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Optochemokine tandem for light-control of intracellular Ca2+

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Feldbauer,  Katrin
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;

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Weissbecker,  Juliane
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;

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Wood,  Philip G.
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;

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Bamberg,  Ernst
Department of Biophysical Chemistry, Max Planck Institute of Biophysics, Max Planck Society;
Chemical and Pharmaceutical Sciences Department, Johann Wolfgang Goethe University;

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Citation

Feldbauer, K., Schlegel, J., Weissbecker, J., Sauer, F., Wood, P. G., Bamberg, E., et al. (2016). Optochemokine tandem for light-control of intracellular Ca2+. PLoS One, 10(11): e0165344. doi:doi:10.1371/journal.pone.0165344.


Cite as: http://hdl.handle.net/11858/00-001M-0000-002D-1CFB-5
Abstract
An optochemokine tandem was developed to control the release of calcium from endosomes into the cytosol by light and to analyze the internalization kinetics of G-protein coupled receptors (GPCRs) by electrophysiology. A previously constructed rhodopsin tandem was re-engineered to combine the light-gated Ca2+-permeable cation channel Channelrhodopsin-2(L132C), CatCh, with the chemokine receptor CXCR4 in a functional tandem protein tCXCR4/CatCh. The GPCR was used as a shuttle protein to displace CatCh from the plasma membrane into intracellular areas. As shown by patch-clamp measurements and confocal laser scanning microscopy, heterologously expressed tCXCR4/CatCh was internalized via the endocytic SDF1/CXCR4 signaling pathway. The kinetics of internalization could be followed electrophysiologically via the amplitude of the CatCh signal. The light-induced release of Ca2+ by tandem endosomes into the cytosol via CatCh was visualized using the Ca2+-sensitive dyes rhod2 and rhod2-AM showing an increase of intracellular Ca2+ in response to light.