Abstract
AbstractCell-based transcriptional reporters are invaluable in high-throughput compound and CRISPR screens for identifying compounds or genes that can impact a pathway of interest. However, many transcriptional reporters have weak activities and transient responses. This can result in overlooking therapeutic targets and compounds that are difficult to detect, necessitating the resource-consuming process of running multiple screens at various timepoints. Here, we present RADAR, a digitizer circuit for amplifying reporter activity and retaining memory of pathway activation. Reporting on the AP-1 pathway, our circuit identifies compounds with known activity against PKC-related pathways and shows an enhanced dynamic range with improved sensitivity compared to a classical reporter in compound screens. In the first genome-wide pooled CRISPR screen for the AP-1 pathway, RADAR identifies canonical genes from the MAPK and PKC pathways, as well as non-canonical regulators. Thus, our scalable system highlights the benefit and versatility of using genetic circuits in large-scale cell-based screening.
Funder
U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences
U.S. Department of Health & Human Services | NIH | National Institute of Biomedical Imaging and Bioengineering
U.S. Department of Health & Human Services | NIH | NIH Office of the Director
NSF | Directorate for Computer & Information Science & Engineering | Division of Computing and Communication Foundations
National Science Foundation
U.S. Department of Health & Human Services | NIH | National Heart, Lung, and Blood Institute
Publisher
Springer Science and Business Media LLC
Subject
General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry
Cited by
3 articles.
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