Ubiquitin specific peptidase 4 stabilizes interferon regulatory factor protein and promotes its function to facilitate interleukin-4 expression in T helper type 2 cells

Guo,Zhixiang; Xu,Peng; Ge,Shangqing; Zhang,Chengxin; Zheng,Xiaoyan; Xu,Jinguo; Liu,Zhuang; Li,Bin; Ge,Shenglin

doi:10.3892/ijmm.2017.3087

Ubiquitin specific peptidase 4 stabilizes interferon regulatory factor protein and promotes its function to facilitate interleukin-4 expression in T helper type 2 cells

Authors:
- Zhixiang Guo
- Peng Xu
- Shangqing Ge
- Chengxin Zhang
- Xiaoyan Zheng
- Jinguo Xu
- Zhuang Liu
- Bin Li
- Shenglin Ge
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Affiliations: Department of Cardiovascular Surgery, The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui 230022, P.R. China, Department of Oncology, The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui 230022, P.R. China, Key Laboratory of Molecular Virology and Immunology, Unit of Molecular Immunology, Institute Pasteur of Shanghai, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200025, P.R. China
Published online on: August 3, 2017 https://doi.org/10.3892/ijmm.2017.3087
Pages: 979-986
Copyright: © Guo et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

We speculated that ubiquitin specific peptidase 4 (USP4) may deubiquitinate interferon regulatory factor 4 (IRF4) and affect T helper type 2 (Th2) cell function. This study aimed to validate this hypothesis. Here, the interaction between USP4 and IRF4 were analyzed by co-immunoprecipitation assay. The deubiquitin effect of USP4 on IRF4 was analyzed by the Ni-NTA pull down assay. Luciferase reporter gene constructs were used to analyze the effects of USP4, IRF4 and nuclear factor of activated T cell-2 (NFATc2) on the activation of the interleukin-4 (IL-4) promoter. Then, the Th2 cells were infected with sh-USP4 to analyze the effects of USP4 on the expression levels of IRF4 and Th2-related cytokines. Western blotting and RT-qPCR were used to detect the protein and mRNA expression levels, respectively. To determine the levels of IL-4 and IRF4 in rheumatic heart disease (RHD) patients, peripheral blood mononuclear cells (PBMCs) were separated by density gradient centrifugation from RHD patients and healthy controls, and flow cytometric analysis was performed. Our results validated the interaction between USP4 and IRF4, and effects of USP4 on stabilization and deubiquitination of IRF4 were also found. Importantly, USP4 and IRF4 synergized with NFATc2 to specifically enhance NFAT-mediated activation of the IL-4 promoter. USP4 knockdown not only decreased the expression level of IRF4, but also affected the expression level of Th2-related cytokines. Finally, the increased level of IL-4 and IRF4 in PBMCs of RHD patients were observed. On the whole, our data indicate that USP4 interacts with and deubiquitinates IRF4, and also stabilizes IRF4 protein and promotes IRF4 function to facilitate IL-4 expression in Th2 cells, which may be related to the pathological process of RHD.

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