Extended Data Fig. 2: Depletion of KDM5B in mouse melanoma cells activates the type I interferon response through both RNA and DNA sensing pathways.
From: KDM5B promotes immune evasion by recruiting SETDB1 to silence retroelements
a, Gene set enrichment analysis comparing RNA-seq data from Kdm5b−/− (Kdm5b sg) YUMMER1.7 cells with control YUMMER1.7 cells. Shown are selected top upregulated pathways (FDR<=0.05). b, Western blot analyses of DNA and RNA sensors, as well as signalling proteins involved in the type I IFN pathway. T-, total; p-, phosphorylated. For gel source data, see Supplementary Fig. 1. c, RT-qPCR analyses of ISGs in Ifngr1−/− YUMMER1.7 mouse melanoma cells with Ctl sg, Kdm5b sg1 and Kdm5b sg2. d, RT-qPCR analyses of ISGs (left) and western blot analyses (right) of YUMM1.7 mouse melanoma cells with Ctl sg and Kdm5b sg. For gel source data, see Supplementary Fig. 1. e, RT-qPCR analyses of ISGs in 1445 mouse melanoma cells with Ctl shRNA and Kdm5b shRNA. f, Western blot analyses of the indicated cell lines used in Figure 2b and Extended Data Fig. 2g. For gel source data, see Supplementary Fig. 1. g, RT-qPCR analyses of selected ISGs in the indicated YUMMER1.7 cells treated with 10 ng/ml IFNγ for 24h. Data are mean ± SD. h, Western blot analyses of the indicated cell lines, used in Figure 2c. For gel source data, see Supplementary Fig. 1. i, RT-qPCR analyses (left) and western blot analyses (right) of YUMM1.7-Kdm5b sg cells with Ctl sg, Sting1 sg, Ddx58 (RIG-I gene) sg, Mda5 sg, Mavs sg, or Cgas sg. For gel source data, see Supplementary Fig. 1. j, RT-qPCR analyses of IRF9 in1445-Kdm5b sg cells with Ctl sg, Sting1 sg, Ddx58 sg, Mda5 sg, Mavs sg, or Cgas sg. k, RT-qPCR analyses of ISGs from YUMM1.7-Kdm5b sg cells treated with Ctl siRNA, Sting1 siRNA or Mavs siRNA for 3 days. l, RT-qPCR analyses of ISGs from 1445 cells with Ctl shRNA treated with Ctl siRNA, 1445 cells with Kdm5b shRNA treated with Ctl siRNA, Sting1 siRNA, or Mavs siRNA cells for 3 days. Data are means ± SEM. m, Tumor growth curve (left) and survival curve (right) of C57BL/6 mice injected with YUMMER1.7 cells of the indicated genotypes, WT (n = 5), Kdm5b−/−-Mda5−/− Cgas−/−-C1 (n = 4), Kdm5b−/−-Mda5−/− Cgas−/−-C3 (n = 4), Kdm5b−/− (n = 5). C1 and C3 are clone numbers. Data represent one experiment. Data are mean ± SEM. 2-way ANOVA analysis for comparison at day 20. n.s., not significant; ****p<0.0001. Log-rank test for survival curves. **p = 0.004 for C1 vs Kdm5b−/−; **p = 0.005 for C3 vs Kdm5b−/−; *p = 0.023 for C3 vs WT. Unpaired two-sided Students’ t tests were performed for all RT-qPCR data (c-e,g, i-l). n.s., not significant; *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.
