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The proinflammatory cytokine TNF? induces DNA demethylation-dependent and -independent activation of interleukin-32 expression.

ABSTRACT: IL-32 is a cytokine involved in proinflammatory immune responses to bacterial and viral infections. However, the role of epigenetic events in the regulation of IL-32 gene expression is understudied. Here we show that IL-32 is repressed by DNA methylation in HEK293 cells. Using ChIP sequencing, locus-specific methylation analysis, CRISPR/Cas9-mediated genome editing, and RT-qPCR (quantitative RT-PCR) and immunoblot assays, we found that short-term treatment (a few hours) with the proinflammatory cytokine tumor necrosis factor ? (TNF?) activates IL-32 in a DNA demethylation-independent manner. In contrast, prolonged TNF? treatment (several days) induced DNA demethylation at the promoter and a CpG island in the IL-32 gene in a TET (ten-eleven translocation) family enzyme- and NF-?B-dependent manner. Notably, the hypomethylation status of transcriptional regulatory elements in IL-32 was maintained for a long time (several weeks), causing elevated IL-32 expression even in the absence of TNF?. Considering that IL-32 can, in turn, induce TNF? expression, we speculate that such feedforward events may contribute to the transition from an acute inflammatory response to chronic inflammation.

SUBMITTER: Zhao Z

PROVIDER: S-EPMC6497958 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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The proinflammatory cytokine TNFα induces DNA demethylation-dependent and -independent activation of interleukin-32 expression.

Zhao Zuodong Z Lan Mengying M Li Jingjing J Dong Qiang Q Li Xiang X Liu Baodong B Li Gang G Wang Hailin H Zhang Zhuqiang Z Zhu Bing B

The Journal of biological chemistry 20190301 17

IL-32 is a cytokine involved in proinflammatory immune responses to bacterial and viral infections. However, the role of epigenetic events in the regulation of IL-32 gene expression is understudied. Here we show that IL-32 is repressed by DNA methylation in HEK293 cells. Using ChIP sequencing, locus-specific methylation analysis, CRISPR/Cas9-mediated genome editing, and RT-qPCR (quantitative RT-PCR) and immunoblot assays, we found that short-term treatment (a few hours) with the pr ...[more]

PMID: 30824537

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Project description:PURPOSE:Cold air is a major environmental factor that exacerbates asthma. Transient receptor potential melastatin family member 8 (TRPM8) is a cold-sensing channel expressed in the airway epithelium. However, its role in airway inflammation remains unknown. We investigated the role of TRPM8 in innate immune responses in bronchial epithelial cells and asthmatic subjects. METHODS:The TRPM8 mRNA and protein expression on BEAS2B human bronchial epithelial cells was examined by real-time polymerase chain reaction (PCR), immunofluorescence staining and western blotting. Additionally, interleukin (IL)-4, IL-6, IL-8, IL-13, IL-25 and thymic stromal lymphopoietin (TSLP) levels before and after menthol, dexamethasone and N-(4-tert-butylphenyl)-4-(3-chloropyridin-2-yl) piperazine-1-carboxamide (BCTC) treatments were measured via real-time PCR. TRPM8 protein levels in the supernatants of induced sputum from asthmatic subjects and normal control subjects were measured using enzyme-linked immunosorbent assay, and mRNA levels in sputum cell lysates were measured using real-time PCR. RESULTS:Treatment with up to 2 mM menthol dose-dependently increased TRPM8 mRNA and protein in BEAS2B cells compared to untreated cells (P < 0.001) and concomitantly increased IL-25 and TSLP mRNA (P < 0.05), but not IL-33 mRNA. BCTC (10 ?M) significantly abolished menthol-induced up-regulation of TRPM8 mRNA and protein and IL-25 and TSLP mRNA (P < 0.01). TRPM8 protein levels were higher in the supernatants of induced sputum from asthmatic subjects (n = 107) than in those from healthy controls (n = 19) (P < 0.001), and IL-25, TSLP and IL-33 mRNA levels were concomitantly increased (P < 0.001). Additionally, TRPM8 mRNA levels correlated strongly with those of IL-25 and TSLP (P < 0.001), and TRPM8 protein levels were significantly higher in bronchodilator-responsive asthmatic subjects than in nonresponders. CONCLUSIONS:TRPM8 may be involved in the airway epithelial cell innate immune response and a molecular target for the treatment of asthma.

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The proinflammatory cytokine TNF? induces DNA demethylation-dependent and -independent activation of interleukin-32 expression.

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The proinflammatory cytokine TNFα induces DNA demethylation-dependent and -independent activation of <i>interleukin-32</i> expression.

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