Dissecting docking and tethering of secretory vesicles at the target membrane

TY - JOUR

T1 - Dissecting docking and tethering of secretory vesicles at the target membrane

AU - Toonen, Ruud F

AU - Kochubey, Olexiy

AU - de Wit, Heidi

AU - Gulyas-Kovacs, Attila

AU - Konijnenburg, Bas

AU - Sørensen, Jakob B

AU - Klingauf, Jurgen

AU - Verhage, Matthijs

N1 - Keywords: Actins; Animals; Cell Membrane; Cells, Cultured; Chromaffin Cells; Exocytosis; Gene Dosage; Membrane Fusion; Mice; Mice, Knockout; Microscopy, Electron, Transmission; Munc18 Proteins; Qa-SNARE Proteins; R-SNARE Proteins; SNARE Proteins; Secretory Vesicles

PY - 2006

Y1 - 2006

N2 - 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.

AB - 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.

U2 - 10.1038/sj.emboj.7601256

DO - 10.1038/sj.emboj.7601256

M3 - Journal article

C2 - 16902411

SN - 0261-4189

VL - 25

SP - 3725

EP - 3737

JO - EMBO Journal

JF - EMBO Journal

IS - 16

ER -