biostudies-literatureGoglia AGNIBIB NIH HHSNCI NIH HHSNational Institutes of HealthNIGMS NIH HHSNational Science Foundation240-253.e6https://www.ebi.ac.uk/biostudies/studies/S-EPMC7540725biostudies-literatureUnknown10(3)Complex, time-varying responses have been observed widely in cell signaling, but how specific dynamics are generated or regulated is largely unknown. One major obstacle has been that high-throughput screens are typically incompatible with the live-cell assays used to monitor dynamics. Here, we address this challenge by screening a library of 429 kinase inhibitors and monitoring extracellular-regulated kinase (Erk) activity over 5 h in more than 80,000 single primary mouse keratinocytes. Our screen reveals both known and uncharacterized modulators of Erk dynamics, including inhibitors of non-epidermal growth factor receptor (EGFR) receptor tyrosine kinases (RTKs) that increase Erk pulse frequency and overall activity. Using drug treatment and direct optogenetic control, we demonstrate that Cell systemsA Live-Cell Screen for Altered Erk Dynamics Reveals Principles of Proliferative Control.PMC7540725DP2 EB024247T32 GM007388F30 CA206408Toettcher JEBasta LPJena SGSilbert JDevenport DGoglia AGWilson MZfalseA Live-Cell Screen for Altered Erk Dynamics Reveals Principles of Proliferative Control.Complex, time-varying responses have been observed widely in cell signaling, but how specific dynamics are generated or regulated is largely unknown. One major obstacle has been that high-throughput screens are typically incompatible with the live-cell assays used to monitor dynamics. Here, we address this challenge by screening a library of 429 kinase inhibitors and monitoring extracellular-regulated kinase (Erk) activity over 5 h in more than 80,000 single primary mouse keratinocytes. Our screen reveals both known and uncharacterized modulators of Erk dynamics, including inhibitors of non-epidermal growth factor receptor (EGFR) receptor tyrosine kinases (RTKs) that increase Erk pulse frequency and overall activity. Using drug treatment and direct optogenetic control, we demonstrate that 2020-01-01T00:00:00Z2020 Mar2025-04-04T01:45:45.012Z2025-04-04T01:45:45.012ZS-EPMC75407253219187410.1016/j.cels.2020.02.005