Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: The highly expressed TTP was identified as a crucial factor in response to the IgE-mediated inflammatory syndrome. A , the robust rank aggregation (RRA) algorithm integrated upregulated genes (Log 2 Foldchange>1 and p < 0.05) from three datasets of three IgE-mediated inflammatory syndromes (allergic rhinitis, asthma, and atopic dermatitis) and their corresponding normal samples. B , the ranking graph displayed the genes that were upregulated in at least two datasets. Among them, the genes corresponding to the red dots (Frequency = 3) showed a significant increase in all three disease datasets, including TTP. C , intersection of the genes encoding CCCH-ZF (zinc finger)-type proteins with the genes upregulated in the nasal mucosa of patients with allergic rhinitis. D , the assessment of evolutionary conservation of amino acid positions, especially two CCCH-ZF repeat regions, in the mouse TTP protein. Conservation scores ranged from 1 to 9, where one was the most highly variable, five was of intermediate conservation, and nine was the most highly conserved position. E , experimental scheme of ragweed pollen (RW)-induced AR model, divided into the sensitization stage (0 and 7 days) and the stimulation stage (14–17 days). The control mice were treated with PBS instead of RW (N = 6 per group). F , relative RNA expressions of TTP in nasal tissues of RW-induced AR mice and control mice. G , compare the frequencies of sneezing and nose-rubbing within 10 min between control mice and AR mice after 14 days. H , immunofluorescence staining of TTP protein ( red ) and CD4 protein ( green ) was performed in normal and RW nasal tissues. DAPI ( blue ) was used for nuclear staining (Scale bar = 50 μm). I , quantitative analyses of CD4 + T cell percentages and TTP fluorescence intensity in nasal mucosal tissues from sham and AR model mice. Data are shown as means ± SD (N = 6), Student's t test was used for statistical analysis of control mice and AR mice.

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Control, Immunofluorescence, Staining, Fluorescence

Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: TTP suppressed T helper 2 (Th2)-type inflammation in AR mice. A , schematics of splenocytes isolation from AR-mice and the differentiation of splenic naive CD4+ T cells model in vitro . B , the levels of Interleukin (IL)-4, IL-5, and IL-13, which belong to the Th2 cytokine family, were measured by ELISA assays in the cell supernatant. The cell supernatant was collected from mononuclear cells of the spleen that had been treated with ionomycin and phorbol 12-myristate 13-acetate (PMA) for 6 h. C , after treatment with ionomycin, PMA, and GolgiPlug, the cells with the double stain of anti-CD4 and anti-IL-4-APC were detected by flow cytometry to analyze the percentage of Th2 cells. Data are shown as means ± SD (N = 3 or 6). One-way analysis of variance (ANOVA) was used for statistical analysis of control mice, AR mice, and AR mice transduced with control vectors or TTP overexpression vectors.

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Isolation, In Vitro, Enzyme-linked Immunosorbent Assay, Staining, Flow Cytometry, Control, Transduction, Over Expression

Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: TTP overexpression inhibited Th2 differentiation in vitro . A , schematics of naive CD4+ isolation, activation, and Th2 differentiation. B , a representative flow cytometric plot and the percentage of Th2 cells was detected using flow cytometry. C , the protein and mRNA expression of TTP in CD4+ T cells. D , the protein and mRNA expression of TTP in Th2 cells after virus infection. E , the levels of IL-5 were detected using real-time PCR and ELISA, respectively, in Th2 cells after virus infection. F , the percentage of Th2 cells was detected using flow cytometry in Th2 cells after virus infection. Data are shown as means ± SD (N = 3). Student's t test and one-way ANOVA were used for statistical analysis.

