Fig. S1.

Single vesicle fusion experiments. Ca²⁺-triggered fusion experiments using the same SV and PM vesicles preparations as used for the cryo-ET experiments shown in Figs. 2–4. Fusion between single vesicles was observed by using a content-mixing assay as previously described (12, 19). (A) Fusion experiments without and with 2 μM Cpx. (B) Fusion experiments in the presence of 2 μM Cpx and without and with 0.5 μM C1C2BMUN fragment, as described in ref. 12. Note that the experiments shown in B included 0.5% DAG, resulting in a slightly increased Ca²⁺-triggered amplitude (compare B, Top Right vs. A, Bottom Right). After SV-PM vesicle association, vesicle pairs either undergo Ca²⁺-independent fusion or remain associated until fusion is triggered by 500 μM Ca²⁺ addition. The histograms were normalized with respect to the corresponding number of analyzed SV-PM vesicle pairs (Table S6).