A central role for G9a and EZH2 in the epigenetic silencing of cyclooxygenase‐2 in idiopathic pulmonary fibrosis

ABSTRACT Selective silencing of the cyclooxygenase‐2 (COX‐2) gene with the loss of the antifibrotic mediator prostaglandin E2 contributes to the fibrotic process in idiopathic pulmonary fibrosis (IPF). This study explored the role of G9a‐ and enhancer of zeste homolog 2 (EZH2)‐mediated methylation o...

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Published in:The FASEB journal 2014-07, Vol.28 (7), p.3183-3196
Main Authors: Coward, William R., Feghali‐Bostwick, Carol A., Jenkins, Gisli, Knox, Alan J., Pang, Linhua
Format: Article
Language:English
Subjects:
Acetylation
Adult
Aged
antifibrotic gene
Cells, Cultured
Chromatin Immunoprecipitation - methods
Cyclooxygenase 2 - genetics
Cyclooxygenase 2 - metabolism
DNA methylation
DNA Methylation - genetics
Enhancer of Zeste Homolog 2 Protein
Epigenesis, Genetic - genetics
Fibroblasts - metabolism
Gene Silencing - physiology
Histocompatibility Antigens - genetics
Histocompatibility Antigens - metabolism
histone deacetylation
histone hypermethylation
Histone-Lysine N-Methyltransferase - genetics
Histone-Lysine N-Methyltransferase - metabolism
Histones - genetics
Histones - metabolism
Humans
Idiopathic Pulmonary Fibrosis - genetics
Idiopathic Pulmonary Fibrosis - metabolism
lung fibroblasts
Male
Middle Aged
Polycomb Repressive Complex 2 - genetics
Polycomb Repressive Complex 2 - metabolism
Promoter Regions, Genetic - genetics
Research Communications
RNA, Messenger - genetics
Young Adult
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Summary:ABSTRACT Selective silencing of the cyclooxygenase‐2 (COX‐2) gene with the loss of the antifibrotic mediator prostaglandin E2 contributes to the fibrotic process in idiopathic pulmonary fibrosis (IPF). This study explored the role of G9a‐ and enhancer of zeste homolog 2 (EZH2)‐mediated methylation of histone H3 lysine 9 (H3K9me3) and histone H3 lysine 27 (H3K27me3) in COX‐2 silencing in IPF. Chromatin immunoprecipitation (ChIP) and re‐ChIP assays demonstrated marked increases in H3K9me3, H3K27me3, and DNA methylation, together with their respective modifying enzymes G9a, EZH2, and DNA methyltransferases (Dnmts) and respective binding proteins heterochromatin protein 1 (HP1), polycomb protein complex 1 (PRC1) and methyl CpG binding protein 2 (MeCP2), at the COX‐2 promoter in lung fibroblasts from patients with IPF (F‐IPFs) compared with fibroblasts from nonfibrotic lungs. HP1, EZH2, and MeCP2 in turn were associated with additional repressive chromatin modifiers in F‐IPFs. G9a and EZH2 inhibitors and small interfering RNAs and the Dnmt1 inhibitor markedly reduced H3K9me3 (49–79%), H3K27me3 (44–81%), and DNA methylation (61–97%) at the COX‐2 promoter. These reductions were correlated with increased histone H3 and H4 acetylation, resulting in COX‐2 mRNA and protein reexpression in F‐IPFs. Our results support a central role for G9a‐ and EZH2‐mediated histone hypermethylation and a model of bidirectional, mutually reinforcing, and interdependent crosstalk between histone hypermethylation and DNA methylation in COX‐2 epigenetic silencing in IPF.—Coward, W. R., Feghali‐Bostwick, C. A., Jenkins, G., Knox, A. J., Pang, L. A central role for G9a and EZH2 in the epigenetic silencing of cyclooxygenase‐2 in idiopathic pulmonary fibrosis. FASEB J. 28, 3183–3196 (2014). www.fasebj.org
ISSN:0892-6638
1530-6860
DOI:10.1096/fj.13-241760