Figure 5. SKI requires SMAD4 to modulate Rorc gene transcription.
(a) CD4+ T cells isolated from wild-type (WT) or (b) Cd4-cre;Smad4fl/fl (SMAD4 KO) mice were cultured with IL-21 and TGF-β, and then transduced with control retrovirus (RV) or SKI-expressing retrovirus (RV SKI). IL-17A+ cells were analyzed by flow cytometry. Representative results and statistical analysis are shown. (c) CD4+ T cells isolated from wild-type (WT) mice were cultured with IL-21 and TGF-β, and then transduced with control retrovirus (RV), SKI-expressing retrovirus (RV SKI), or SKI and RORγt co-expressing retrovirus (RV SKI+RORγt). IL-17A+ cells were analyzed by flow cytometry. Representative results and statistical analysis are shown. (d) CD4+ T cells isolated from wild-type (WT) mice were cultured under indicated conditions, cells were harvested after 3 days. ChIP assay was performed with control IgG antibody or SKI antibody. The relative enrichment of SKI binding to the Rorc locus was determined. (e) CD4+ T cells isolated from wild-type (WT) and Cd4-cre;Smad4fl/fl (SMAD4 KO) mice were cultured with IL-21 and TGF-β receptor I inhibitor, cells were harvested after 3 days. ChIP assay was performed with control IgG antibody or SKI antibody. The relative enrichment of SKI binding to the Rorc locus was determined.