Synaptotagmin-7 places dense-core vesicles at the cell membrane to promote Munc13-2- and Ca2+-dependent priming

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Abstract

Synaptotagmins confer calcium-dependence to the exocytosis of secretory vesicles, but how coexpressed synaptotagmins interact remains unclear. We find that synaptotagmin-1 and synaptotagmin-7 when present alone act as standalone fast and slow Ca²⁺-sensors for vesicle fusion in mouse chromaffin cells. When present together, synaptotagmin-1 and synaptotagmin-7 are found in largely non-overlapping clusters on dense-core vesicles. Synaptotagmin-7 stimulates Ca²⁺-dependent vesicle priming and inhibits depriming, and it promotes ubMunc13-2- and phorbolester-dependent priming, especially at low resting calcium concentrations. The priming effect of synaptotagmin-7 increases the number of vesicles fusing via synaptotagmin-1, while negatively affecting their fusion speed, indicating both synergistic and competitive interactions between synaptotagmins. Synaptotagmin-7 places vesicles in close membrane apposition (<6 nm); without it, vesicles accumulate out of reach of the fusion complex (20-40 nm). We suggest that a synaptotagmin-7-dependent movement toward the membrane is involved in Munc13-2/phorbolester/Ca²⁺-dependent priming as a prelude to fast and slow exocytosis triggering.

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All data generated or analysed during this study are or will be included in the manuscript and supporting files.

Article and author information

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Bassam Tawfik

Center for Neuroscience, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-1193-8494
Joana S Martins

Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark

Competing interests
No competing interests declared.
Sébastien Houy

Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0003-3639-1931
Cordelia Imig

Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, Göttingen, Germany

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-7351-8706
Paulo S Pinheiro

Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark

Competing interests
No competing interests declared.
Sonja M Wojcik

Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, Göttingen, Germany

Competing interests
No competing interests declared.
Nils Brose

Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, Göttingen, Germany

Competing interests
Nils Brose, Reviewing editor, eLife.
Benjamin H Cooper

Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, Göttingen, Germany

Competing interests
No competing interests declared.
Jakob Balslev Sørensen

Department of Neuroscience, University of Copenhagen, Copenhagen, Denmark

For correspondence
jakobbs@sund.ku.dk

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-5465-3769

Funding

University of Copenhagen 2016 excellence program (KU2016)

Jakob Balslev Sørensen

Novo Nordisk Foundation (NNF19OC0058298)

Jakob Balslev Sørensen

Lundbeckfonden (R221-2016-1202)

Jakob Balslev Sørensen

Independent Research Fund Denmark (0134-00141A)

Jakob Balslev Sørensen

Lundbeckfonden (R34-A3740)

Paulo S Pinheiro

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

Ethics

Animal experimentation: Mice were kept in an AAALAC-accredited stable at the University of Copenhagen operating a 12h/12h light/dark cycle with access to water and food ad libitum. Permission to keep and breed KO mice were obtained from the Danish Animal Experiments Inspectorate (permissions 2006/562-43 and 2018-15-0202-00157).

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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.