Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light

Frontiers in Neuroscience
Toshifumi AsanoHiromu Yawo

Abstract

As one of the ubiquitous second messengers, the intracellular Ca2+, has been revealed to be a pivotal regulator of various cellular functions. Two major sources are involved in the initiation of Ca2+-dependent signals: influx from the extracellular space and release from the intracellular Ca2+ stores such as the endoplasmic/sarcoplasmic reticulum (ER/SR). To manipulate the Ca2+ release from the stores under high spatiotemporal precision, we established a new method termed "organelle optogenetics." That is, one of the light-sensitive cation channels (channelrhodopsin-green receiver, ChRGR), which is Ca2+-permeable, was specifically targeted to the ER/SR. The expression specificity as well as the functional operation of the ER/SR-targeted ChRGR (ChRGRER) was evaluated using mouse skeletal myoblasts (C2C12): (1) the ChRGRER co-localized with the ER-marker KDEL; (2) no membrane current was generated by light under whole-cell clamp of cells expressing ChRGRER; (3) an increase of fluorometric Ca2+ was evoked by the optical stimulation (OS) in the cells expressing ChRGRER in a manner independent on the extracellular Ca2+ concentration ([Ca2+]o); (4) the ΔF/F0 was sensitive to the inhibitor of sarco/endoplasmic reticulum Ca2+-ATPase (S...Continue Reading

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Citations

Nov 2, 2019·Traffic·Sunaina SuranaGiampietro Schiavo
Feb 9, 2021·Reviews in the Neurosciences·Alla B SalminaOlga L Vlasova
Feb 23, 2020·Seminars in Cell & Developmental Biology·Olof Idevall-Hagren, Anders Tengholm
Dec 29, 2020·ACS Synthetic Biology·Peiling WuJifeng Yuan

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Methods Mentioned

BETA
PCR
Transfection
confocal microscopy
super-resolution microscopy

Software Mentioned

KDEL
pCLAMP
Venus
ImageJ
Excel

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