1. Cell Biology
  2. Neuroscience
Download icon

Phosphatidylinositol 4,5-bisphosphate optical uncaging potentiates exocytosis

  1. Alexander M Walter  Is a corresponding author
  2. Rainer Müller
  3. Bassam Tawfik
  4. Keimpe DB Wierda
  5. Paulo S Pinheiro
  6. André Nadler
  7. Anthony W McCarthy
  8. Iwona Ziomkiewicz
  9. Martin Kruse
  10. Gregor Reither
  11. Jens Rettig
  12. Martin Lehmann
  13. Volker Haucke
  14. Bertil Hille
  15. Carsten Schultz
  16. Jakob Balslev Sorensen  Is a corresponding author
  1. University of Copenhagen, Denmark
  2. European Molecular Biology Laboratory (EMBL), Germany
  3. Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Germany
  4. University of Washington School of Medicine, United States
  5. Saarland University, Germany
Research Article
  • Cited 11
  • Views 2,336
  • Annotations
Cite this article as: eLife 2017;6:e30203 doi: 10.7554/eLife.30203

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 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 bypasses 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.

Article and author information

Author details

  1. Alexander M Walter

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    For correspondence
    awalter@fmp-berlin.de
    Competing interests
    No competing interests declared.
  2. Rainer Müller

    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3464-494X
  3. Bassam Tawfik

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1193-8494
  4. Keimpe DB Wierda

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8784-9490
  5. Paulo S Pinheiro

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    Competing interests
    No competing interests declared.
  6. André Nadler

    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
    Competing interests
    No competing interests declared.
  7. Anthony W McCarthy

    Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany
    Competing interests
    No competing interests declared.
  8. Iwona Ziomkiewicz

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    Competing interests
    Iwona Ziomkiewicz, Performed experiments as an employee of University of Copenhagen and is now an employee of AstraZeneca; IZ has no financial investments in AstraZeneca.
  9. Martin Kruse

    Department of Physiology and Biophysics, University of Washington School of Medicine, Seattle, United States
    Competing interests
    No competing interests declared.
  10. Gregor Reither

    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
    Competing interests
    No competing interests declared.
  11. Jens Rettig

    Center for Integrative Physiology and Molecular Medicine, Saarland University, Homburg, Germany
    Competing interests
    No competing interests declared.
  12. Martin Lehmann

    Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany
    Competing interests
    No competing interests declared.
  13. Volker Haucke

    Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Berlin, Germany
    Competing interests
    No competing interests declared.
  14. Bertil Hille

    Department of Physiology and Biophysics, University of Washington School of Medicine, Seattle, United States
    Competing interests
    No competing interests declared.
  15. Carsten Schultz

    Cell Biology and Biophysics Unit, European Molecular Biology Laboratory (EMBL), Heidelberg, Germany
    Competing interests
    No competing interests declared.
  16. Jakob Balslev Sorensen

    Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark
    For correspondence
    jakobbs@sund.ku.dk
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5465-3769

Funding

European Union 7th Framework programme (HEALTH-F2-2009-242167)

  • Jakob Balslev Sorensen

The independent Research Fond Denmark

  • Jakob Balslev Sorensen

The Novo Nordisk Foundation

  • Jakob Balslev Sorensen

The Lundbeck Foundation

  • Paulo S Pinheiro
  • Jakob Balslev Sorensen

European Molecular Biology Laboratory

  • Carsten Schultz

Deutsche Forschungsgemeinschaft (Transregio83)

  • Alexander M Walter
  • Carsten Schultz

Deutsche Forschungsgemeinschaft (Emmy Noether Programme)

  • Alexander M Walter

National Institutes of Health (R37NS008174)

  • Bertil Hille

Alexander von Humboldt-Stiftung

  • Martin Kruse

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Permission to keep and breed knockout mice for this study was obtained from The Danish Animal Experiments Inspectorate (2006/562−43, 2012−15−2935−00001). The animals were maintained in an AAALAC-accredited stable in accordance with institutional guidelines as overseen by the Institutional Animal Care and Use Committee (IACUC).

Reviewing Editor

  1. Suzanne R Pfeffer, Stanford University School of Medicine, United States

Publication history

  1. Received: July 6, 2017
  2. Accepted: October 24, 2017
  3. Accepted Manuscript published: October 25, 2017 (version 1)
  4. Version of Record published: December 1, 2017 (version 2)

Copyright

© 2017, Walter et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,336
    Page views
  • 641
    Downloads
  • 11
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)