title: High-resolution interrogation of functional elements in the noncoding genome.
abstract: The noncoding genome affects gene regulation and disease, yet we lack tools for rapid identification and manipulation of noncoding elements. We developed a CRISPR screen using ~18,000 single guide RNAs targeting >700 kilobases surrounding the genes NF1, NF2, and CUL3, which are involved in BRAF inhibitor resistance in melanoma. We find that noncoding locations that modulate drug resistance also harbor predictive hallmarks of noncoding function. With a subset of regions at the CUL3 locus, we demonstrate that engineered mutations alter transcription factor occupancy and long-range and local epigenetic environments, implicating these sites in gene regulation and chemotherapeutic resistance. Through our expansion of the potential of pooled CRISPR screens, we provide tools for genomic discovery and for elucidating biologically relevant mechanisms of gene regulation.
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. nsanjana@nygenome.org zhang@broadinstitute.org.
authors.forename: Neville E
authors.initials: NE
authors.lastname: Sanjana
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Wright
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Kaijie
authors.initials: K
authors.lastname: Zheng
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Ophir
authors.initials: O
authors.lastname: Shalem
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA.
authors.forename: Pierre
authors.initials: P
authors.lastname: Fontanillas
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Julia
authors.initials: J
authors.lastname: Joung
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. Howard Hughes Medical Institute, David H. Koch Institute of Integrative Cancer Biology, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
authors.forename: Christine
authors.initials: C
authors.lastname: Cheng
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. Howard Hughes Medical Institute, David H. Koch Institute of Integrative Cancer Biology, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
authors.forename: Aviv
authors.initials: A
authors.lastname: Regev
authors.affiliation: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA. McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. nsanjana@nygenome.org zhang@broadinstitute.org.
authors.forename: Feng
authors.initials: F
authors.lastname: Zhang
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use; D001426:Bacterial Proteins; D000076987:CRISPR-Associated Protein 9; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D044842:Cullin Proteins / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004720:Endonucleases; D004742:Enhancer Elements, Genetic / Q000235:genetics / Q000502:physiology*; D005786:Gene Expression Regulation; D015972:Gene Expression Regulation, Neoplastic; D016514:Genes, Neurofibromatosis 1; D016515:Genes, Neurofibromatosis 2; D056426:Genetic Loci; D015894:Genome, Human; D023281:Genomics / Q000379:methods*; D006801:Humans; D007211:Indoles / Q000494:pharmacology / Q000627:therapeutic use; D008545:Melanoma / Q000188:drug therapy / Q000235:genetics; D016296:Mutagenesis; D048493:Proto-Oncogene Proteins B-raf / Q000037:antagonists & inhibitors; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics; D013449:Sulfonamides / Q000494:pharmacology / Q000627:therapeutic use; D000077484:Vemurafenib
chemical_list: D000970:Antineoplastic Agents; D001426:Bacterial Proteins; C503645:CUL3 protein, human; D044842:Cullin Proteins; D007211:Indoles; D000094704:RNA, Guide, CRISPR-Cas Systems; D013449:Sulfonamides; D000077484:Vemurafenib; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D000076987:CRISPR-Associated Protein 9; C000606107:Cas9 endonuclease Streptococcus pyogenes; D004720:Endonucleases