Visualizing Dynamics of Cell Signaling In Vivo with a Phase Separation-Based Kinase Reporter

Mol Cell. 2018 Jan 18;69(2):334-346.e4. doi: 10.1016/j.molcel.2017.12.008. Epub 2018 Jan 4.


Visualizing dynamics of kinase activity in living animals is essential for mechanistic understanding of cell and developmental biology. We describe GFP-based kinase reporters that phase-separate upon kinase activation via multivalent protein-protein interactions, forming intensively fluorescent droplets. Called SPARK (separation of phases-based activity reporter of kinase), these reporters have large dynamic range (fluorescence change), high brightness, fast kinetics, and are reversible. The SPARK-based protein kinase A (PKA) reporter reveals oscillatory dynamics of PKA activities upon G protein-coupled receptor activation. The SPARK-based extracellular signal-regulated kinase (ERK) reporter unveils transient dynamics of ERK activity during tracheal metamorphosis in live Drosophila. Because of intensive brightness and simple signal pattern, SPARKs allow easy examination of kinase signaling in living animals in a qualitative way. The modular design of SPARK will facilitate development of reporters of other kinases.

Keywords: Drosophila; ERK; G protein coupled receptor; GFP; kinase; kinase reporter; phase separation; protein kinase A; protein-protein interaction; receptor tyrosine kinase.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Drosophila
  • Enzyme Activation
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Green Fluorescent Proteins / metabolism
  • Humans
  • MAP Kinase Signaling System / physiology
  • Optical Imaging / methods*
  • Phosphorylation
  • Phosphotransferases / metabolism
  • Phosphotransferases / physiology*
  • Signal Transduction / physiology*


  • Green Fluorescent Proteins
  • Phosphotransferases
  • Cyclic AMP-Dependent Protein Kinases
  • Extracellular Signal-Regulated MAP Kinases