Histone methyltransferase G9a regulates expression of nuclear receptors and cytochrome P450 enzymes in HepaRG cells at basal level and in fatty acid induced steatosis

Abstract

Obesity and nonalcoholic fatty liver disease (NAFLD) affect expression and function of cytochrome P450 genes (P450s). The increased expression of inflammatory cytokines is a major driver of the downregulation of P450 expression in NAFLD. Decrease in P450 expression could potentially lead to drug-drug interaction, inefficient pharmacological effect of a drug, or hepatotoxicity. An epigenetic modifier, histone 3 lysine 9 methyl transferase enzyme (G9a), known to increase histone 3 lysine 9 methylation, is downregulated in diet-induced obesity animal models. In a liver-specific G9a knockout animal model, expression of P450s was downregulated. Currently, the role of G9a in regulation of P450s in steatosis is unknown. Our hypothesis is that in steatosis G9a plays a role in downregulation of P450 expression. In this study, we used HepaRG cells to induce steatosis using a combination of free fatty acids oleic acid and palmitic acid. The G9a was knocked down and overexpressed using small interfering RNA and adenovirus mediated approaches, respectively. Knockdown and overexpression of G9a in the absence of steatosis decreased and increased expression of nuclear receptors constitutive androstane receptor (CAR), pregnane X receptor, small heterodimer partner, and CYP2B6, 2E1, 2C8, 2C9, and 3A4, respectively. In steatotic conditions, overexpression of G9a prevented fatty acid mediated decreased expression of CAR, CYP2C19, 2C8, 7A1, and 3A4. Our current study suggests that G9a might serve as a key regulator of P450 expression at both the basal level and in early steatotic conditions. Single nucleotide polymorphism of G9a leading to loss/gain of function could lead to the poor metabolizer or ultrarapid metabolizer phenotypes. SIGNIFICANCE STATEMENT The current study demonstrates that histone modification enzyme G9a is involved in the regulation of expression of nuclear receptors constitutive androstane receptor, pregnane X receptor, and small heterodimer partner as well as drug-metabolizing cytochrome P450s (P450s) at basal conditions and in fatty acid induced cellular model of steatosis. Histone 3 lysine 9 methylation should be considered together with histone 3 lysine 4 and histone 3 lysine 27 methylation as the epigenetic mechanisms controlling gene expression of P450s.