Background/Purpose: Signals that trigger epigenetic changes in CD4⁺ T-cells are unknown. To examine a role for FcγRIIIa mediated co-signal in causing epigenetic changes in human CD4⁺ T-cells. GWAS studies have shown hypomethylation of IFN promoter region in CD4⁺ T-cells. During IFN signaling multiple genome sites are hypomethylated. Procainamide and hydrazine, which inhibit DNA (cytosine-5)-methyltransferase 1 (DNMT1), predispose individuals to drug-induced systemic lupus erythematosus (SLE). We examined whether immune complexes (ICs) by ligating FcγRIIIa on CD4⁺ T-cells cause epigenetic changes. 

Methods:

We activated peripheral human naïve CD4⁺ T-cells using plate bound anti-CD3+anti-CD28 and anti-CD3+ICs+C5b-9. Thereafter, we cultured these cells in T_H17 polarizing conditions. On day nine, we harvested the cells and prepared cDNA. PCR array was performed using chromatin modification array (n=6) (Applied Biosystems) and Human Epigenetic Chromatin Modification Enzyme RT²Profiler PCR array (n=5) (SA Biosciences). We compared the relative change in DNA and histone modifying enzyme genes from ICs+C5b-9 co-stimulation over CD28 co-stimulation. Epigenetic marks were examined by ChiP assay.

Results:

In six donors analyzed by AB array, hierarchical cluster analysis showed a strong correlation among DNMT1, RBBP7, CHD4, and MBD2, which are part of deacetylases complex. All enzymes of this complex were repressed by ICs ligation to CD4⁺ T-cells. A dramatic decrease (4.5 fold) was observed in co-activator associated arginine methyl transferase and nuclear receptor co-activator (NCOA6) (7.14 fold), which remodels chromatin with HAT1. A 7.37-fold increase was observed in activating transcription factor 2 (ATF2), which acetylates H2B and H4 histones during DNA damage response, from ICs ligation of FcγRIIIa on CD4⁺ T-cells compared to CD28 co-stimulation. ATF2/c-Jun regulated IFN-β gene and IFN-γ production. K(lysine) acetyl transferase 6A (KAT6A) was increased 6-fold, which acetylate H3 and H4 histones. KAT6A is a transcriptional co-activator of Runx1 and Runx2. Both T-bet and Runx generate pathogenic IFN-γ producing T_H17 cells. In addition, a 6.16-fold increase in SET domain containing lysine methyltransferases (SETD)-7, a transcription activator that generates H3K4 mark was observed. Also other SET domain proteins SET4, SET8 that generate H4K2me1 mark and SET1 that generates H3K9 mark were upregulated. Relative increase in the expression of ATF2, KAT6A, and SEDT7 was seen in both arrays. 

Conclusion:

Epigenetic modifications contribute to multistep differentiation and plasticity of in vitro and in vivo-derived helper T-cells. Our results show that signal via FcγRIIIa in CD4⁺ T-cells modulates both histones and DNA modifying enzymes. Ligation of FcγRIIIa by ICs lead to the development of IFN-γ^high subset. ATF2 and c-Jun bind to -53 CpG in the IFN-γ promoter in T_H1 and not in T_H2 cells. Persistent signaling from IC ligation to FcγRIIIa by up-regulating ATF2 will continue to generate IFN-γ producing cells.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/fci%c2%b3riiia-mediated-signal-cause-epigenetic-changes-in-human-naive-cd4-t-cells/