TCR- and IL-2-dependent transcriptomes of human regulatory T cells identifies IL-2R signaling as largely enhancing TCR/CD28-dependet gene expression

Although regulatory T cells (Tregs) constitutively express high levels of the IL-2R, the contribution of IL-2R signaling in the expansion of human Tregs is poorly understood. Genome-wide transcriptome analyses was performed to identify IL-2R- and TCR/CD28-dependent processes for human Tregs. Human Tregs were cultured for 4 and 16 hr in media alone or with IL-2, anti-CD3/anti-CD28 plus anti-IL-2 to inhibit potential endogenous IL-2 from any contaminating Teff cells, or a combination of anti-CD3/anti-CD28 and IL-2. RNA-seq of the Tregs from these cultures revealed that in comparison to cells cultured in only media, 2616 and 6239 unique genes were up- or down-regulated 1.5-fold (FDR<0.01) in one or more culture conditions at 4 and 16 hr, respectively. At both time points, IL-2 regulated many fewer genes than stimulation through TCR/CD28. By 16 hr nearly 80% of the IL-2-dependent genes were also activated by TCR/CD28 whereas approximately 3% are solely responsive to IL-2. Thus, the role of IL-2 is largely to enhance the up- or down-regulation of many genes induced after stimulation through TCR/CD28. Overall, fully activated Tregs (through TCR/CD28/IL-2R ) express genes in pathways that are required to prepare these cells for proliferation, differentiation and expression of some immune functions.