Genetically encoded calcium indicators allow monitoring subcellular
Ca2+ signals inside organelles. Most genetically encoded calcium
indicators are fusions of endogenous calcium-binding proteins
whose functionality in vivo may be perturbed by competition with
cellular partners.We describe here a novel family of fluorescent Ca2+
sensors based on the fusion of two Aequorea victoria proteins, GFP
and apo-aequorin (GAP). GAP exhibited a unique combination
of features: dual-excitation ratiometric imaging, high dynamic
range, good signal-to-noise ratio, insensitivity to pH and Mg2+,
tunable Ca2+ affinity, uncomplicated calibration, and targetability
to five distinct organelles. Moreover, transgenic mice for endoplasmic
reticulum-targeted GAP exhibited a robust long-term
expression that correlated well with its reproducible performance in
various neural tissues. This biosensor fills a gap in the actual repertoire
of Ca2+ indicators for organelles and becomes a valuable tool
for in vivo Ca2+ imaging applications.
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