Extended Data Fig. 7: Membrane repair negatively regulated PFO translocation to the TGN.

a, Schematic of Ca²⁺ influx-driven membrane repair. Left: in Ca²⁺-containing medium, PFO induces Ca²⁺ influx, which leads to membrane repair. Right: in Ca²⁺-free medium, PFO cannot induce Ca²⁺ influx, and thus is unable to trigger membrane repair. b,c, Representative fluorescence and phase contrast images (b) and quantification of propidium iodide-positive cells (c) in HeLa cells incubated with rPFO in Ca²⁺-containing medium or Ca²⁺-free medium supplemented with 50 μg/mL propidium iodide for 80 min. Scale bar, 25 μm. c, The percentage of cells with propidium iodide signal was quantified from 100 cells (n = 3; mean ± s.d.; two-sided t-test; NS, not significant). Representative from 3 independent experiments. d, Representative immunofluorescence images (left) and quantification of rPFO colocalization with the TGN (right) in HeLa cells stably expressing scFv-sfGFP incubated with 10xGCN4-rPFO in Ca²⁺-containing medium or Ca²⁺-free medium for 80 min. Areas of rPFO foci colocalizing with the TGN were measured with ImageJ (n = 40 cells per condition; N.D., not detectable). Scale bar, 5 μm. Representative from 3 independent experiments.

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