Figure 4. TGF-β-induced IκB-ζ represses Ifng gene expression.

(A) Expression of IκB-ζ in naïve CD4⁺ T cells cultured with/without TGF-β1 stimulation. (B) Expression of IκB-ζ in naive CD4⁺ T cells from Smad2^flox/flox mice or Smad2^flox/flox Lck-Cre mice that were cultured for 24 hours. (C and D) CD4⁺ T cells were retrovirally transduced to express IκB-ζ and GFP. (C) The numbers indicate the percentage of IFN-γ-producing GFP⁺CD4⁺ T cells. Data are representative of 3 independent experiments. (D) Relative IFN-γ-producing GFP⁺CD4⁺ T cells. The percentage of IFN-γ-producing GFP⁺CD4⁺ T cells from the IRES-GFP control is set as “1.” Data shown represent the means ± sd (n = 3). (E) Relative histone acetylation of the IFN-γ promoter region or CNS region in naïve CD4⁺ T cells from control or cKO mice. Purified naïve CD4⁺ T cells were cultured for 3 d in the presence of TGF-β1. Histone acetylation was analyzed by ChIP assays, performed by use of an acetyl-histone (AcH) H3 antibody and control IgG. Data represent means ± sd with triplicate samples. (F) IFN-γ reporter activity in HEK293 cells. Reporter activity in the absence of p65 and IκB-ζ expression vector was set to 1. Data shown reflect means ± sd (triplicate samples). Data are representative of 3 (A–E) or 2 (F) independent experiments. *P < 0.05; **P < 0.01.