DUSP8 induces TGF-β–stimulated IL-9 transcription and Th9-mediated allergic inflammation by promoting nuclear export of Pur-α
Huai-Chia Chuang, … , Ming-Han Chen, Tse-Hua Tan
Huai-Chia Chuang, … , Ming-Han Chen, Tse-Hua Tan
Published November 1, 2023
Citation Information: J Clin Invest. 2023;133(21):e166269. https://doi.org/10.1172/JCI166269.
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DUSP8 induces TGF-β–stimulated IL-9 transcription and Th9-mediated allergic inflammation by promoting nuclear export of Pur-α

Abstract

Dual-specificity phosphatase 8 (DUSP8) is a MAPK phosphatase that dephosphorylates and inactivates the kinase JNK. DUSP8 is highly expressed in T cells; however, the in vivo role of DUSP8 in T cells remains unclear. Using T cell–specific Dusp8 conditional KO (T-Dusp8 cKO) mice, mass spectrometry analysis, ChIP-Seq, and immune analysis, we found that DUSP8 interacted with Pur-α, stimulated interleukin-9 (IL-9) gene expression, and promoted Th9 differentiation. Mechanistically, DUSP8 dephosphorylated the transcriptional repressor Pur-α upon TGF-β signaling, leading to the nuclear export of Pur-α and subsequent IL-9 transcriptional activation. Furthermore, Il-9 mRNA levels were induced in Pur-α–deficient T cells. In addition, T-Dusp8–cKO mice displayed reduction of IL-9 and Th9-mediated immune responses in the allergic asthma model. Reduction of Il-9 mRNA levels in T cells and allergic responses of T-Dusp8–cKO mice was reversed by Pur-α knockout. Remarkably, DUSP8 protein levels and the DUSP8–Pur-α interaction were indeed increased in the cytoplasm of T cells from people with asthma and patients with atopic dermatitis. Collectively, DUSP8 induces TGF-β–stimulated IL-9 transcription and Th9-induced allergic responses by inhibiting the nuclear translocation of the transcriptional repressor Pur-α. DUSP8 may be a T-cell biomarker and therapeutic target for asthma and atopic dermatitis.

Authors

Huai-Chia Chuang, Chia-Hsin Hsueh, Pu-Ming Hsu, Ching-Yi Tsai, Ying-Chun Shih, Hsien-Yi Chiu, Yi-Ming Chen, Wen-Kuang Yu, Ming-Han Chen, Tse-Hua Tan

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Figure 2

In vitro Th9 differentiation is reduced by Dusp8 cKO.

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In vitro Th9 differentiation is reduced by Dusp8 cKO. (A–G) Flow cytomet...
(A–G) Flow cytometry analyses of Th9 (CD4+IL-9+) (in vitro differentiated from splenic CD4+ T cells, (A), Th9 (CD4+IL-9+) (in vitro differentiated from splenic naive CD4+CD62L+ T cells, (B), Th1 (CD4+IFN-γ+) (C), Th2 (CD4+IL-4+) (D), Th17 (CD4+IL-17A+) (E), and Treg (CD4+Foxp3+) (F) cells of in vitro differentiated T cells. Mean ± SEM are shown. Data (C–F) were not significantly changed in T-Dusp8–cKO T cells. (G) Real-time PCR of Il-9 mRNA levels in T-Dusp8 cKO or WT T cells stimulated with IL-4 (20 ng/mL) and/or TGF-β (50 ng/mL) for 2 or 8 hours. The expression levels of Il-9 were normalized to Srp72 levels. Mean ± SEM are shown. n = 3. *P < 0.05 (2-tailed Student’s t test); §P < 0.05 (1-tailed Student’s t test). (H and I) Real-time PCR of IL-9 mRNA levels in DUSP8 shRNA knocked-down human primary Th9 cells (H) or DUSP8-overexpressing human primary Th9 cells (I). Human primary T cells were cultured under Th9 polarizing conditions for 5 days, followed by transfection with indicated plasmids. The expression levels of IL-9 were normalized to GAPDH levels. Mean ± SEM are shown. n = 3. (J) In vitro phosphatase assays of purified DUSP8 proteins isolated from protein lysates of Myc-DUSP8 or Myc-DUSP8 (C246S) phosphatase-dead mutant-expressing J-TAg T cells stimulated with TGF-β (10 or 20 ng/mL). n = 3. Results (mean ± SEM) are presented relative to those of vector controls. T-Dusp8 cKO, T cell-specific Dusp8 cKO (Dusp8fl/fl;Cd4-Cre); WT (Dusp8fl/fl); DUSP8 (C246S), DUSP8 phosphatase-dead mutant. For (H–J), *P < 0.05; **P < 0.01; ***P < 0.001 (1-way ANOVA and Tukey’s posthoc test). Data shown are representative of 3 independent experiments.