Extended Data Fig. 3: Loss of ZBP1 attenuates IFN response and promotes crisis bypass.
From: Telomere-to-mitochondria signalling by ZBP1 mediates replicative crisis
a, Line chart showing growth curves analysis of IMR90E6E7 or WI38SV40LT that have bypassed senescence expressing control sgRNAs (sgLUC, sgGFP) or three individual sgRNAs targeting ZBP1 (sgZBP1#1, sgZBP1#2, sgZBP1#3). Three independent experiments were performed. b, Crystal violet viability assay of IMR90E6E7 or WI38SV40LT as in a. Cells were seeded at day 30 and fixed at day 40. One experiment was performed. c, Immunoblotting of IMR90E6E7 or WI38SV40LT as in a. Protein extracts were collected at day 30 when control cells (sgLUC, sgGFP) entered crisis and upregulated ZBP1. GAPDH loading control. Two independent experiments were performed. d, Line chart showing IncuCyte analysis of cell death of IMR90E6E7 or WI38SV40LT as in a, measured in real time by Cytotox green incorporation at day 30 of the experiment. Bars represent mean ± s.e.m. from technical replicate. One-way ANOVA, *** p < 0.001. Two independent experiments were performed. e, Scatter plots with bars showing RT-qPCR analysis of IFNB1 in IMR90E6E7 or WI38SV40LT as in a. RNA extracts were collected at day 30 of the experiment. Expression levels were normalized to control cells with sgLUC. Bars represent mean ± s.d. of technical replicates. n: number of technical replicates. One-way ANOVA, ns: not significant, *** p < 0.001. Two independent experiments were performed. f, Scatter plots with bars showing ELISA assays of IFN-β levels secreted by IMR90E6E7 or WI38SV40LT as in a. Culture supernatant were collected at day 30 of the experiment. Bars represent mean ± s.d. of technical replicates. n: number of technical replicates. One-way ANOVA, ns: not significant, *** p < 0.001. One experiment was performed. Abbreviations: GFP: Green fluorescent protein; LUC: luciferase; PDs: population doublings.
