Arkadia-SKI/SnoN signaling differentially regulates TGF-β–induced iTreg and Th17 cell differentiation

TGF-β signaling is fundamental for both Th17 and regulatory T (Treg) cell differentiation. However, these cells differ in requirements for downstream signaling components, such as SMAD effectors. To further characterize mechanisms that distinguish TGF-β signaling requirements for Th17 and Treg cell differentiation, we investigated the role of Arkadia (RNF111), an E3 ubiquitin ligase that mediates TGF-β signaling during development. Inactivation of Arkadia in CD4+ T cells resulted in impaired Treg cell differentiation in vitro and loss of RORγt+FOXP3+ iTreg cells in the intestinal lamina propria, which increased susceptibility to microbiota-induced mucosal inflammation. In contrast, Arkadia was dispensable for Th17 cell responses. Furthermore, genetic ablation of two Arkadia substrates, the transcriptional corepressors SKI and SnoN, rescued Arkadia-deficient iTreg cell differentiation both in vitro and in vivo. These results reveal distinct TGF-β signaling modules governing Th17 and iTreg cell differentiation programs that could be targeted to selectively modulate T cell functions.

Prostaglandin E2 directly inhibits the conversion of inducible regulatory T cells through EP2 and EP4 receptors via antagonizing TGF-β signalling

Background and Purpose: Regulatory T (Treg) cells are essential for control of inflammatory processes by suppressing Th1 and Th17 cells. The bioactive lipid mediator prostaglandin E2 (PGE2) promotes inflammatory Th1 and Th17 cells and exacerbates T cell-mediated autoimmune diseases. However, the actions of PGE2 on the development and function of Treg cells, particularly under inflammatory conditions, are debated. In this study, we examined whether PGE2 had a direct action on T cells to modulate de novo differentiation of Treg cells. Experimental Approach: We employed an in vitro T cell culture system of TGF-β-dependent Treg induction from naive T cells. PGE2 and selective agonists for its receptors, and other small molecular inhibitors were used. Mice with specific lack of EP4 receptors in T cells were used to assess Treg cell differentiation in vivo. Human peripheral blood T cells from healthy individuals were used to induce differentiation of inducible Treg cells. Key Results: TGF-β-induced Foxp3 expression and Treg cell differentiation in vitro was markedly inhibited by PGE2, which was due to interrupting TGF-β signalling. EP2 or EP4 agonism mimicked suppression of Foxp3 expression in WT T cells, but not in T cells deficient in EP2 or EP4, respectively. Moreover, deficiency of EP4 in T cells impaired iTreg cell differentiation in vivo. PGE2 also appeared to inhibit the conversion of human iTreg cells. Conclusion and Implications: Our results show a direct, negative regulation of iTreg cell differentiation by PGE2, highlighting the potential for selectively targeting the PGE2-EP2/EP4 pathway to control T cell-mediated inflammation.

Arkadia-SKI/SnoN signaling cascade differentially regulates TGF-β-induced iTreg and Th17 cell differentiation

TGF-β signaling is fundamental for both Th17 and regulatory T cell (Treg) differentiation. However, these cells differ in requirements for downstream signaling components, such as various SMAD effectors, for their differentiation. To further characterize the mechanisms that distinguish TGF-β signaling requirements for Th17 and Treg cell differentiation, we investigated the role of Arkadia (RNF111), a RING type E3 ubiquitin ligase known to enhance TGF-β signaling during development. We find that Arkadia mediates the differentiation of induced-Treg (iTreg) but not Th17 cells both in vitro and in vivo. Inactivation of Arkadia in CD4+ T cells resulted in impairment of Treg cell differentiation in vitro and loss of RORγt+FOXP3+ iTreg cells in the intestinal lamina propria in vivo, which increased susceptibility to microbiota-induced mucosal inflammation. Furthermore, genetic ablation of two substrates of Arkadia, the transcriptional co-repressors SKI and SnoN, rescued in vitro and in vivo iTreg cell-differentiation of Arkadia-deficient cells. These results reveal distinct TGF-β signaling modules that govern Th17 and iTreg cell differentiation programs and that could be exploited therapeutically to selectively modulate T cell subsets in pathological settings such as autoimmunity or cancer.

