title: CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome.
abstract: Large genome-mapping consortia and thousands of genome-wide association studies have identified non-protein-coding elements in the genome as having a central role in various biological processes. However, decoding the functions of the millions of putative regulatory elements discovered in these studies remains challenging. CRISPR-Cas9-based epigenome editing technologies have enabled precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR-Cas9-based epigenomic regulatory element screening (CERES) for improved high-throughput screening of regulatory element activity in the native genomic context. Using dCas9KRABrepressor and dCas9p300activator constructs and lentiviral single guide RNA libraries to target DNase I hypersensitive sites surrounding a gene of interest, we carried out both loss- and gain-of-function screens to identify regulatory elements for the β-globin and HER2 loci in human cells. CERES readily identified known and previously unidentified regulatory elements, some of which were dependent on cell type or direction of perturbation. This technology allows the high-throughput functional annotation of putative regulatory elements in their native chromosomal context.
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Tyler S
authors.initials: TS
authors.lastname: Klann
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Joshua B
authors.initials: JB
authors.lastname: Black
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Malathi
authors.initials: M
authors.lastname: Chellappan
authors.affiliation: Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Alexias
authors.initials: A
authors.lastname: Safi
authors.affiliation: Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Lingyun
authors.initials: L
authors.lastname: Song
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
authors.forename: Isaac B
authors.identifier: 0000-0002-3064-8532
authors.initials: IB
authors.lastname: Hilton
authors.affiliation: Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.|Department of Pediatrics, Division of Medical Genetics, Duke University Medical Center, Durham, North Carolina, USA.
authors.forename: Gregory E
authors.initials: GE
authors.lastname: Crawford
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.|Department of Biostatistics and Bioinformatics, Duke University Medical Center, Durham, North Carolina, USA.
authors.forename: Timothy E
authors.identifier: 0000-0002-7629-061X
authors.initials: TE
authors.lastname: Reddy
authors.affiliation: Department of Biomedical Engineering, Duke University, Durham, North Carolina, USA.|Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.|Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina, USA.
authors.forename: Charles A
authors.initials: CA
authors.lastname: Gersbach
mesh_terms: D064130:CRISPR-Associated Proteins / Q000235:genetics*; D064113:CRISPR-Cas Systems; D002874:Chromosome Mapping / Q000379:methods; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D057890:Epigenomics / Q000379:methods; D000072669:Gene Editing / Q000379:methods*; D015894:Genome, Human / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D050436:Regulatory Elements, Transcriptional / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods*
chemical_list: D064130:CRISPR-Associated Proteins