Differential occupancy of SATB1, GATA3, and HDAC1 on IL-5 proximal promoter in Th1 and Th2 cells during T-cell differentiation. (A) Naive CD4⁺ cells were polarized for 24 hours and subjected to ChIP assay. Occupancy of IL-5 promoter by SATB1, GATA3, and HDAC1 during Th1 () and Th2 (▬) differentiation was monitored by quantitative RT-PCR using primers corresponding to fragment A of the IL-5 promoter. Data represent fold change in occupancy of indicated proteins compared with their corresponding IgG controls, after normalizing for the input chromatin. Each error bar represents SD calculated from triplicates. (B) ChIP-PCR analysis for occupancy of SATB1, GATA3, and HDAC1 on proximal IL-5 promoter in Th1 and Th2 cells was performed as described in “ChIP and ChIP-on-chip.” ChIP-PCR products (∼ 200 bp) from 2 representative cord blood (CB) samples are depicted. Vertical lines have been inserted to indicate a repositioned gel lane. (C) Naive CD4⁺ T cells nucleofected with scrambled (Scr) control or SATB1-siRNA (siR) were cultured in Th2-polarizing conditions for 24 hours and subjected to ChIP assay. Occupancy of IL-5 promoter by SATB1, GATA3, and HDAC1 during Th2 differentiation in the presence of Scr () or siR (▬) was monitored by quantitative RT-PCR using primers corresponding to fragment A of IL-5 promoter as described in “Quantitative RT-PCR analyses.” Error bar represents SD calculated from triplicates. *P < .005. **P < .01.