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Over Expression, In Vitro, Isolation, Activation Assay, Flow Cytometry, Expressing, Virus, Infection, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: TTP knockdown exacerbated allergic phenotypes and promoted Th2-type inflammation in AR mice. A , experimental scheme of RW-induced AR mice with or without TTP knockdown via intravenous injection of lentiviral vectors (Lv-shNC or Lv-shTTP). B , expression of TTP mRNA in AR mice with or without TTP knockdown. C , immunofluorescence staining for TTP protein ( red ) and CD4 protein ( green ) in normal and RW-exposed nasal tissues. DAPI ( blue ) was used for nuclear staining (Scale bar = 50 μm). D , quantitative analysis of TTP fluorescence intensity in nasal mucosal tissues from lentivirus-injected mice. E , hematoxylin-eosin (H&E)-stained nasal mucosal tissue sections, showing nasal mucosa thickness (Scale bar = 100 μm). Dashed lines indicate the boundary between the epithelial layer and lamina propria. F and G , changes in the number of sneezes and nasal rubbings after TTP knockdown in AR mice. H , schematic of splenocyte isolation from AR mice and in vitro treatment. I – K , Levels of IL-4, IL-5, and IL-13 in the cell culture supernatant of splenocytes treated with ionomycin and PMA for 6 h. Data are presented as means ± SD (N = 3 or 6). Student's t test was used for statistical analysis of AR mice transduced with shNC or shTTP.

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Knockdown, Injection, Expressing, Immunofluorescence, Staining, Fluorescence, Isolation, In Vitro, Cell Culture, Transduction

Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: Transcriptional profiles of CD4+ T cell isolated from AR mouse spleen reveal categories of genes associated with TTP. A , RNA transcriptome microarray analysis of CD4+ T cell isolated from mouse. B , a volcano plot of all DEGs between CD4+ T cells isolated from LV-TTP and LV-Vector infected AR mouse. C , GSEA analysis identified significant enrichment of signaling pathways based on KEGG and GO analysis following TTP overexpression. D , KEGG pathway enrichment and GO functional classification analysis of DEGs. Data are shown as means ± SD (N = 3).

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Isolation, Microarray, Plasmid Preparation, Infection, Protein-Protein interactions, Over Expression, Functional Assay

Journal: The Journal of Biological Chemistry

Article Title: RNA-binding protein tristetraprolin inhibits Th2 cell activation and differentiation in allergic rhinitis by promoting TRIM18 mRNA decay

doi: 10.1016/j.jbc.2026.111240

Figure Lengend Snippet: The TTP protein acted as a negative regulator of TRIM18-mediated Th2-type inflammation by reducing TRIM18 mRNA stability. A , protein–protein interaction network of 16 potential substrates of E3 ubiquitin ligase TRIM18, constructed using the STRING database and supplemented with functional annotations via GO (Gene Ontology) pathway enrichment analyses, highlighting key biological processes and signaling pathways associated with these substrates. B and C , the qRT-PCR and ELISA were performed to detect the mRNA level and secreted protein concentration of IL-5, respectively, in naive CD4+ T cells or in vitro differentiated Th2 cells transfected with TRIM18 overexpression vector or empty vector. D , schematic diagram of the experimental workflow: Naive CD4+ T cells were induced to differentiate into Th2 cells under in vitro and were co-transfected with combinations of TTP overexpression vector, TRIM18 overexpression vector, or corresponding empty vectors. E , the mRNA level of TRIM18 in differentiated Th2 cells from the three groups (Control, TTP-OE, TTP+TRIM18-OE) was detected by qRT-PCR assays. F and G , The mRNA level and secreted protein concentration of IL-5 were measured by qRT-PCR and ELISA, respectively, in differentiated Th2 cells from the three groups (Control, TTP-OE, TTP+TRIM18-OE). H , flow cytometric analysis of the percentage of CD4+IL-4+ Th2 cells among the three groups after in vitro differentiation, and the proportion of double-positive cells was quantified to assess the impact of TTP and/or TRIM18 overexpression on Th2 cell differentiation. Data are shown as means ± SD (N = 3), One-way ANOVA was used for statistical analysis.

Article Snippet: Then, the sections were co-incubated with antibodies against TTP (1:100 dilution, Proteintech, Cat No.66938-1-Ig) and CD4 (1:50 dilution, Abclonal, Cat No.A0363) overnight at 4 °C.

Techniques: Ubiquitin Proteomics, Construct, Functional Assay, Protein-Protein interactions, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Protein Concentration, In Vitro, Transfection, Over Expression, Plasmid Preparation, Control, Cell Differentiation