A live-cell screen for altered Erk dynamics reveals principles of proliferative control

Abstract

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 for identifying pathway components are typically incompatible with the live-cell assays used to monitor dynamics. Here, we address this challenge by performing a drug screen for altered Erk signaling dynamics in primary mouse keratinocytes. We screened a library of 429 kinase inhibitors, monitoring Erk activity over 5 h in more than 80,000 single live cells. The screen revealed both known and uncharacterized modulators of Erk dynamics, including inhibitors of non-EGFR receptor tyrosine kinases (RTKs) that increased Erk pulse frequency and overall activity. Using drug treatment and direct optogenetic control, we demonstrate that drug-induced changes to Erk dynamics alter the conditions under which cells proliferate. Our work opens the door to highthroughput screens using live-cell biosensors and reveals that cell proliferation integrates information from Erk dynamics as well as additional permissive cues. Ras | Erk | MAP kinase | cell signaling | dynamics | drug screen

Resonance energy transfer sensitises and monitors in situ switching of LOV2-based optogenetic actuators

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Live-imaging of endothelial Erk activity reveals dynamic and sequential signalling events during regenerative angiogenesis

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