Figure S5.
Horizontal bar graph a show the top 5 pathways from C2:REACTOME per regulon, with the x-axis showing negative log10 adjusted p-value ranging from 0 to 4, where Common All Down regulon bars extend to approximately 1 for pathways including RMTS methylate histone arginines, regulation of endogenous retroelements by PIWI interacting RNAs, regulation of endogenous retroelements, and chromatin organization, while Common All Up regulon bars show notably longer extensions with constitutive signaling by AKT1 E17K in cancer and signaling by interleukins reaching up to 4. Horizontal bar graph b show top 5 pathways from C2:REACTOME for three regulons including IRF2.IRF8 specific down, IRF4.IRF8 specific up, and IRF8.IRF4 specific down, on a negative log10 adjusted p-value x-axis ranging from 0 to 15, where IRF8.IRF4 specific down bars dominate with cellular responses to stimuli, viral infection pathways, and infectious disease extending beyond 10, while IRF4.IRF8 specific up and IRF2.IRF8 specific down bars remain below 5 across their respective pathways. Horizontal bar graphs c show two sets of pathway enrichment results, the first from Hallmark gene sets and the second from C2:REACTOME, both plotted across six regulons including IRF2 and IRF4 and IRF8 specific up and down versus control groups, with the Hallmark x-axis extending to 30 showing interferon gamma response and interferon alpha response reaching beyond 25 for IRF8 specific up versus control, and the C2:REACTOME x-axis extending to 25 showing cellular responses to stimuli and infectious disease and viral infection pathways reaching above 20 for IRF4 specific down versus control and IRF8 specific up versus control regulons. Sequence logos d display DNA binding motifs for IRF2, IRF4, and IRF8 with their corresponding q-values of 0.0000, 0.0006, and 0.0181 respectively, alongside their known motif references from Homer ChIP-seq datasets, where all three motifs share a core GAAGT sequence with flanking variations distinguishing each factor.
Overrepresentation analysis reveals enriched pathways upon IRF KO in CD8 TILs. (a and c) Overrepresentation of pathways from MSigDB Hallmark and MSigDB C2:REACTOME gene sets across up- and downregulated genes. (a and b) In a, enriched pathways from C2:REACTOME using up- and downregulated genes common to all KOs and common to IRF2 and IRF8 KO and to IRF4 and IRF8 KO in b are shown. (c) In c, enriched pathways using up- and downregulated genes specific to IRF2, IRF4, and IRF8 KO are shown for Hallmark (top) and C2:REACTOME (bottom). (d) Homer known motif enrichment analysis done for CUT&RUN data for quality purpose for each of the IRF target, showing significant target enrichment.