Extended Data Fig. 10: Observations in Pfa-1 cells are consistent with those in HT-1080 cells.
a-c. Temporal correlation of the onset and progression of lipid peroxyl radical production, cell morphology change and membrane permeability at basal conditions, with RSL3 (1 µM), or 48 hours after TAM (1 µM) treatment (replicates of Fig. 2h-i). Lipid peroxyl radical generation was recorded via H4B-PMHC enhancement (10 nM, green curve). Cell death was marked visually by the time of blebbing (black curve). Membrane integrity was assessed by PI (1 µM, orange curve) fluorescence. Each experiment was repeated twice on different days with similar results. d. Tabulated values of parameters describing CTCF of fluorogenic RTAs and morphology change curves in the presence of RSL3 (1 μM) or 48 hours after TAM (1 μM) treatment. e-f. Colocalization (100x HILO) of H4B-PMHC (10 nM, cyan) with ER Tracker Red (1 μM, not shown due to poor staining) in GPX4 proficient Pfa-1 cells with RSL3 (1 μM) treatment (e) or GPX4 KO Pfa-1 cells (f). Scale bars = 12 μm. Scale bar of the enlarged region = 4 μm. Colocalization of all channels (H4B-PMHC and DIC) is shown in the “merged” panel. Each experiment was repeated twice on different days with similar results. In each experiment, 4 FOVs were monitored.
