Green Fluorescent Protein-Based Chloride Ion Sensors for In Vivo Imaging

Fluorescence quenching-based optical sensors offer a powerful tool for noninvasive monitoring of halide transmembrane fluxes and intracellular distribution. The observation that the native fluorescence of yellow fluorescent protein (YFP) is quenched by many anions led to the development of intracellular halide sensors genetically encoded and thus able to target specific subcellular compartments. Cyan (C)FP-YFP-based sensors have opened the way for ratiometric estimation of intracellular Cl- concentrations. Moreover, the complete loss of fluorescence induced by halide binding via static quenching in specific GFP variants led to the possibility of development of combined chloride-pH sensors. In this chapter, the thermodynamics, binding-pocket structure, and spectroscopic properties of halide-sensing GFP variants are reviewed in terms of the detailed mechanism governing the dependence of fluorescence on local ions. Engineering of GFP-based probes and their advantages in a range of applications will be discussed, with emphasis on the measurement in vivo of chloride ion fluxes, which regulate many physiological functions and cellular parameters such as membrane excitability, cell volume, charge balance, and resting potential.