A Low Affinity GCaMP3 Variant (GCaMPer) for Imaging the Endoplasmic Reticulum Calcium Store

被引：51

作者：

Henderson, Mark J. ^{[1
]}

Baldwin, Heather A. ^{[1
]}

Werley, Christopher A. ^{[2
]}

Boccardo, Stefano ^{[2
]}

Whitaker, Leslie R. ^{[1
]}

Yan, Xiaokang ^{[1
]}

Holt, Graham T. ^{[6
]}

Schreiter, Eric R. ^{[6
]}

Looger, Loren L. ^{[6
]}

Cohen, Adam E. ^{[2
,3
,4
,5
]}

Kim, Douglas S. ^{[6
]}

Harvey, Brandon K. ^{[1
]}

机构：

[1] NIDA, NIH, Baltimore, MD 21224 USA

[2] Harvard Univ, Dept Chem & Chem Biol, Cambridge, MA 02138 USA

[3] Harvard Univ, Dept Phys, Cambridge, MA 02138 USA

[4] Harvard Univ, Harvard Stem Cell Inst, Cambridge, MA 02138 USA

[5] Harvard Univ, Howard Hughes Med Inst, Cambridge, MA 02138 USA

[6] Howard Hughes Med Inst, Ashburn, VA 20147 USA

来源：

PLOS ONE | 2015年 / 10卷 / 10期

基金：

美国国家卫生研究院;

关键词：

FLUORESCENT PROTEINS; CA2+; RELEASE; EXPRESSION; REGULATOR; INDICATOR; NEURONS; VECTOR; KDEL;

D O I：

10.1371/journal.pone.0139273

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Endoplasmic reticulum calcium homeostasis is critical for cellular functions and is disrupted in diverse pathologies including neurodegeneration and cardiovascular disease. Owing to the high concentration of calcium within the ER, studying this subcellular compartment requires tools that are optimized for these conditions. To develop a single-fluorophore genetically encoded calcium indicator for this organelle, we targeted a low affinity variant of GCaMP3 to the ER lumen (GCaMPer (10.19)). A set of viral vectors was constructed to express GCaMPer in human neuroblastoma cells, rat primary cortical neurons, and human induced pluripotent stem cell-derived cardiomyocytes. We observed dynamic changes in GCaMPer (10.19) fluorescence in response to pharmacologic manipulations of the ER calcium store. Additionally, periodic calcium efflux from the ER was observed during spontaneous beating of cardiomyocytes. GCaMPer (10.19) has utility in imaging ER calcium in living cells and providing insight into luminal calcium dynamics under physiologic and pathologic states.

引用

页数：17

共 47 条

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