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High-resolution interrogation of functional elements in the noncoding genome

Neville E. Sanjana, Jason Wright, Kaijie Zheng, Ophir Shalem, Pierre Fontanillas, Julia Joung, Christine Cheng, Aviv Regev, Feng Zhang
doi: https://doi.org/10.1101/049130
Neville E. Sanjana
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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  • For correspondence: nsanjana@nygenome.org zhang@broadinstitute.org
Jason Wright
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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Kaijie Zheng
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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Ophir Shalem
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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Pierre Fontanillas
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
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Julia Joung
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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Christine Cheng
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
3Howard Hughes Medical Institute, David H. Koch Institute of Integrative Cancer Biology, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA
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Aviv Regev
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
3Howard Hughes Medical Institute, David H. Koch Institute of Integrative Cancer Biology, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA
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Feng Zhang
1Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA 02142, USA
2McGovern Institute for Brain Research, Department of Brain and Cognitive Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA
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  • For correspondence: nsanjana@nygenome.org zhang@broadinstitute.org
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Abstract

The noncoding genome plays a major role in gene regulation and disease yet we lack tools for rapid identification and manipulation of noncoding elements. Here, we develop a large-scale CRISPR screen employing ~18,000 sgRNAs targeting >700 kb of noncoding sequence in an unbiased manner surrounding three genes (NF1, NF2, and CUL3) involved in resistance to the BRAF inhibitor vemurafenib in the BRAF-mutant melanoma cell line A375. We identify specific noncoding locations near genes that modulate drug resistance when mutated. These sites have predictive hallmarks of noncoding function, such as physical interaction with gene promoters, evolutionary conservation and tissue-specific chromatin accessibility. At a subset of identified elements at the CUL3 locus, we show that engineered mutations lead to a loss of gene expression associated with changes in transcription factor occupancy and in long-range and local epigenetic environments, implicating these sites in gene regulation and chemotherapeutic resistance. This demonstration of an unbiased mutagenesis screen across large noncoding regions expands the potential of pooled CRISPR screens for fundamental genomic discovery and for elucidating biologically relevant mechanisms of gene regulation.

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Posted April 18, 2016.
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High-resolution interrogation of functional elements in the noncoding genome
Neville E. Sanjana, Jason Wright, Kaijie Zheng, Ophir Shalem, Pierre Fontanillas, Julia Joung, Christine Cheng, Aviv Regev, Feng Zhang
bioRxiv 049130; doi: https://doi.org/10.1101/049130
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High-resolution interrogation of functional elements in the noncoding genome
Neville E. Sanjana, Jason Wright, Kaijie Zheng, Ophir Shalem, Pierre Fontanillas, Julia Joung, Christine Cheng, Aviv Regev, Feng Zhang
bioRxiv 049130; doi: https://doi.org/10.1101/049130

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