Fluorescence resonance energy transfer imaging of cell signaling from in vitro to in vivo: basis of biosensor construction, live imaging, and image processing

Abstract

The progress in imaging technology with fluorescent proteins has uncovered a wide range of biological processes in developmental biology. In particular, genetically-encoded biosensors based on the principle of fluorescence resonance energy transfer (FRET) have been used to visualize spatial and temporal dynamics of intracellular signaling in living cells. However, development of sensitive FRET biosensors and their application to developmental biology remain challenging tasks, which has prevented their widespread use in developmental biology. In this review, we first overview general procedures and tips of imaging with FRET biosensors. We then describe recent advances in FRET imaging - namely, the use of optimized backbones for intramolecular FRET biosensors and transposon-mediated gene transfer to generate stable cell lines and transgenic mice expressing FRET biosensors. Finally, we discuss future perspectives of FRET imaging in developmental biology.