Abstract
Modification of histones by lysine methylation plays a role in many biological processes, and it is dynamically regulated by several histone methyltransferases and demethylases. The polycomb repressive complex contains the H3K27 methyltransferase EZH2 and controls dimethylation and trimethylation of H3K27 (H3K27me2/3), which trigger gene suppression. JMJD3 and UTX have been identified as H3K27 demethylases that catalyze the demethylation of H3K27me2/3, which in turns lead to gene transcriptional activation. EZH2, JMJD3 and UTX have been extensively studied for their involvement in development, immune system, neurodegenerative disease, and cancer. However, their role in molecular mechanisms underlying the differentiation process of hepatic cells is yet to be elucidated. Here, we show that EZH2 methyltransferase and JMJD3/UTX demethylases were deregulated during hepatic differentiation of human HepaRG cells resulting in a strong reduction of H3K27 methylation levels. Inhibition of JMJD3 and UTX H3K27 demethylase activity by GSK-J4 epi-drug reverted phenotype of HepaRG DMSO-differentiated cells and human primary hepatocytes, drastically decreasing expression of hepatic markers and inducing cell proliferation. In parallel, inhibition of EZH2 H3K27me3 activity by GSK-126 epi-drug induced upregulation of hepatic markers and downregulated the expression of cell cycle inhibitor genes. To conclude, we demonstrated that modulation of H3K27 methylation by inhibiting methyl-transferase and dimethyl-transferase activity influences the differentiation status of hepatic cells, identifying a possible new role of EZH2, JMJD3 and UTX epi-drugs to modulate hepatic cell plasticity.
Highlights
Chromatin remodeling represents a highly dynamic and reversible process in which there is continual laying down and removal of modifications of histones Nterminal tails by chromatin-remodeling enzymes
Enhancer of Zeste Homolog 2 (EZH2), JMJD3, and UTX are modulated during hepatic differentiation In order to investigate the role of histone methylation, during the process of hepatic differentiation, we first evaluated protein and transcript levels of methyltransferase EZH2 and demethylases JMJD3 and UTX in differentiating HepaRG cells
In cells differentiated for 14 days (DM) transcript levels of hepatic genes Cyp3A4, Albumin, Cyp2E1, E-cadherin and HNF4 were upregulated as compared to proliferating cells (GM) (Fig. 1b)
Summary
Chromatin remodeling represents a highly dynamic and reversible process in which there is continual laying down and removal of modifications of histones Nterminal tails by chromatin-remodeling enzymes. The N-terminal tails of histones contain lysine (K) and arginine (R) residues that can undergo different posttranslational modifications. Enhancer of Zeste Homolog 2 (EZH2) methyltransferase is a component of Polycomb Repressive Complex 2 (PRC2) complex and functions as a histone methyltransferase that induces H3K27me[3] to the targeted genes. PRC2 has been shown to deregulate gene expression promoting cancer cell growth and Official journal of the Cell Death Differentiation Association. It has been shown that EZH2 is essential for expansion of hepatic progenitor population and its loss of function results in decreased expression of hepatic differentiation marker genes[6,7]
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