In vitro fusion of single synaptic and dense core vesicles reproduces key physiological properties

Alex J. B. Kreutzberger, Volker Kiessling, Christopher Stroupe, Binyong Liang, Julia Preobraschenski, Marcelo Ganzella, Mark A. B. Kreutzberger, Robert Nakamoto, Reinhard Jahn, J. David Castle and Lukas K. Tamm ()
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Alex J. B. Kreutzberger: University of Virginia
Volker Kiessling: University of Virginia
Christopher Stroupe: University of Virginia
Binyong Liang: University of Virginia
Julia Preobraschenski: Max Planck Institute for Biophysical Chemistry
Marcelo Ganzella: Max Planck Institute for Biophysical Chemistry
Mark A. B. Kreutzberger: University of Virginia
Robert Nakamoto: University of Virginia
Reinhard Jahn: Max Planck Institute for Biophysical Chemistry
J. David Castle: University of Virginia
Lukas K. Tamm: University of Virginia

Nature Communications, 2019, vol. 10, issue 1, 1-11

Abstract: Abstract Regulated exocytosis of synaptic vesicles is substantially faster than of endocrine dense core vesicles despite similar molecular machineries. The reasons for this difference are unknown and could be due to different regulatory proteins, different spatial arrangements, different vesicle sizes, or other factors. To address these questions, we take a reconstitution approach and compare regulated SNARE-mediated fusion of purified synaptic and dense core chromaffin and insulin vesicles using a single vesicle-supported membrane fusion assay. In all cases, Munc18 and complexin are required to restrict fusion in the absence of calcium. Calcium triggers fusion of all docked vesicles. Munc13 (C1C2MUN domain) is required for synaptic and enhanced insulin vesicle fusion, but not for chromaffin vesicles, correlating inversely with the presence of CAPS protein on purified vesicles. Striking disparities in calcium-triggered fusion rates are observed, increasing with curvature with time constants 0.23 s (synaptic vesicles), 3.3 s (chromaffin vesicles), and 9.1 s (insulin vesicles) and correlating with rate differences in cells.

Date: 2019
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DOI: 10.1038/s41467-019-11873-8

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