Histone acetylation plays an important role in transcriptional activation. We have investigated the effect of ethanol on nuclear histone H3 acetylation in rat hepatocytes. Hepatocytes were incubated with ethanol (5-200 mM) for 24 h and then acetylation states of nuclear histone H3 at specific lysine residues (Lys(9) and Lys(14)) were measured by immunoblot analysis using site-specific antibodies. Ethanol increased acetylation of histone H3 at Lys(9) in a dose-dependent manner; 3-fold at 5 mM and maximum of 8-fold at 100 mM. Sensitivity to low dose of ethanol was remarkable. This ethanol-induced acetylation was also time-dependent, showing a maximal response at 24 h. Ethanol did not alter the level of histone H3 expression. Trichostatin A, a histone deacetylase inhibitor, was used as a positive control and it also increased acetylation. However, acetylation at Lys(14) was not affected by ethanol. Treatment of cells with ethanol metabolizing enzyme inhibitors (4-methylpyrazole and cyanamide) decreased ethanol-induced histone H3 acetylation at Lys(9). This is the first report of ethanol-induced selective, post-translational acetylation of histone H3 at Lys(9). This is not due to increased histone expression or a direct physical effect of ethanol but is dependent on ethanol metabolism. (C) 2003 Elsevier Science (USA). All rights reserved.