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PMID: 16902411 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Dissecting docking and tethering of secretory vesicles at the target membrane.

The EMBO journal ·Vol. 25 ·No. 16 ·2006-08-23 ·Pages 3725-37

Toonen RF, Kochubey O, de Wit H, Gulyas-Kovacs A, Konijnenburg B, Sørensen JB, Klingauf J, Verhage M

Abstract

Secretory vesicles dock at their target in preparation for fusion. Using single-vesicle total internal reflection fluorescence microscopy in chromaffin cells, we show that most approaching vesicles dock only transiently, but that some are captured by at least two different tethering modes, weak and strong. Both vesicle delivery and tethering depend on Munc18-1, a known docking factor. By decreasing the amount of cortical actin by Latrunculin A application, morphological docking can be restored artificially in docking-deficient munc18-1 null cells, but neither strong tethering nor fusion, demonstrating that morphological docking is not sufficient for secretion. Deletion of the t-SNARE and Munc18-1 binding partner syntaxin, but not the v-SNARE synaptobrevin/VAMP, also reduces strong tethering and fusion. We conclude that docking vesicles either undock immediately or are captured by minimal tethering machinery and converted in a munc18-1/syntaxin-dependent, strongly tethered, fusion-competent state.

MeSH Terms
Actins/metabolism Animals Cell Membrane/metabolism,ultrastructure Cells, Cultured Chromaffin Cells/physiology,ultrastructure Exocytosis/physiology Gene Dosage/physiology Membrane Fusion/physiology Mice Mice, Knockout Microscopy, Electron, Transmission Munc18 Proteins/genetics,metabolism Qa-SNARE Proteins/genetics,metabolism R-SNARE Proteins/metabolism SNARE Proteins/metabolism Secretory Vesicles/physiology,ultrastructure
Chemicals
Actins Munc18 Proteins Qa-SNARE Proteins R-SNARE Proteins SNARE Proteins Stxbp1 protein, mouse
Authors & Affiliations
8 authors, click to expand affiliations / ORCID
Toonen Ruud F
Department of Functional Genomics, Center for Neurogenomics and Cognitive Research (CNCR), Vrije Universiteit Amsterdam (VUA) and VU Medical Center (VUmc), Amsterdam, The Netherlands.
Kochubey Olexiy
de Wit Heidi
Gulyas-Kovacs Attila
Konijnenburg Bas
Sørensen Jakob B
Klingauf Jurgen
Verhage Matthijs
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
DOI
10.1038/sj.emboj.7601256
Published
2006-08-23
Epub
2006-00-10
Pages
3725-37
Language
English
Region
England
NLM ID
8208664
PMCID
PMC1553188
Subset
IM
Grants
HSRD VA · CDA 10-036 · United States
External Links
PubMed source DOI: 10.1038/sj.emboj.7601256 PMC Full Text View journal details
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