Figure 8. c-Rel regulates the expression of a restricted set of genes at early stages of CD4⁺ T cell activation.

(A) Heatmap showing log₂ FC of expression in PMA-stimulated naive CD4⁺ T cell at 2 hours. Only genes both differentially expressed in response to stimulation in controls (adjusted P < 0.05 and | log₂ FC | >1) and differentially expressed in P (adjusted P < 0.05 and | log₂ FC | >2) are shown. For selected genes, the log₂ FC between controls and P is indicated. (B) DNA binding motifs at PMA-induced c-Rel binding peaks. The de novo motif discovery P value and family of transcription factors are shown. (C) Bubble histogram showing the association of P’s dysregulated genes with c-Rel genomic binding sites as defined by ChIP-Seq in PMA-stimulated naive CD4⁺ T cells and located within 10 kb of their transcription start site. The association was computed for all c-Rel peaks and for a subgroup of peaks for which c-Rel binding increased by more than 5-fold following stimulation (1309 peaks). The circle size represents the significance of the association (–log₁₀ Fisher’s exact test P value). The color gradient shows the ratio of the number of peaks proximal (within 10 kb) to these genes relative to randomly selected gene sets. (D) Genomic snapshots of selected genes with lower levels of expression in P. The top 3 tracks show the control input DNA and ChIP-Seq for c-Rel in nonstimulated (NS) and PMA-treated naive CD4⁺ T cells from 3 controls; blue boxes indicate significant c-Rel binding peaks. Below the gene structure are shown the normalized RNA-Seq profiles for naive CD4⁺ T cells from controls (overlaid) and P, at steady state or after stimulation. (E) RT-qPCR after 2 hours of activation of naive CD4⁺ T cells from 4 controls and P with anti-CD3 mAb with or without anti-CD28 mAb. Data are displayed as 2-ΔCt after normalization to GUS expression. Error bars represent the SD. n = 1.