RRC ID 59729
Author Farhana I, Hossain MN, Suzuki K, Matsuda T, Nagai T.
Title Genetically Encoded Fluorescence/Bioluminescence Bimodal Indicators for Ca2+ Imaging.
Journal ACS Sens
Abstract Fluorescent and bioluminescent genetically encoded Ca2+ indicators (GECIs) are an indispensable tool for monitoring Ca2+ dynamics in numerous cellular events. Although fluorescent GECIs have a high spatiotemporal resolution, their application is often confined to short-term imaging due to the external illumination that causes phototoxicity and autofluorescence from specimens. Bioluminescent GECIs overcome these pitfalls with enhanced compatibility to optogenetic manipulation and photophysiological processes; however, they are compromised for spatiotemporal resolution. Therefore, there has been a push toward the use of Ca2+ indicators that possess the advantages of both fluorescent and bioluminescent GECI for a wide range of applications. To address this, we developed a high-affinity bimodal GECI, GLICO, using a single fluorescent protein-based GECI combined with a split luciferase. Through this novel design, the fusion protein becomes bimodal and possesses Ca2+ sensing properties similar to those of its fluorescent ancestor and confers bioluminescence-based Ca2+ imaging. GLICO in bioluminescence mode has the highest dynamic range (2200%) of all bioluminescent GECIs. We demonstrated the performance of GLICO in studying cytosolic Ca2+ dynamics in different cultured cells in each mode. With the purpose of Ca2+ imaging in high Ca2+ content organelle, we also created a low-affinity variant, ReBLICO and performed Ca2+ imaging of the ER in both fluorescence and bioluminescence modes. The ability to switch between fluorescence and bioluminescence modes with a single indicator would benefit transgenic applications by presenting an opportunity for a wide range of live Ca2+ imaging in physiological and pathophysiological conditions.
Volume 4(7)
Pages 1825-1834
Published 2019-7-26
DOI 10.1021/acssensors.9b00531
PMID 31276380
MeSH Animals Calcium / analysis* Calcium / metabolism Calmodulin / genetics Calmodulin / isolation & purification Calmodulin / metabolism* Cell Line, Tumor Channelrhodopsins / metabolism Fluorescence Green Fluorescent Proteins / genetics Green Fluorescent Proteins / isolation & purification Green Fluorescent Proteins / metabolism* Humans Neurons / metabolism Optical Imaging Protein Engineering Rats Recombinant Fusion Proteins / genetics Recombinant Fusion Proteins / isolation & purification Recombinant Fusion Proteins / metabolism*
IF 6.944
Times Cited 5
Human and Animal Cells HeLa