Decreased RORC-dependent silencing of prostaglandin receptor EP2 induces autoimmune Th17 cells
David M. Kofler, … , Vijay K. Kuchroo, David A. Hafler
David M. Kofler, … , Vijay K. Kuchroo, David A. Hafler
Published May 8, 2014
Citation Information: J Clin Invest. 2014;124(6):2513-2522. https://doi.org/10.1172/JCI72973.
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Research Article Immunology

Decreased RORC-dependent silencing of prostaglandin receptor EP2 induces autoimmune Th17 cells

Abstract

Prostaglandin E2 (PGE2) promotes Th17 expansion while otherwise inhibiting other CD4+ T cell subsets. Here, we identified a PGE2-dependent pathway that induces pathogenic Th17 cells in autoimmune disease and is regulated by the transcription factor RORC. Compared with other CD4+ cell types from healthy subjects, there is a surprising lack of the prostaglandin receptor EP2 on Th17 cells; therefore, we examined the hypothesis that RORγt, which is highly expressed in Th17 cells, mediates EP2 downregulation. Chromatin immunoprecipitation followed by DNA sequencing revealed that RORγt binds directly to Ptger2 (the gene encoding EP2 receptor) in Th17 cells isolated from WT mice. In Th17 cells isolated from humans, RORC repressed EP2 by directly silencing PTGER2 transcription, and knock down of RORC restored EP2 expression in Th17 cells. Compared with Th17 cells from healthy individuals, Th17 cells from patients with MS exhibited reduced RORC binding to the PTGER2 promoter region, resulting in higher EP2 levels and increased expression of IFN-γ and GM-CSF. Finally, overexpression of EP2 in Th17 cells from healthy individuals induced a specific program of inflammatory gene transcription that produced a pathogenic Th17 cell phenotype. These findings reveal that RORC directly regulates the effects of PGE2 on Th17 cells, and dysfunction of this pathway induces a pathogenic Th17 cell phenotype.

Authors

David M. Kofler, Alexander Marson, Margarita Dominguez-Villar, Sheng Xiao, Vijay K. Kuchroo, David A. Hafler

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Figure 1

Prostaglandin receptor expression in human CD4+ T cell subsets.

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Prostaglandin receptor expression in human CD4+ T cell subsets. (A) Quan...
(A) Quantitative RT-PCR analysis of EP1, EP2, EP3, and EP4 expression in distinct CD4+ T cell subsets. P was calculated by 1-way ANOVA test. n = 10 individual donors. CD4+ subsets were differentiated from naive CD4+ cells in the presence of 2.5 μg/ml anti-CD3 antibodies, 1 μg/ml anti-CD28 antibodies, and the respective cytokines required for the induction of specific CD4+ subsets. Representative example (B) and cumulative results (C) of flow cytometric analysis of EP2 expression in Th0, Th1, Th2, Th9, Th17, and Tregs. P was calculated by 1-way ANOVA test. n = 8 individual donors. (D) IL-17A, RORC, and EP2 expression in ex vivo–isolated IL-17+CD4+ T cells and IL-17–CD4+ T cells. Cells were sorted by FACSAria based on cell surface expression of IL-17. ***P < 0.001. P was calculated by an unpaired Student’s t test; mean ± SEM. n = 10 individual donors. (E) Representative example of flow cytometric analysis of EP2 expression in IFN-γ+ and IL-17+ memory CD4+ T cells. Cells were analyzed for EP2 expression directly after ex vivo isolation without prior purification by FACSAria cell sorting.