Extended Data Fig. 9: Sensing of TERRA by ZBP1 triggers an IFN response.
From: Telomere-to-mitochondria signalling by ZBP1 mediates replicative crisis
a, Immunoblotting of growing (PD45) IMR90E6E7 expressing either WT ZBP1(S)-Flag or mutant ZBP1(S)-Flag lacking Zα2 with either TRF1(ΔN) or VP16-TRF1(ΔN). Protein extracts were collected at day 4 post-doxycycline treatment (1 μg / ml). GAPDH loading control. Two independent experiments were performed. b, RNA-dot blot performed on RNA isolated from cells as in a using 32P-dCTP-labelled probes targeting either TERRA or GAPDH transcripts. RNA extracts were collected at day 4 post-doxycycline treatment (1 μg ml−1). GAPDH loading control. RNase A treatment was used to assess possible DNA contamination. Two independent experiments were performed. c, Scatter plot with bars showing RT-qPCR analysis with 5 pairs of subtelomere-specific primers designed to measure TERRA transcribed from individual chromosome ends. Expression levels were normalized to control cells expressing TRF1(ΔN). RNA extracts were collected from cells as in a. Bars represent mean ± s.d. from technical replicates. n: number of technical replicates. One-way ANOVA, ** p < 0.01, *** p < 0.001. Two independent experiment were performed. Abbreviations: ZBP1(S): short isoform; Dox: doxycycline; WT: wild-type; fRIP: formaldehyde RNA immunoprecipitation; PDs: population doublings.
