Syt1 promotes the formation of migrasome-like structures in vitro. (A) Recombinant Syt1-GFP was analyzed on an SDS-PAGE gel and stained with Coomassie Brilliant Blue. (B) Control GUVs and GUVs embedded with Syt1-GFP treated with or without Ca²⁺ were subjected to the in vitro reconstitution assay. Green signal, Syt1-GFP. Magenta signal, Rhodamine-PE. Scale bar, 2 µm. (C) Quantification of the number of migrasome-like structures per 100 µm of the tether from images in B. Data shown represent the mean ± SEM; P values were calculated using a two-tailed, unpaired t test. n = 102 (no Syt1, no Ca²⁺), 108 (no Syt1, with Ca²⁺), 172 (with Syt1, no Ca²⁺), and 213 (with Syt1, with Ca²⁺) tethers. Quantifications are pooled from three independent experiments. Each biological replicate is color-coded: the statistical data from one experimental run is red, another independent experiment is represented by gray, and a third experiment is shown as blue. The dots, squares, triangles, and inverted triangles represent the four groups respectively. The bordered shapes represent those three means. (D) Image of Syt1-containing GUVs in vitro reconstitution assay. The mean intensity was generated using ImageJ. Scale bar, 2 µm. (E) Mean fluorescence intensity of Syt1 along the white lines in D. Source data are available for this figure: SourceData F2.