Figure 5. Munc13-1 C1C2BMUNC2C can induce Ca2+-independent fusion of V- and T-liposomes in the absence of NSF-αSNAP.
Lipid mixing (A,C) between V- and T-liposomes was measured from the fluorescence de-quenching of Marina Blue-labeled lipids and content mixing (B,D) was monitored from the development of FRET between PhycoE-Biotin trapped in the T-liposomes and Cy5-Streptavidin trapped in the V-liposomes. The assays were performed in the presence of different combinations of Munc18-1 (M18), Syt1 C2AB fragment and Munc13-1 C1C2BMUN or C1C2BMUNC2C as indicated. Experiments were started in the presence of 100 μM EGTA and 5 μM streptavidin, and Ca2+ (600 μM) was added after 300 s.
Figure 5—figure supplement 1. Quantification of lipid and content mixing experiments of Figure 5.
Panels (A–D) correspond to panels (A–D) of Figure 5, respectively. Bars represent averages of the normalized fluorescence observed after 500 s (200 s after Ca2+ addition) in experiments performed at least in triplicate. Error bars represent standard deviations.