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Summary: T helper 17 (Th17) cells specifically transcribe the Il17 and Il17f genes, which are localized in the same chromosome region, but the underlying mechanism is unclear. Here, we report a cis element that we previously named conserved noncoding sequence 2 (CNS2) physically interacted with both Il17 and Il17f gene promoters and was sufficient for regulating their selective transcription in Th17 cells. Targeted deletion of CNS2 resulted in impaired retinoic acid-related orphan receptor gammat (RORγt)-driven IL-17 expression in vitro. CNS2-deficient T cells also produced substantially decreased amounts of IL-17F. These cytokine defects were associated with defective chromatin remodeling in the Il17-Il17f gene locus, possibly because of effects on CNS2-mediated recruitment of histone-modifying enzymes p300 and JmjC domain-containing protein 3 (JMJD3). CNS2-deficient animals were also shown to be resistant to experimental autoimmune encephalomyelitis (EAE). Our results thus suggest that CNS2 is sufficient and necessary for Il17 and optimal Il17f gene transcription in Th17 cells. [ABSTRACT FROM AUTHOR]

T follicular helper (Tfh) cells play essential roles in regulating humoral immunity, especially germinal center reactions. However, how CD4+ T cells integrate the antigenic and costimulatory signals in Tfh cell development is still poorly understood. Here, we found that phorbol 12-myristate 13-acetate (PMA) + ionomycin (P+I) stimulation, together with interleukin-6 (IL-6), potently induce Tfh cell-like transcriptomic programs in vitro. The ERK kinase pathway was attenuated under P+I stimulation; ERK2 inhibition enhanced Tfh cell development in vitro and in vivo. We observed that inducible T cell costimulator (ICOS), but not CD28, lacked the ability to activate ERK, which was important in sustaining Tfh cell development. The transcription factor Zfp831, whose expression was repressed by ERK, promoted Tfh cell differentiation by directly upregulating the expression of the transcription factors Bcl6 and Tcf7. We have hence identified an ERK-Zfp831 axis, regulated by costimulation signaling, in critical regulation of Tfh cell development. [Display omitted] • PMA + ionomycin promote Tfh cell generation in vitro • ERK kinase pathway negatively regulates Tfh cell differentiation program • CD28 and ICOS signaling differentially regulate ERK during Tfh cell differentiation • Repressed by ERK, Zfp831 is a transcription factor that promotes Tfh cell development Tfh cell essentially regulates humoral immunity, but its differentiation process is not fully understood. Wan et al. reveal that ICOS and CD28 differentially regulate the ERK kinase pathway and ERK is Tfh-repressive. Repressed by ERK, the transcriptional factor Zfp831 promotes T follicular helper cell differentiation. [ABSTRACT FROM AUTHOR]

RORγt is the lineage-specific transcription factor for T helper 17 (Th17) cells whose upregulation in developing Th17 cells is critically regulated by interleukin-6 (IL-6) and TGF-β, the molecular mechanisms of which remain largely unknown. Here we identified conserved non-coding sequences (CNSs) 6 and 9 at the Rorc gene, essential for its expression during Th17 cell differentiation but not required for RORγt expression in innate lymphocytes and γδ T cells. Mechanistically, the IL-6-signal transducer and activator of transcription 3 (STAT3) axis appeared to be largely dependent on CNS9 and only partially on CNS6 in controlling RORγt expression and epigenetic activation of the Rorc locus. TGF-β alone was sufficient to induce RORγt expression in a CNS6- but not CNS9-dependent manner through CNS6 binding by SMAD proteins. Our study reveals an important synergistic mechanism downstream of IL-6 and TGF-β in regulation of RORγt expression and Th17 cell commitment via distinct cis -regulatory elements. • CNS6 and CNS9 are not required for RORγt expression in innate immune cells • Deficiency in CNS6 or CNS9 decreases Th17 cell differentiation and EAE disease • CNS9 and, to a lesser extent, CNS6 are required for IL-6-induced RORγt expression • CNS6, but not CNS9, is indispensable for TGF-β-induced RORγt expression IL-6 and TGF-β induce Th17 cell differentiation. Chang et al. demonstrate that these two cytokines initiate Th17 cell differentiation by inducing de novo Rorc transcription via the distinct conserved non-coding sequences CNS6 and CNS9. [ABSTRACT FROM AUTHOR]