Durham, North Carolina, USA.

    • Anthony M D'Ippolito
  4. Department of Cell Biology, Duke University Medical Center, Durham, North Carolina, USA.

    • Christopher M Vockley
  5. Department of Pediatrics, Division of Medical Genetics, Duke University Medical Center, Durham, North Carolina, USA.

    • Gregory E Crawford
  6. Department of Biostatistics & Bioinformatics, Duke University Medical Center, Durham, North Carolina, USA.

    • Timothy E Reddy
  7. Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina, USA.

    • Charles A Gersbach

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Contributions

I.B.H., A.M.D., C.M.V., P.I.T., G.E.C., T.E.R. and C.A.G. designed experiments. I.B.H., A.M.D. and C.M.V. performed the experiments. I.B.H., A.M.D., C.M.V., P.I.T., G.E.C., T.E.R. and C.A.G. analyzed the data. I.B.H. and C.A.G. wrote the manuscript with contributions by all authors.

Competing interests

C.A.G., I.B.H. and P.I.T. have filed patent applications related to genome engineering technologies. C.A.G. is a scientific advisor to Editas Medicine, a company engaged in therapeutic development of genome engineering technologies.

Corresponding authors

Correspondence to Timothy E Reddy or Charles A Gersbach.

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DOI

https://doi.org/10.1038/nbt.3199

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