CD40-signalling abrogates induction of RORγt+ Treg cells by intestinal CD103+ DCs and causes fatal colitis

Immune homeostasis in intestinal tissues depends on the generation of regulatory T (Treg) cells. CD103+ dendritic cells (DCs) acquire microbiota-derived material from the gut lumen for transport to draining lymph nodes and generation of receptor-related orphan γt+ (RORγt+) Helios−-induced Treg (iTreg) cells. Here we show CD40-signalling as a microbe-independent signal that can induce migration of CD103+ DCs from the lamina propria (LP) to the mesenteric lymph nodes. Transgenic mice with constitutive CD11c-specific CD40-signalling have reduced numbers of CD103+ DCs in LP and a low frequency of RORγt+Helios− iTreg cells, exacerbated inflammatory Th1/Th17 responses, high titres of microbiota-specific immunoglobulins, dysbiosis and fatal colitis, but no pathology is detected in other tissues. Our data demonstrate a CD40-dependent mechanism capable of abrogating iTreg cell induction by DCs, and suggest that the CD40L/CD40-signalling axis might be able to intervene in the generation of new iTreg cells in order to counter-regulate immune suppression to enhance immunity.

Thymus-Derived Regulatory T Cell Development Is Regulated by C-Type Lectin-Mediated BIC/MicroRNA 155 Expression

ABSTRACT Thymus-derived regulatory T (tTreg) cells are key to preventing autoimmune diseases, but the mechanisms involved in their development remain unsolved. Here, we show that the C-type lectin receptor CD69 controls tTreg cell development and peripheral Treg cell homeostasis through the regulation of BIC/microRNA 155 (miR-155) and its target, suppressor of cytokine signaling 1 (SOCS-1). Using Foxp3-mRFP/cd69 +/− or Foxp3-mRFP/cd69 −/− reporter mice and short hairpin RNA (shRNA)-mediated silencing and miR-155 transfection approaches, we found that CD69 deficiency impaired the signal transducer and activator of transcription 5 (STAT5) pathway in Foxp3+ cells. This results in BIC/miR-155 inhibition, increased SOCS-1 expression, and severely impaired tTreg cell development in embryos, adults, and Rag2−/− γc−/− hematopoietic chimeras reconstituted with cd69 −/− stem cells. Accordingly, mirn155 −/− mice have an impaired development of CD69+ tTreg cells and overexpression of the miR-155-induced CD69 pathway, suggesting that both molecules might be concomitantly activated in a positive-feedback loop. Moreover, in vitro-inducible CD25+ Treg (iTreg) cell development is inhibited in Il2rγ−/−/cd69 −/− mice. Our data highlight the contribution of CD69 as a nonredundant key regulator of BIC/miR-155-dependent Treg cell development and homeostasis.

Migratory, and not lymphoid-resident, dendritic cells maintain peripheral self-tolerance and prevent autoimmunity via induction of iTreg cells

There is evidence that dendritic cells (DCs) induce peripheral tolerance. Nevertheless, it is not known whether immature DCs in general are able to tolerize CD4+ T cells or if this is a prerogative of specialized subtypes. Here we show that, when autoantigen presentation is extended to all conventional mouse DCs, immature lymphoid tissue resident DCs are unable to induce autoantigen-specific regulatory T (iTreg) cell conversion. In contrast, this is an exclusive prerogative of steady-state migratory DCs. Because only lymph nodes host migratory DCs, iTreg cells develop and are retained solely in lymph nodes, and not in the spleen. Mechanistically, in cutaneous lymph nodes, DC-derived CCL22 contributes to the retention of iTreg cells. The importance of the local generation of iTreg cells is emphasized by their essential role in preventing autoimmunity.