Fig. 2: PrPC sensitizes cells to ferroptosis via lipid remodeling.
A Cell survival of HT-1080 PrPC OE (OE) and empty vector control (C) cells treated with cell death inhibitors. Cells were infected for 3 days, selected with puromycin for 24 h, then cultured with inhibitors 10 μM α-tocopherol (aToc) or 2 µM ferrostatin-1 as a ferroptosis inhibitor, 10 μM z-VAD-FMK (zVAD) as an apoptosis inhibitor, and 10 μM necrostatin-1 (Nec-1) as a necroptosis inhibitor, respectively, for 3 days. B (Left) Dose-response curves illustrating sensitivity of PrPC-expressing HT-1080 cells (PrPC OE) compared to empty vector control cells (control OE) to IKE treatment over 16 h. (Right) Survival of PrPC OE cells compared to control against ferroptosis inducer 0.3 μM IKE (16 h). (Lower panels) Sensitivity of GPX8 knockout (KO) cells with (+dox) and without (-dox) PrPC OE. Inset shows GPX8 protein levels by Western and total protein loading compared to knockout control cells. Viability data are plotted as representative mean ± SD of n = 3 technical replicates for independent experiments repeated at least three times with similar outcomes. C PrPC OE effect on lipid peroxidation induced by 0.1 μM RSL3 induction in HT-1080 OE and control cells measured by BODIPY 581/591 C11 (BODIPY-C11). A typical FACS histogram of n = 3 technical replicates of three independent repetitions is depicted. The bar graph shown on the right illustrates the BODIPY-C11 median intensity ± SEM. D Comparative analysis of lipid levels in PrPC OE HT-1080 cells compared to control cells, highlighting fatty acid acyl chain saturation in normalized lipidomics data. Normalization was performed by glucose peak area for each sample and each feature. XCMS data analysis and tentative annotation (of 617 known lipids) were according to the Bruker UPLC-Q-TOF protocol in positive mode, detecting 10,521 total features. Significant lipids have -log10 P > 1.30. Viability data are representative means of n = 9 (A) biological or n = 3 (B) technical replicates for experiments repeated independently at least three times. P-values of two-way ANOVA multiple comparisons with Tukey post-test are shown (A–C). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Lipid statistics (D) were determined by P-values (P < 0.05, n = 5, two-tailed Welch’s t-test). n = 5 PrPC OE and n = 5 control samples. Source data are provided as a Source Data file.
