Compelling evidence suggests that the transcription factor Foxp3 acts as a master switch governing the development and function of CD4+ regulatory T cells (Tregs). However, whether transcriptional control of Foxp3 expression itself contributes to the development of a stable Treg lineage has thus far not been investigated. We here identified an evolutionarily conserved region within the foxp3 locus upstream of exon-1 possessing transcriptional activity. Bisulphite sequencing and chromatin immunoprecipitation revealed complete demethylation of CpG motifs as well as histone modifications within the conserved region in ex vivo isolated Foxp3+CD25+CD4+ Tregs, but not in naïve CD25−CD4+ T cells. Partial DNA demethylation is already found within developing Foxp3+ thymocytes; however, Tregs induced by TGF-β in vitro display only incomplete demethylation despite high Foxp3 expression. In contrast to natural Tregs, these TGF-β–induced Foxp3+ Tregs lose both Foxp3 expression and suppressive activity upon restimulation in the absence of TGF-β. Our data suggest that expression of Foxp3 must be stabilized by epigenetic modification to allow the development of a permanent suppressor cell lineage, a finding of significant importance for therapeutic applications involving induction or transfer of Tregs and for the understanding of long-term cell lineage decisions.

Although T helper 17 (TH17) cells have been acknowledged as crucial mediators of autoimmune tissue damage, the effector cytokines responsible for their pathogenicity still remain poorly defined, particularly in humans. In mouse models of autoimmunity, the pathogenicity of TH17 cells has recently been associated with their production of granulocyte-macrophage colony-stimulating factor (GM-CSF). We analyzed the regulation of GM-CSF expression by human TH cell subsets. Surprisingly, the induction of GM-CSF expression by human TH cells is constrained by the interleukin-23 (IL-23)/ROR-γt/TH17 cell axis but promoted by the IL-12/T-bet/TH1 cell axis. IL-2-mediated signal transducer and activator of transcription 5 (STAT5) signaling induced GM-CSF expression in naïve and memory TH cells, whereas STAT3 signaling blocked it. The opposite effect was observed for IL-17 expression. Ex vivo, GM-CSF(+) TH cells that coexpress interferon-γ and T-bet could be distinguished by differential chemokine receptor expression from a previously uncharacterized subset of GM-CSF-only-producing TH cells that did not express TH1, TH2, and TH17 signature cytokines or master transcription factors. Our findings demonstrate distinct and counterregulatory pathways for the generation of IL-17- and GM-CSF-producing cells and also suggest a pathogenic role for GM-CSF(+) T cells in the inflamed brain of multiple sclerosis (MS) patients. This provides not only a scientific rationale for depleting T cell-derived GM-CSF in MS patients but also multiple new molecular checkpoints for therapeutic GM-CSF suppression, which, unlike in mice, do not associate with the TH17 but instead with the TH1 axis.

After being activated by antigen, helper T lymphocytes switch from a resting state to clonal expansion. This switch requires inactivation of the transcription factor Foxo1, a suppressor of proliferation expressed in resting helper T lymphocytes. In the early antigen-dependent phase of expansion, Foxo1 is inactivated by antigen receptor-mediated post-translational modifications. Here we show that in the late phase of expansion, Foxo1 was no longer post-translationally regulated but was inhibited post-transcriptionally by the interleukin 2 (IL-2)-induced microRNA miR-182. Specific inhibition of miR-182 in helper T lymphocytes limited their population expansion in vitro and in vivo. Our results demonstrate a central role for miR-182 in the physiological regulation of IL-2-driven helper T cell-mediated immune responses and open new therapeutic possibilities.

In the bone marrow, a population of memory T cells has been described that promotes efficient secondary immune responses and has been considered to be preactivated, owing to its expression of CD69 and CD25. Here we show that human bone marrow professional memory T cells are not activated but are resting in terms of proliferation, transcription, and mobility. They are in the G0 phase of the cell cycle, and their transcriptome is that of resting T cells. The repertoire of CD4(+) bone marrow memory T cells compared with CD4(+) memory T cells from the blood is significantly enriched for T cells specific for cytomegalovirus-pp65 (immunodominant protein), tetanus toxoid, measles, mumps, and rubella. It is not enriched for vaccinia virus and Candida albicans-MP65 (immunodominant protein), typical pathogens of skin and/or mucosa. CD4(+) memory T cells specific for measles are maintained nearly exclusively in the bone marrow. Thus, CD4(+) memory T cells from the bone marrow provide long-term memory for systemic pathogens.

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