TET2 regulates immune tolerance in chronically activated mast cells
Riccardo Rigo, Rabie Chelbi, Julie Agopian, Sebastien Letard, Aurélien Griffon, Hussein Ghamlouch, Julien Vernerey, Vasileios Ladopoulos, Edwige Voisset, Paulo De Sepulveda, Geoffrey Guittard, Jacques A. Nunès, Ghislain Bidaut, Berthold Göttgens, Michael Weber, Olivier A. Bernard, Patrice Dubreuil, Erinn Soucie
Riccardo Rigo, Rabie Chelbi, Julie Agopian, Sebastien Letard, Aurélien Griffon, Hussein Ghamlouch, Julien Vernerey, Vasileios Ladopoulos, Edwige Voisset, Paulo De Sepulveda, Geoffrey Guittard, Jacques A. Nunès, Ghislain Bidaut, Berthold Göttgens, Michael Weber, Olivier A. Bernard, Patrice Dubreuil, Erinn Soucie
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Research Article Inflammation

TET2 regulates immune tolerance in chronically activated mast cells

Abstract

Mutation of the TET2 DNA-hydroxymethylase has been associated with a number of immune pathologies. The disparity in phenotype and clinical presentation among these pathologies leads to questions regarding the role of TET2 mutation in promoting disease evolution in different immune cell types. Here we show that, in primary mast cells, Tet2 expression is induced in response to chronic and acute activation signals. In TET2-deficient mast cells, chronic activation via the oncogenic KITD816V allele associated with mastocytosis, selects for a specific epigenetic signature characterized by hypermethylated DNA regions (HMR) at immune response genes. H3K27ac and transcription factor binding is consistent with priming or more open chromatin at both HMR and non-HMR in proximity to immune genes in these cells, and this signature coincides with increased pathological inflammation signals. HMR are also associated with a subset of immune genes that are direct targets of TET2 and repressed in TET2-deficient cells. Repression of these genes results in immune tolerance to acute stimulation that can be rescued with vitamin C treatment or reiterated with a Tet inhibitor. Overall, our data support a model where TET2 plays a direct role in preventing immune tolerance in chronically activated mast cells, supporting TET2 as a viable target to reprogram the innate immune response for innovative therapies.

Authors

Riccardo Rigo, Rabie Chelbi, Julie Agopian, Sebastien Letard, Aurélien Griffon, Hussein Ghamlouch, Julien Vernerey, Vasileios Ladopoulos, Edwige Voisset, Paulo De Sepulveda, Geoffrey Guittard, Jacques A. Nunès, Ghislain Bidaut, Berthold Göttgens, Michael Weber, Olivier A. Bernard, Patrice Dubreuil, Erinn Soucie

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Figure 7

TET2 is required to directly regulate mast cell activation genes and rescue acute mast cell activation.

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TET2 is required to directly regulate mast cell activation genes and res...
(A) ChIP-qPCR analyses showing enrichment in signal in TET2 ChIP over ChIP using a nonspecific IgG antibody at HMR and at regions 4 kb upstream the HMR in Tet2+/+KITD816V (gray) and Tet2–/–KITD816V cells (blue; negative control for TET2 antibody). Each point is derived from 3 independent mast cell cultures for each genotype that were pooled at the time of assay. Graphs show the combined results from 2 independent experiments (mean ± SEM). (B) Expression levels of genes relative to Hprt1 in untreated Tet2+/+KITD816V and Tet2 +/–KITD816V mast cells or after 24 hours of treatment with 250 μM vitamin C (n = 3; mean ± SD; the means of all groups were significantly different by 1-way ANOVA; *P < 0.05, **P < 0.01, ***P < 0.001 for Dunnett’s multiple-comparison test, each treatment versus control, or unpaired, 2-tailed t test to compare Tet2 +/–KITD816V with or without vitamin C. Only significant differences are shown in the figure). (C) Representative methylation profile of HMR associated regions in Tet2+/+ KITD816V and Tet2 +/–KITD816V mast cells obtained by performing bisulfite sequencing. Samples were untreated or treated for 24 hours with 250 μM vitamin C. Black circles represent methylated CpG dinucleotide, while gray and white circles indicate partially methylated or unmethylated CpGs, respectively (mean methylation of n = 3). (D) Enrichment of 5hmC at HMR associated loci in Tet2 +/–KITD816V mast cells with or without vitamin C treatment (250 μM for 24 hours) evaluated by using hydroxy-methyl DNA immunoprecipitation (hMeDIP) (n = 3; mean ± SD; *P < 0.05, **P < 0.01 unpaired, 2-tailed t test). (E) Quantification analysis for acute mast cell activation assay measuring TNF and IL6 levels by intracellular staining. Cells were pretreated for 24 hours with 250 μM vitamin C, prior to acute mast cell activation assay using 10 ng/mL IL10 or 0.5 μg/mL LPS as in Figure 5A. Fold change is relative to control for each cell population. Each point is derived from 3 independent mast cell cultures for each genotype that were pooled at the time of assay. Graph show the combined results from 2 independent experiments (mean ± SD; **P < 0.01 unpaired, 2-tailed t test).