Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis

Alexander M Walter; Rainer Müller; Bassam Tawfik; Keimpe Db Wierda; Paulo S Pinheiro; André Nadler; Anthony W McCarthy; Iwona Ziomkiewicz; Martin Kruse; Gregor Reither; Jens Rettig; Martin Lehmann; Volker Haucke; Bertil Hille; Carsten Schultz; Jakob Balslev Sorensen

doi:10.7554/eLife.30203

Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis

Alexander M Walter, Rainer Müller, Bassam Tawfik, Keimpe Db Wierda, Paulo S Pinheiro, André Nadler, Anthony W McCarthy, Iwona Ziomkiewicz, Martin Kruse, Gregor Reither, Jens Rettig, Martin Lehmann, Volker Haucke, Bertil Hille, Carsten Schultz, Jakob Balslev Sorensen

Neuronal Signalling

18 Citations (Scopus)

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Abstract

Phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] is essential for exocytosis. Classical ways of manipulating PI(4,5)P2 levels are slower than its metabolism, making it difficult to distinguish effects of PI(4,5)P2 from those of its metabolites. We developed a membrane-permeant, photoactivatable PI(4,5)P2, which is loaded into cells in an inactive form and activated by light, allowing sub-second increases in PI(4,5)P2 levels. By combining this compound with electrophysiological measurements in mouse adrenal chromaffin cells, we show that PI(4,5)P2 uncaging potentiates exocytosis and identify synaptotagmin-1 (the Ca2+ sensor for exocytosis) and Munc13-2 (a vesicle priming protein) as the relevant effector proteins. PI(4,5)P2 activation of exocytosis did not depend on the PI(4,5)P2-binding CAPS-proteins, suggesting that PI(4,5)P2 uncaging may bypass CAPS-function. Finally, PI(4,5)P2 uncaging triggered the rapid fusion of a subset of readily-releasable vesicles, revealing a rapid role of PI(4,5)P2 in fusion triggering. Thus, optical uncaging of signaling lipids can uncover their rapid effects on cellular processes and identify lipid effectors.

Original language	English
Article number	e30203
Journal	eLife
Volume	6
Number of pages	41
ISSN	2050-084X
DOIs	https://doi.org/10.7554/eLife.30203
Publication status	Published - 25 Oct 2017

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10.7554/eLife.30203Licence: CC BY

Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosisFinal published version, 2.61 MBLicence: CC BY

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title = "Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis",

abstract = "Phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] is essential for exocytosis. Classical ways of manipulating PI(4,5)P2 levels are slower than its metabolism, making it difficult to distinguish effects of PI(4,5)P2 from those of its metabolites. We developed a membrane-permeant, photoactivatable PI(4,5)P2, which is loaded into cells in an inactive form and activated by light, allowing sub-second increases in PI(4,5)P2 levels. By combining this compound with electrophysiological measurements in mouse adrenal chromaffin cells, we show that PI(4,5)P2 uncaging potentiates exocytosis and identify synaptotagmin-1 (the Ca2+ sensor for exocytosis) and Munc13-2 (a vesicle priming protein) as the relevant effector proteins. PI(4,5)P2 activation of exocytosis did not depend on the PI(4,5)P2-binding CAPS-proteins, suggesting that PI(4,5)P2 uncaging may bypass CAPS-function. Finally, PI(4,5)P2 uncaging triggered the rapid fusion of a subset of readily-releasable vesicles, revealing a rapid role of PI(4,5)P2 in fusion triggering. Thus, optical uncaging of signaling lipids can uncover their rapid effects on cellular processes and identify lipid effectors.",

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AU - Wierda, Keimpe Db

AU - Pinheiro, Paulo S

AU - Nadler, André

AU - McCarthy, Anthony W

AU - Ziomkiewicz, Iwona

AU - Kruse, Martin

AU - Reither, Gregor

AU - Rettig, Jens

AU - Lehmann, Martin

AU - Haucke, Volker

AU - Hille, Bertil

AU - Schultz, Carsten

AU - Sorensen, Jakob Balslev

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AB - Phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2] is essential for exocytosis. Classical ways of manipulating PI(4,5)P2 levels are slower than its metabolism, making it difficult to distinguish effects of PI(4,5)P2 from those of its metabolites. We developed a membrane-permeant, photoactivatable PI(4,5)P2, which is loaded into cells in an inactive form and activated by light, allowing sub-second increases in PI(4,5)P2 levels. By combining this compound with electrophysiological measurements in mouse adrenal chromaffin cells, we show that PI(4,5)P2 uncaging potentiates exocytosis and identify synaptotagmin-1 (the Ca2+ sensor for exocytosis) and Munc13-2 (a vesicle priming protein) as the relevant effector proteins. PI(4,5)P2 activation of exocytosis did not depend on the PI(4,5)P2-binding CAPS-proteins, suggesting that PI(4,5)P2 uncaging may bypass CAPS-function. Finally, PI(4,5)P2 uncaging triggered the rapid fusion of a subset of readily-releasable vesicles, revealing a rapid role of PI(4,5)P2 in fusion triggering. Thus, optical uncaging of signaling lipids can uncover their rapid effects on cellular processes and identify lipid effectors.

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Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis

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