KDM5 histone demethylases repress immune response via suppression of STING
Fig 3
Activation of ISGs by KDM5 inhibition is dependent on the cGAS-STING-TBK1-IRF3 signaling pathway.
(A) Schematic of the pattern recognition receptor pathways. (B–D) Western blot (panel B and C) and RT-qPCR (panel D) analyses of MCF7 cells with knockout of the indicated genes after treatment with DMSO or 1 μM KDM5-C70 for 6 days. (E, F) RT-qPCR (panel E) and western blot (panel F) analyses of MCF7 cells with knockout of the indicated genes 5 days after transfection with the indicated siRNAs. (G) RT-qPCR analysis of MCF7 cells with knockout of the indicated genes after treatment with DMSO or 1 μM KDM5-C70 for 6 days. (H) Flow cytometry plots (left panel) and quantification of percentage of GFP-positive cells (right panel) in the indicated MCF7 knockout cells 24 hours after infection with VSV-GFP virus at MOI 0.5. Error bar denotes SEM. Representative data from triplicate experiments are shown in panel D, E, and G. Three biological replicates are shown in panel H. #p < 0.01 for inhibitors versus DMSO (panel D, G, and H), knockdown of KDM5B and KDM5C versus control (panel E). ^p < 0.01 for knockout sgRNA versus control sgRNA (panel D and G). The numerical values used to generate graphs in panel D, E, G, and H are available in S1 Data. cGAS, cyclic GMP-AMP synthase; IRF3, interferon regulatory factor 3; ISG, interferon-stimulated gene; MOI, multiplicity of infection; RT-qPCR, reverse transcription followed by quantitative PCR; sgRNA, single guide RNA; siRNA, small interfering RNA; STING, stimulator of interferon genes; TBK1, TANK-binding kinase 1; VSV-GFP, vesicular stomatitis virus carrying a green fluorescent protein reporter.