Fig. 2: H3L induces DNA damage via upregulating IL1A.

A Signaling pathway enrichment analyses of differentially expressed genes. Enrichment analyses were run on Reactome. Heatmap showing differentially expressed genes involved in interferon signaling pathway (B–D) and interleukin signaling pathway (D). E RNA-seq read counts of signature genes involved in interferon and interleukin signaling pathways. Four biological replicates were applied for RNA-seq. *p < 0.05 (vs. Control). F Relative expression levels of signature genes involved in interferon and interleukin signaling pathways in RNA-seq. G RT-qPCR showing relative expression levels of signature genes involved in interferon and interleukin signaling pathways. *p < 0.05 (vs. Control). H The enzyme-linked immunosorbent assay (ELISA) showing relative expression levels of IL1A, IFN-γ, IFN-α, and IFN-β. *p < 0.05 (vs. Control). Relative level in the Y-axis meant the read count on the absorption at 450 nm by the equipment. I Representative ChIP-seq peaks on human IL1A. There was a potential binding site on upstream distal enhancer of IL1A showed by ChIP-seq. TSS, transcriptional start site. Data was from the published datasets in UCSC genome browser. J ChIP-qPCR showing IRF4 enrichment on the binding site on upstream distal enhancer of IL1A in WT hESCs. *p < 0.05 (vs. IgG). K RT-qPCR showing the expression of IRF4 in negative control and shRNAs knockdown hESCs. *p < 0.05 (vs. Negative shRNA control). L Flow cytometry quantification of IL1A⁺ cells in negative control and IRF4-shRNA knockdown hESCs. *p < 0.05 (vs. Negative shRNA control). M Flow cytometry quantification of γ-H2AX⁺ hESCs treated with different concentration of IL1A. IL1A antibody was used to block IL1A activity. *p < 0.05 (vs. 0 ng/ml), ^#p < 0.05. N Flow cytometry quantification of TUNEL⁺ hESCs treated with different concentration of IL1A. IL1A antibody was used to block IL1A activity. *p < 0.05 (vs. 0 ng/ml), ^#p < 0.05. O Working model of H3L-driven IL1A in inducing DNA damage and cell death.