Hypomethylation of the proximal and intronic regulatory regions of the IFN-γ gene is not essential for its transcription by naive CD4⁺ T cells cultured with IL-4

Recently, long-term preculture with IL-4 or IL-7 has been reported to induce IFN-γ-producing ability in naive CD4⁺ T cells without stimulation via TCR. The mechanism of IFN-γ-transcription in naive CD4⁺ T cells precultured with IL-4 was analyzed and compared with that in typical Th1 cells by focusing on the TATA proximal and first intronic regulatory regions of the IFN-γ gene. Both regulatory regions in these IL-4-primed naive CD4⁺ T cells, which produce a large amount of IFN-γ upon stimulation with PMA and ionomycin, were completely methylated in contrast to the same hypomethylated regions in Th1 cells. DNase I hypersensitive site analysis suggested that both regulatory regions in IL-4-primed naive CD4⁺ T cells were not active for IFN-γ-expression. Moreover, we demonstrated that the composition of transcriptional factors that can bind to the proximal regulatory region is different between IL-4-primed naive CD4⁺ T cells and Th1 cells. These results indicated that the transcriptional machinery involved in the expression of the IFN-γ gene by CD4⁺ T cells varied depending on their modes of differentiation in both the responsive regulatory regions and the specific nuclear factors.