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Research Project: DISCOVERY AND DEVELOPMENT OF NATURAL PRODUCTS FOR PHARMACEUTICAL AND AGROCHEMICAL APPLICATIONS

Location: Natural Products Utilization Research

Title: DNA methyltransferase expressions in Japanese rice fish (Oryzias latipes) embryogenesis is developmentally regulated and modulated by ethanol and 5-azacytidine

Author
item Dasmahapatra, Asok - University Of Mississippi
item Khan, Ikhlas - University Of Mississippi

Submitted to: Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/9/2015
Publication Date: 7/13/2015
Citation: Dasmahapatra, A.K., Khan, I.A. 2015. DNA methyltransferase expressions in Japanese rice fish (Oryzias latipes) embryogenesis is developmentally regulated and modulated by ethanol and 5-azacytidine. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology. 13:176-177 (2015).

Interpretive Summary: Alcohol consumption by women during pregnancy often induces fetal alcohol spectrum disorder (FASD) in children. Molecular mechanism or prevention of FASD, other than women abstaining from alcohol drinking during pregnancy, is not known. FASD is regulated by both genetic and epigenetic mechanisms. Compared to genetic mechanisms, epigenetic mechanisms are enzyme based and reversible. Moreover, due to limitation of the use of synthetic anti-alcoholic drugs during pregnancy, our long term goal is to investigate on botanicals that have therapeutic potential for the treatment of alcoholism and thus FASD. We used Japanese rice fish (Oryzias latipes) as an animal model of FASD and focused on DNA methyl transferase enzymes as epigenome modulator during development. Our study indicates that Japanese rice fish embryos expressed three human ortholog dnmt genes during development and alcohol is able to attenuate the expression of dnmt1 gene. Therefore dnmt1gene should be a potential epigenetic target of FASD and will be used for screening botanicals for the discovery of antialcoholic drugs.

Technical Abstract: We aimed to investigate the impact of the epigenome in inducting fetal alcohol spectrum disorder (FASD) phenotypes in Japanese rice fish embryogenesis. One of the significant events in epigenome is DNA methylation which is catalyzed by DNA methyl transferase (DNMT) enzymes. We analyzed DNMT enzyme mRNA expressions in Japanese rice fish development starting from fertilized eggs to hatching and also in embryos exposed for first 48 h of development either to ethanol (300 mM) or to 5-azacytidine (5-azaC; 2mM), an inhibitor of DNMT enzyme activity. As observed in FASD phenotypes, 5-azaC exposure was able to induce microcephaly and craniofacial cartilage deformities in Japanese rice fish. Moreover, we have observed that expression of DNMTs (dnmt1, dnmt3aa, and dnmt3bb.1) are developmentally regulated; high mRNA copies were found in early stages (1-2 day-post-fertilization, dpf), followed by gradual reduction until hatched. In ethanol-treated embryos, compared to controls, dnmt1 mRNA is in reduced level in 2 dpf and in enhanced level in 6dpf embryos. While dnmt3aa and 3bb.1 remained unaltered. In contrast, embryos exposed to 5-azaC have an enhanced level of dnmt1 and dnmt3bb.1 mRNAs both in 2 and 6 dpf embryos while dnmt3aa is enhanced only in 6 dpf embryos. Moreover, endocannabinoid receptor 1a (cnr1a) mRNA which was found to be reduced by ethanol remained unaltered and cnr1b and cnr2 mRNAs, which were remained unaltered by ethanol, were increased significantly by 5-azaC in 6 dpf embryos. This study indicates that the craniofacial defects observed in FASD phenotypes are the results of dysregulations in DNMT expressions.