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The mouth of a dense-core vesicle opens and closes in a concerted action regulated by calcium and amphiphysin

¹ Medical Research Council Laboratory of Molecular Biology, Cambridge CB2 2QH, England, UK
² Laboratori de Neurobiologia, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas, Institut d'Investigació Biomèdica de Bellvitge, Universitat de Barcelona, L'Hospitalet de Llobregat 08907, Spain

Correspondence to Artur Llobet: allobet{at}ub.edu; or Leon Lagnado: ll1{at}mrc-lmb.cam.ac.uk

Secretion of hormones and peptides by neuroendocrine cells occurs through fast and slow modes of vesicle fusion but the mechanics of these processes are not understood. We used interference reflection microscopy to monitor deformations of the membrane surface and found that both modes of fusion involve the tightly coupled dilation and constriction of the vesicle. The rate of opening is calcium dependent and occurs rapidly at concentrations <5 µM. The fast mode of fusion is blocked selectively by a truncation mutant of amphiphysin. Vesicles do not collapse when fusion is triggered by strontium, rather they remain locked open and membrane scission is blocked. In contrast, constriction of the vesicle opening continues when endocytosis is blocked by inhibiting the function of dynamin. Thus, fast and slow modes of fusion involve similar membrane deformations and vesicle closure can be uncoupled from membrane scission. Regulation of these processes by calcium and amphiphysin may provide a mechanism for controlling the release of vesicle contents.

Abbreviations used in this paper: ANP, atrial natriuretic peptide; BAR, Bin-Amphiphysin-Rv; CCD, charge-coupled device; IRM, interference reflection microscopy; NP, nitrophenyl; ROI, region of interest; TIRF, total internal reflection fluorescence.

© 2008 Llobet et al. This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Vesicles roar, and don't collapse

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This article has been cited by other articles:

• Bhat, P., Thorn, P. (2009). Myosin 2 Maintains an Open Exocytic Fusion Pore in Secretory Epithelial Cells. Mol. Biol. Cell 20: 1795-1803 [Abstract] [Full Text]  
• Llobet, A., Wu, M., Lagnado, L. (2008). The mouth of a dense-core vesicle opens and closes in a concerted action regulated by calcium and amphiphysin. JGP 132: i3-i3 [Full Text]  
• Robinson, R., Williams, R. (2008). Vesicles roar, and don't collapse. JCB 182: 818-819 [Full Text]  

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