Fig. 9
The IL-6-STAT3 pathway regulates epigenetic modifications of the RFX1 locus. a Genome location of the RFX1 locus and the STAT3-binding sites in intron 7 of the RFX1 gene. b ChIP-qPCR assay of pSTAT3 binding in intron 7 of RFX1 in CD4+ T cells with or without IL-6 stimulation. Primer 1 and Primer 2 were used to amplify the fragment containing STAT3-binding sites. c Luciferase reporter assay analysis of STAT3 regulation on the enhancer activity of intron 7 in 293T cells transfected with STAT3 expression vector (STAT3-over) or empty expression vector (EV). A, B, and C represent the STAT3-binding sites in intron 7. d Luciferase reporter assay analysis of the enhancer activity of intron 7 in CD4+ T cells with IL-6 stimulation and negative control. e Luciferase reporter assay analysis of the enhancer activity of intron 7 in CD4+ T cells with IL-6 stimulation transfected with STAT3 expression vector (STAT3-over) or empty expression vector (EV). f ChIP-qPCR analysis of H3 acetylation level and H4 acetylation level of intron 7 of the RFX1 gene in CD4+ T cells with or without IL-6 stimulation. g BSP analysis of the DNA methylation level of CpG island in intron 7 of the RFX1 gene in CD4+ T cells with or without IL-6 stimulation. The left panel shows the methylation level of each CpG site, and the right panel shows the mean methylation level of all 30 CpG sites in the CpG island. Data are representative of three independent experiments (mean ± s.d.; n = 3). *P < 0.05, **P < 0.01, compared between the indicated groups. n.s., not significant. P-values were determined using two-tailed Student’s t-tests
