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Optochemokine Tandem for Light-Control of Intracellular Ca(2+)

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-g...

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Autores principales: Feldbauer, Katrin, Schlegel, Jan, Weissbecker, Juliane, Sauer, Frank, Wood, Phillip G., Bamberg, Ernst, Terpitz, Ulrich
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5074463/
https://www.ncbi.nlm.nih.gov/pubmed/27768773
http://dx.doi.org/10.1371/journal.pone.0165344
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author Feldbauer, Katrin
Schlegel, Jan
Weissbecker, Juliane
Sauer, Frank
Wood, Phillip G.
Bamberg, Ernst
Terpitz, Ulrich
author_facet Feldbauer, Katrin
Schlegel, Jan
Weissbecker, Juliane
Sauer, Frank
Wood, Phillip G.
Bamberg, Ernst
Terpitz, Ulrich
author_sort Feldbauer, Katrin
collection PubMed
description 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 Ca(2+)-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 Ca(2+) by tandem endosomes into the cytosol via CatCh was visualized using the Ca(2+)-sensitive dyes rhod2 and rhod2-AM showing an increase of intracellular Ca(2+) in response to light.
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spelling pubmed-50744632016-11-04 Optochemokine Tandem for Light-Control of Intracellular Ca(2+) Feldbauer, Katrin Schlegel, Jan Weissbecker, Juliane Sauer, Frank Wood, Phillip G. Bamberg, Ernst Terpitz, Ulrich PLoS One Research Article 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 Ca(2+)-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 Ca(2+) by tandem endosomes into the cytosol via CatCh was visualized using the Ca(2+)-sensitive dyes rhod2 and rhod2-AM showing an increase of intracellular Ca(2+) in response to light. Public Library of Science 2016-10-21 /pmc/articles/PMC5074463/ /pubmed/27768773 http://dx.doi.org/10.1371/journal.pone.0165344 Text en © 2016 Feldbauer et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Feldbauer, Katrin
Schlegel, Jan
Weissbecker, Juliane
Sauer, Frank
Wood, Phillip G.
Bamberg, Ernst
Terpitz, Ulrich
Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title_full Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title_fullStr Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title_full_unstemmed Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title_short Optochemokine Tandem for Light-Control of Intracellular Ca(2+)
title_sort optochemokine tandem for light-control of intracellular ca(2+)
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5074463/
https://www.ncbi.nlm.nih.gov/pubmed/27768773
http://dx.doi.org/10.1371/journal.pone.0165344
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