Dual observation of the ATP-evoked small GTPase activation and Ca²⁺ transient in astrocytes using a dark red fluorescent protein.

ABSTRACT: Intracellular signal transduction involves a number of biochemical reactions, which largely consist of protein-protein interactions and protein conformational changes. Monitoring Förster resonance energy transfer (FRET) by fluorescence lifetime imaging microscopy (FLIM), called FLIM-FRET, is one of the best ways to visualize such protein dynamics. Here, we attempted to apply dark red fluorescent proteins with significantly smaller quantum yields. Application of the dark mCherry mutants to single-molecule FRET sensors revealed that these dark mCherry mutants are a good acceptor in a pair with mRuby2. Because the FRET measurement between mRuby2 and dark mCherry requires only the red region of wavelengths, it facilitates dual observation with other signaling sensors such as genetically encode

SUBMITTER: Nakahata Y 

PROVIDER: S-EPMC5177924 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature