An ERK biosensor identifies phosphatases regulating ERK pathway in tissue development and carcinoma cells

Extracellular signal-regulated kinases (ERKs) play essential roles in development and are implicated in numerous diseases. Phosphatases act as key regulators of ERK activation. To obtain a view of the phosphorylation network regulating ERK activity, we describe a binary approach using mammalian cell culture and forward genetic screens in a model organism. We analyzed a spectrum of phosphatases in human cells with ERK-SPARK reporters. Further validation of selected components in Drosophila revealed a functional map of ERK pathway phosphatases. EYA family phosphatases were selected and characterized as novel and critical regulators of ERK signaling in vivo and in mammalian cells. Chemical screens using phosphatase inhibitors showed that pharmacological inhibition of Drosophila Eyes Absent Homologue 2 (EYA2) promotes the progression of ERK-dependent carcinoma. The kinase reporter enabled high-throughput screening of genetic and pharmacological regulators under physiological and pathological condition, an important approach for investigating the regulation of kinase signaling.