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. 2019 Dec 2;116(51):25790–25799. doi: 10.1073/pnas.1907224116

Fig. 5.

Fig. 5.

Hhex binds to and suppresses the Foxp3 locus and promoters of Treg signature genes. (A and B) Binding of Hhex to the Foxp3 locus and promoters of Treg signature genes was measured by ChIP. iTreg cells transduced with empty (ctrl), Hhex-FLAG, or HhexΔHD-FLAG (without the homeodomain) expression vectors were used. Enrichment of the target DNA was quantified by qPCR following immunoprecipitation with a FLAG antibody. (C and D) Transactivation of the promoters of Foxp3 and Treg signature genes by Hhex was measured by transient reporter assay. EL4 cells were transfected with Foxp3P-, Foxp3P+CNS1-, CD25P-, Ctla4P-, GitrP-, or IcosP–LUC reporter constructs together with an empty vector (ctrl) or a Hhex expression vector. After 16 h, cells were left untreated (nontreated) or were stimulated with PMA and ionomycin (stim.). Promoter activity is shown as the fold change (FC) relative to the nontreated ctrl. All error bars indicate the SD, and P values were calculated using Student’s t tests. *P < 0.05, **P < 0.01, ***P < 0.001. ns, not significant.