T Cell Receptor Stimulation-Induced Epigenetic Changes and Foxp3 Expression Are Independent and Complementary Events Required for Treg Cell Development

Abstract

The transcription factor Foxp3 is essential for the development of regulatory T (Treg) cells, yet its expression is insufficient for establishing the Treg cell lineage. Here we showed that Treg cell development was achieved by the combination of two independent processes, i.e., the expression of Foxp3 and the establishment of Treg cell-specific CpG hypomethylation pattern. Both events were induced by T cell receptor stimulation. The Treg cell-type CpG hypomethylation began in the thymus and continued to proceed in the periphery and could be fully established without Foxp3. The hypomethylation was required for Foxp3(+) T cells to acquire Treg cell-type gene expression, lineage stability, and full suppressive activity. Thus, those T cells in which the two events have concurrently occurred are developmentally set into the Treg cell lineage. This model explains how Treg cell fate and plasticity is controlled and can be exploited to generate functionally stable Treg cells.

Foxp3+ T cells expressing RORγt represent a stable regulatory T-cell effector lineage with enhanced suppressive capacity during intestinal inflammation

“…54 However, the overall methylation pattern in the Treg-specific epigenetic signature genes in Foxp3 þ RORgt þ T cells may rather support the idea that a fraction of Foxp3 þ RORgt þ T cells are peripherally induced and are on the way to acquire full demethylation in their Tnfrsf18 and Ikzf4 loci, as recently suggested. 40 Further supporting this hypothesis, we also found a mixed pattern of Helios expression in Foxp3 þ RORgt þ T cells, indicating that the Foxp3 þ RORgt þ T cells observed under physiologic conditions may be a mixture of not only thymic but also peripherally induced origin, especially in the gut-associated tissues. It should be noted however that Helios expression may not unambiguously distinguish thymus-derived Tregs from pTregs, as several studies demonstrated that Foxp3 þ pTregs may under certain conditions also express Helios.…”

“…In order to assess the stability of the Treg phenotype of Foxp3 þ RORgt þ T cells, we focused on the CpG methylation status in different gene loci that have been described to be of significance for Treg lineage stability. 36,38,40 Our data showed full demethylation of the TSDR in Foxp3 þ RORgt þ T cells and substantial demethylation of Ctla4, Tnfrsf18, and Ikzf4, a clear indication of full Treg lineage stability and function according to Ohkura et al 40 Of note, the degree of demethylation in Ikzf4 was lower in Foxp3 þ RORgt þ T cells compared with Foxp3 þ Tregs. It has been argued that downmodulation of Eos (encoded by Ikzf4) transcription is indicative of Tregs undergoing reprogramming into T helper-like cells.…”

“…39 In addition, a number of Treg-specific epigenetic signature genes (e.g., Ctla4, Tnfrsf18 (GITR) and Ikzf4 (Eos)) need to be fully demethylated to enable Foxp3 þ T cells to acquire Treg-specific gene expression, lineage stability, and full suppressive activity. 40 Interestingly, Foxp3 þ RORgt þ T cells also showed substantial demethylation at Ctla4, Tnfrsf18, and Ikzf4, although the degree of demethylation at Tnfrsf18 and Ikzf4 was somewhat lower as compared with Foxp3 þ Tregs ( Figure 3b). As expected, these loci remained largely methylated in RORgt þ T cells, except for Ctla4 that was substantially demethylated in these cells.…”

Regulatory T cells in cancer immunotherapy

“…For example, whereas TCR stimulation upregulates the expression of ZAP-70 in Tconv cells, it downregulates ZAP-70 expression in Treg cells [33]. Foxp3 binds to the promoter region of ZAP-70 gene [33][34][35] and retroviral expression of Foxp3 in Tconv cells reduces their ZAP-70 expression [33]. It remains to be determined how TCR signaling attenuation at the level of ZAP-70 may contribute to Treg-specific functions.…”

Harnessing the plasticity of CD4+ T cells to treat immune-mediated disease

“…Importantly, TCR signal strength drives the first divergence of inflammatory versus regulatory subsets during T cell development in the thymus. An increased strength of signal, which is imparted by TCRs that recognize self-antigen, imprints a distinct epi genetic state and induces FOXP3 expression to generate the T Reg cell lineage 61,62 .…”