Extended Data Fig. 7: Ku depletion induces PKR and OAS activation beyond IFN.
From: Ku limits RNA-induced innate immunity to allow Alu expansion in primates
a, IFNβ (up to 40 ng/ml) decrease in viability ~10% in 3 days. b. Poly I:C of 10 ng/ml and 100 ng/ml causes 40% and 99% loss of viability, regardless of Ku-AID tag (no IAA or Dox). The bars represent the mean and SEM from multiple repeats. c, Ku80-AID cells showed increased OP-puro low populations after IAA and Dox treatment, suggesting reduced translation. Bars represent mean ± SD of n = 3 samples from three independent clones. Two-way ANOVA test with Sidak’s correction for multiple comparisons: p < 0.0001 ****. Dox alone slightly decreased OP-puro in parental AID cells, consistent with its mild effect on mitochondrial translation79. To confirm the translation defect was Ku-specific, 4OHT-treated Ku86flox/−:CreERT2 and Ku86flox/− controls were tested and showed minimal background changes (Fig. 3c). d, Multiple members of the OAS family are transcriptionally induced (up to 128-fold for OAS1) upon Ku degradation (day 4 after Ku degradation). The error bars represent the standard errors from three independently derived cell lines. To plot all the OAS family genes with varying expression levels on the same plot, the Log2 fold changes were calculated. The untreated sample, with a Log2FC of 0, was excluded from the plot. Thus, no brackets can be drawn. e, The 28S versus 5S and 5.8S rRNA ratios in DMSO-treated cells (Ctrl, n = 5) versus IAA&Dox-treated Ku80-AID cells (n = 5 for CDK4/6i treated samples, otherwise n = 3), and poly I:C (100 ng/ml) transfected cells (n = 3). f, The representative histogram shows the relative reduction of 28S size and the relative decrease of 28S rRNA abundance after Ku80 degradation. g, The 28S versus 18S rRNA ratio in 4OHT treated control Ku86flox/− cells and Ku86flox/−:CreERT2 cells (Day7). For panel d, e, and g, at least three independent samples were measured at each time point from multiple clones. The means and SDs were plotted. The p-values were calculated via an unpaired two-tailed Student’s t-test, p < 0.0001 ****; p > 0.05 n.s.
