Fig. 4: USP18 loss in human cancer cells alters the landscape of ISGs.

a Western blots depicting proteins related to IFN-signaling and the DNA damage response in WT, USP18^+/− (2 different clones) and USP18^−/− (2 different clones) THP-1, and WT, and USP18^−/− (2 different clones) MDA-MB-231 cells. b Percentage of annexin V⁺ THP-1 cells (WT, USP18^+/− or USP18^−/−) when treated with IFNα (1000 U/ml) for the indicated times. (n = 3 independent samples each). Data represent mean ± s.d. c Volcano plot of RNAseq data comparing WT vs WT + IFNα and USP18^−/− (KO) vs KO + IFNα (1000U/ml) THP-1 cells (n = 3 independent samples each). p-value was determined by two-sided Wald test adjusted with Benjamini–Hochberg approach. d Heat map representing RNAseq data for MDA-MB-231 WT and USP18^−/− cells with or without IFNα (1000 U/ml) for 6 h (n = 3 independent samples each). e Heat map representing RNAseq data from THP-1 WT and USP18^−/− cells with or without IFNα treatment for 6 h. IPA pathway analysis and representative genes in clusters A and D are shown. p-value was determined by right-tailed Fisher’s exact test adjusted with Benjamini–Hochberg approach. f Dot plot and schematic representing how ISGs were classified as ‘typical’ versus ‘atypical’. Genes upregulated in USP18^−/− + IFNα vs USP18^−/− and not in WT + IFNα vs WT were labeled ‘Atypical ISGs’ (605 genes) and genes upregulated in both were labeled ‘Typical ISGs’ (448 genes). Within atypical ISGs, 262 genes have been reported as IFNα (1000 U/ml) inducible in human cells. However, 343 genes are not reported as ISGs by Interferome analysis. These genes were further divided into ‘hidden atypical ISGs’ or ‘Non-canonical atypical ISGs’ depending on the respective presence or absence of an ISRE and/or STAT1 binding motif/data in their promoter/enhancers. g Typical ISGs and atypical ISGs were analyzed by GO biological process analysis. p-value was determined by one-sided Fisher’s exact test adjusted with Benjamini–Hochberg approach. h qRT-PCR analysis of WT and USP18^−/− THP-1 cells (n = 2 independent samples each) treated with or without IFNα (1000 U/ml) for indicated times. Data represent mean ± s.d. p-value was determined by one-way ANOVA test. Source data are provided as a Source Data file.