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Nat Biotechnol. 2018 Feb;36(2):170-178. doi: 10.1038/nbt.4062. Epub 2018 Jan 15.

Dual gene activation and knockout screen reveals directional dependencies in genetic networks.

Author information

1
Department of Microbiology and Immunology, University of California San Francisco Diabetes Center, WM Keck Center for Noncoding RNAs, University of California, San Francisco, San Francisco, California, USA.
2
Institute for Neurodegenerative Diseases, Department of Biochemistry and Biophysics, University of California, San Francisco and Chan Zuckerberg Biohub, San Francisco, California, USA.
3
Helen Diller Family Comprehensive Cancer Center, Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, California, USA.
4
Department of Pharmacology and Emory Chemical Biology Discovery Center, Emory University School of Medicine, Atlanta, Georgia, USA.
5
Department of Medicine, Lung Biology Center, University of California, San Francisco, San Francisco, California, USA.
6
Centre de Bioinformatique, Biostatistique et Biologie Intégrative (C3BI, USR 3756 Institut Pasteur et CNRS), Paris, France.

Abstract

Understanding the direction of information flow is essential for characterizing how genetic networks affect phenotypes. However, methods to find genetic interactions largely fail to reveal directional dependencies. We combine two orthogonal Cas9 proteins from Streptococcus pyogenes and Staphylococcus aureus to carry out a dual screen in which one gene is activated while a second gene is deleted in the same cell. We analyze the quantitative effects of activation and knockout to calculate genetic interaction and directionality scores for each gene pair. Based on the results from over 100,000 perturbed gene pairs, we reconstruct a directional dependency network for human K562 leukemia cells and demonstrate how our approach allows the determination of directionality in activating genetic interactions. Our interaction network connects previously uncharacterized genes to well-studied pathways and identifies targets relevant for therapeutic intervention.

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