Ethanol action in Japanese medaka: Alteration in specific gene methylation

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DESCRIPTION (provided by applicant): The primary focus of the project described herein is to study the effects of ethanol on aldehyde dehydrogenase 1A2 (Aldh1A2) gene expression in Japanese medaka embryogenesis and to determine whether alteration in methylation pattern of the CpG island at the promoter region of this gene is a possible cause of ethanol teratogenesis. Ethanol induces fetal alcohol spectrum disorder (FASD) in the newborn babies of mothers who ingested alcohol during pregnancy. Among FASD, fetal alcohol syndrome (FAS) is the most clinically recognizable form identified by growth retardation, central nervous system (CNS) malformation and dysfunction, and has a distinctive pattern of craniofacial,cardiovascular, and limb defects. The molecular mechanism by which ethanol perturbs fetal development is unknown. Moreover, prevention of FAS, other than women abstaining from drinking alcohol during pregnancy, is not known. Previously, we have demonstrated that medaka embryos exposed to ethanol during embryogenesis have developed precocious FAS features in craniofacial, cardiovascular and skeletal organs which can be compared with FAS features observed in human. These findings prompted us to use medaka as a unique model for searching genes responsible for FAS. Our hypothesis is ethanol metabolism by the enzymes alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) alters cellular NAD + /NADH ratio and induces oxidative stress. This altered redox state is able to induce epigenetic modifications in DNA molecules of specific genes and thus modulate the normal development of the embryo. In this application we will study the DNA methylation pattern in the CpG island in the promoter region of Aldh1A2 gene, which we have identified as an ethanol target gene in medaka embryogenesis. We will also determine the effect of a natural product, kudzu (Pueraria lobota), in preventing these errors. Analyses will be made in embryonic, larval and adult stages of the animal. We will apply histological, histochemical and molecular biological techniques including quantitative RT-PCR and DNA methylation analysis for this study. This study will provide a foundation for identifying an alcohol metabolizing enzyme gene that is primarily responsible for FAS, and determine if a natural product will rescue the embryo from epigenetic ethanol toxicity. In this proposal, we have focused on an ethanol-induced epigenetic event as the major cause of Fetal Alcohol Syndrome and evaluate a natural product as the preventive agent for FAS.