Engineering of NEMO as calcium indicators with large dynamics and high sensitivity

Genetically-encoded calcium indicators (GECI) are indispensable tools for real-time monitoring of intracellular calcium signals and cellular activities in living organisms. Current GECIs face the challenge of sub-optimal peak signal-to-baseline-ratio (SBR) with limited resolution for reporting subtle calcium transients. We report herein the development of a suite of calcium sensors, designated NEMO, with fast kinetics and wide dynamic ranges (>100-fold). NEMO indicators report Ca2+transients with peak SBRs ~20-fold larger than the top-of-the-range GCaMP series. NEMO sensors further enable the quantification of absolution calcium concentration with ratiometric or photochromic imaging. Compared to GCaMPs, NEMOs could detect single action potentials in neurons with a peak SBR two times higher and a median peak SBR four times largerin vivo, thereby outperforming most existing state-of-the-art GECIs. Given their high sensitivity and resolution to report intracellular Ca2+signals, NEMO sensors may find broad applications in monitoring neuronal activities and other Ca2+-modulated physiological processes in both mammals and plants.