JMJD3 regulates CD4+ T cell trafficking by targeting actin cytoskeleton regulatory gene Pdlim4
Chuntang Fu, Qingtian Li, Jia Zou, Changsheng Xing, Mei Luo, Bingnan Yin, Junjun Chu, Jiaming Yu, Xin Liu, Helen Y. Wang, Rong-Fu Wang
Chuntang Fu, Qingtian Li, Jia Zou, Changsheng Xing, Mei Luo, Bingnan Yin, Junjun Chu, Jiaming Yu, Xin Liu, Helen Y. Wang, Rong-Fu Wang
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Research Article Autoimmunity Cell biology Immunology

JMJD3 regulates CD4+ T cell trafficking by targeting actin cytoskeleton regulatory gene Pdlim4

Abstract

Histone H3K27 demethylase JMJD3 plays a critical role in gene expression and T cell differentiation. However, the role and mechanisms of JMJD3 in T cell trafficking remain poorly understood. Here, we show that JMJD3 deficiency in CD4+ T cells resulted in an accumulation of T cells in the thymus and reduction of T cell number in the secondary lymphoid organs. We identified PDLIM4 as a significantly downregulated target gene in JMJD3-deficient CD4+ T cells by gene profiling and ChIP-Seq analyses. We further showed that PDLIM4 functioned as an adaptor protein to interact with sphingosine-1 phosphate receptor 1 (S1P1) and filamentous actin (F-actin), thus serving as a key regulator of T cell trafficking. Mechanistically, JMJD3 bound to the promoter and gene-body regions of the Pdlim4 gene and regulated its expression by interacting with zinc finger transcription factor KLF2. Our findings have identified Pdlim4 as a JMJD3 target gene that affects T cell trafficking by cooperating with S1P1 and have provided insights into the molecular mechanisms by which JMJD3 regulates genes involved in T cell trafficking.

Authors

Chuntang Fu, Qingtian Li, Jia Zou, Changsheng Xing, Mei Luo, Bingnan Yin, Junjun Chu, Jiaming Yu, Xin Liu, Helen Y. Wang, Rong-Fu Wang

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Figure 4

PDLIM4 regulates T cell migration by interaction with S1P1 and modulation of F-actin reorganization.

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PDLIM4 regulates T cell migration by interaction with S1P1 and modulatio...
(A) Immunofluorescence microscopy images of WT and Jmjd3-cKO CD4+ T cells infected with lentivirus containing GFP or PDLIM4-GFP plasmids. Cells were starved 12 hours and treated with 100 nM S1P for 3 hours at 37°C. GFP or PDLIM4-GFP was detected with green fluorescence. Actin filaments were labeled with rhodamine-conjugated phalloidin (red). Nuclei were stained with DAPI (blue). (B) FACS analysis of phalloidin-labeled F-actin in untreated and S1P-treated WT, Jmjd3-cKO, Pdlim4-KO, and Pdlim4-expressing Jmjd3-cKO CD4 SP thymocytes. (C) F-actin (pellet [P]) and G-actin (supernatant [S]) from untreated and treated WT, Jmjd3-cKO CD4 SP thymocytes were detected by Western blot. (D) Immunofluorescence microscopy of untreated and S1P-treated WT CD4+ SP cells stained with FITC-conjugated Abs to detect PDLIM4, Cy5-conjugated Abs to detect S1P1, and rhodamine-conjugated phalloidin to detect actin filaments. Goat IgG and rabbit IgG were used as isotype controls. Merged images indicate colocalization of proteins. (E) Co-IP analysis of endogenous interaction of PDLIM4 with S1P1 in untreated and S1P-treated CD4 SP thymocytes. (F) Cosedimentation assay was performed using GST-PDLIM4, GST-PDLIM4 N-del, or GST-PDLIM4 C-del with F-actin, and subsequent analysis of supernatants and pellets by Western blot analysis. (G) 293T cells were cotransfected with FLAG-Pdlim4-N-del, C-del, or full-length Pdlim4 along with HA-S1p1. WCLs were immunoprecipitated with anti-HA beads and immunoblotted with anti-FLAG Ab. Three independent experiments were repeated with similar results. (H) A schematic diagram of the proposed model showing that PDLIM4 interacts with the S1P1 protein at the N-terminal PDZ domain and binds F-actin by the C-terminal LIM domain.