Fig. 3
From: In vitro fusion of single synaptic and dense core vesicles reproduces key physiological properties
Secretory vesicle docking and calcium-triggered fusion. a Docking of synaptic vesicles (green), dense-core vesicles (red), and insulin vesicles (blue depends on the presence of both target SNARE proteins syntaxin-1a and SNAP-25 in the planar-supported membrane in the absence of calcium (100 µM EDTA)). Syntaxin-1a (183–288) denotes a construct containing the transmembrane domain and the SNARE motif, whereas syntaxin-1a(1-288) is the full-length protein that also contains the regulatory Habc domain. Secretory vesicle docking to planar-supported lipid bilayers containing different combinations of syntaxin-1a1-288, dSNAP-25, complexin-1, and Munc18. A summary of the statistics is shown in Supplementary Table 1. b Single-vesicle fusion events triggered by calcium from a docked state in the presence of syntaxin-1a1-288, dSNAP-25, complexin-1, Munc18, and the C1C2MUN domain of Munc13. Single events for synaptic vesicles, dense-core vesicles, and insulin vesicles are shown. Soluble Alexa647 was included in the buffer containing 100 μM calcium to measure calcium arrival (black trace) to determine delay times of fusion. Orange line is the precise time determined for calcium arrival. The traces represent the release of respective fluorescent content markers of acridine orange for synaptic vesicles (green), NPY-mRuby for dense-core vesicles (red), and C-peptide-GFP (blue) for insulin vesicles. Graphs with expanded times scales around the arrival time of the calcium buffer are shown in Supplementary Fig. 4 and more representative traces are shown in Supplementary Fig. 3. Error bars represent standard errors of the mean
