Histone acetylation by ethanol:Mechanisms & consequence

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DESCRIPTION (provided by applicant): Post-translational modifications in histones are emerging as important mechanisms for the transcriptional activation and gene silencing. We have recently made the original observation that ethanol preferentially caused a spectacular (8-10 fold) increase in acetylation of nuclear histone H3 at Lys 9 and that this increase was remarkably sensitive to as low as 5 mM ethanol. Our long-term goal is to address how and why this acetylation occurs? This project will lay the foundation. Our hypothesis is that "ethanol causes selective acetylation in histones, both in vitro and in vivo, and that this would consequently affect the transcriptional machinery." Primary cultures of hepatocytes and livers from in vivo ethanol fed rats will serve as the main model in this project. There are three aims designed to build the framework for future development of this project. Aim I will determine the characteristics of ethanol induced histone acetylation in hepatocytes. In this aim, time course of this effect and its reversibility will be studied. We will also explore the specificity and diversity of this ethanol induced H3 acetylation response in cells of different tissue origins. Aim II will determine the mechanism(s) of histone acetylation in hepatocytes. The protocols will examine the effects of inhibitors of ethanol metabolism, individual ethanol metabolites or derived components, and the role of changes in levels of acetyl CoA in the acetylation response. This aim will also address whether increased expression and/or activation of histone acetyl transferase (HAT) or an inhibition of histone deacetylase (HDAC) are involved. Aim III is devoted to in vivo studies where effect of liquid diet ethanol feeding on rat liver H3 acetylation will be established. Experiments to study the consequence of acetylation on gene expression using chromatin immunoprecipitation (CHIP) assay will be performed and these experiments will form the basis for future extension of this project. This exploratory R21 proposal deals with a new direction in the alcohol field and offers the promise of providing mechanistic data into the ethanol effects at transcriptional level. Furthermore, "machinery" involved in histone modification will serve as a new target to develop therapeutic tools for prevention and treatment of ethanol induced cellular damage in liver and other organ systems affected by ethanol.