Phosphorylation of IRF4 by ROCK2 regulates IL-17 and IL-21 production and the development of autoimmunity in mice
Partha S. Biswas, … , Govind Bhagat, Alessandra B. Pernis
Partha S. Biswas, … , Govind Bhagat, Alessandra B. Pernis
Published August 9, 2010
Citation Information: J Clin Invest. 2010;120(9):3280-3295. https://doi.org/10.1172/JCI42856.
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Research Article Autoimmunity

Phosphorylation of IRF4 by ROCK2 regulates IL-17 and IL-21 production and the development of autoimmunity in mice

Abstract

Deregulated production of IL-17 and IL-21 plays a key pathogenic role in many autoimmune disorders. A delineation of the mechanisms that underlie the inappropriate synthesis of IL-17 and IL-21 in autoimmune diseases can thus provide important insights into potential therapies for these disorders. Here we have shown that the serine-threonine kinase Rho-associated, coiled-coil–containing protein kinase 2 (ROCK2) becomes activated in mouse T cells under Th17 skewing conditions and phosphorylates interferon regulatory factor 4 (IRF4), a transcription factor that is absolutely required for the production of IL-17 and IL-21. We furthermore demonstrated that ROCK2-mediated phosphorylation of IRF4 regulated the synthesis of IL-17 and IL-21 and the differentiation of Th17 cells. Whereas CD4+ T cells from WT mice activated ROCK2 physiologically under Th17 conditions, CD4+ T cells from 2 different mouse models of spontaneous autoimmunity aberrantly activated ROCK2 under neutral conditions. Moreover, administration of ROCK inhibitors ameliorated the deregulated production of IL-17 and IL-21 and the inflammatory and autoantibody responses observed in these autoimmune mice. Our findings thus uncover a crucial link among ROCK2, IRF4, and the production of IL-17 and IL-21 and support the idea that selective inhibition of ROCK2 could represent an important therapeutic regimen for the treatment of autoimmune disorders.

Authors

Partha S. Biswas, Sanjay Gupta, Emily Chang, Li Song, Roslynn A. Stirzaker, James K. Liao, Govind Bhagat, Alessandra B. Pernis

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Figure 1

Enhanced ROCK2 activation in the absence of Def6.

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Enhanced ROCK2 activation in the absence of Def6. (A and B) Purified CD4...
(A and B) Purified CD4+ T cells from Def6+/+ or Def6trap/trap mice were either left unstimulated (unst) or restimulated (st) with anti-CD3 and anti-CD28 mAbs for 48 hours in the presence or absence of 30 μM Y27632 [Y(30)]. (A) ROCK2 kinase activity was measured by in vitro kinase assay using [γ-32P]ATP. Autophosphorylated ROCK2 was detected by autoradiography. (B) ROCK2 kinase activity was assayed by incubating immunoprecipitated ROCK2 with purified rMYPT1 as ROCK2 substrate, and pMYPT1 was detected by Western blot analysis using anti-pMYPT1 Ab. In A and B, total ROCK2 levels in the input samples are shown below. In B, lanes were run on the same gel but were noncontiguous (white line). (C) Levels of pERM in CD4+CD44intCD62Lhi, CD4+CD44hiCD62Llo, B220+, and CD11b+ cells from Def6+/+DO11.10 or Def6trap/trapDO11.10 mice were detected by intracellular staining with anti-pERM Ab and analyzed by FACS. Solid histograms indicate isotype control. Shown are overlaid histograms after gating on the appropriate populations. (A–C) Data are representative of 3 independent experiments.