Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis

eLife

Jens Rettig¹  Rainer Müller²  Iwona Ziomkiewicz³  Anthony W McCarthy⁴  Volker Haucke⁴  Bassam Tawfik⁴  Bertil Hille⁵  Martin Lehmann⁶  Jakob Balslev Sørensen⁷  Carsten Schultz⁷  Martin Kruse⁷  Gregor Reither⁸  Keimpe DB Wierda⁸  Alexander M Walter⁸  Paulo S Pinheiro⁸  André Nadler⁸ 

[1] Discovery Sciences, AstraZeneca, Cambridge, United Kingdom;Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Berlin, Germany;Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany;Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Heidelberg, Germany;Cellular Neurophysiology, Center for Integrative Physiology and Molecular Medicine, Saarland University, Homburg, Germany;Department of Physiology and Biophysics, School of Medicine, University of Washington, Seattle, United States;Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Berlin, Germany;Neurosecretion group, Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark;

关键词: phosphatidylinositols;    exocytosis;    adrenal chromaffin cell;    synaptotagmin;    Munc13;    optical uncaging;   

DOI  :  10.7554/eLife.30203

来源: DOAJ

【 摘 要 】

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.

【 授权许可】

Unknown