Genetically Encoded Fluorescence/Bioluminescence Bimodal Indicators for Ca²⁺ Imaging

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Genetically encoded calcium indicators (GECIs) are crucial for monitoring cellular calcium dynamics.
• Fluorescent GECIs offer high spatiotemporal resolution but are limited by phototoxicity and autofluorescence.
• Bioluminescent GECIs avoid phototoxicity but have lower spatiotemporal resolution.

Purpose of the Study:

• To develop a bimodal GECI combining the advantages of fluorescent and bioluminescent indicators.
• To create a novel indicator with high dynamic range for calcium imaging.
• To enable flexible calcium imaging in various cellular compartments and conditions.

Main Methods:

• Development of GLICO, a fusion protein combining a fluorescent GECI with a split luciferase.
• Characterization of GLICO's Ca2+ sensing properties in both fluorescence and bioluminescence modes.
• Application of GLICO and its low-affinity variant (ReBLICO) for cytosolic and ER calcium imaging.

Main Results:

• GLICO exhibits bimodal Ca2+ sensing capabilities, retaining fluorescence properties and enabling bioluminescence imaging.
• GLICO demonstrates the highest dynamic range (2200%) among known bioluminescent GECIs.
• GLICO and ReBLICO successfully imaged cytosolic and ER calcium dynamics in cultured cells.

Conclusions:

• GLICO represents a significant advancement in GECI technology, offering a versatile tool for live Ca2+ imaging.
• The ability to switch between fluorescence and bioluminescence modes enhances applicability in transgenic and physiological studies.
• This bimodal approach facilitates diverse live Ca2+ imaging applications under various physiological and pathophysiological conditions.