Reduced susceptibility of DNA methyltransferase 1 hypomorphic (Dnmt1N/+) mice to hepatic steatosis upon feeding liquid alcohol diet.
BACKGROUND
Methylation at C-5 (5-mdC) of CpG base pairs, the most abundant epigenetic modification of DNA, is catalyzed by 3 essential DNA methyltransferases (Dnmt1, Dnmt3a and Dnmt3b). Aberrations in DNA methylation and Dnmts are linked to different diseases including cancer. However, their role in alcoholic liver disease (ALD) has not been elucidated.


RESULTS
Dnmt1 wild type (Dnmt1(+/+)) and hypomorphic (Dnmt1(N/+)) male mice that express reduced level of Dnmt1 were fed Lieber-DeCarli liquid diet containing ethanol for 6 weeks. Control mice were pair-fed calorie-matched alcohol-free liquid diet, and Dnmtase activity, 5-mdC content, gene expression profile and liver histopathology were evaluated. Ethanol feeding caused pronounced decrease in hepatic Dnmtase activity in Dnmt1(+/+) mice due to decrease in Dnmt1 and Dnmt3b protein levels and upregulation of miR-148 and miR-152 that target both Dnmt1 and Dnmt3b. Microarray and qPCR analysis showed that the genes involved in lipid, xenobiotic and glutathione metabolism, mitochondrial function and cell proliferation were dysregulated in the wild type mice fed alcohol. Surprisingly, Dnmt1(N/+) mice were less susceptible to alcoholic steatosis compared to Dnmt1(+/+) mice. Expression of several key genes involved in alcohol (Aldh3b1), lipid (Ppara, Lepr, Vldlr, Agpat9) and xenobiotic (Cyp39a1) metabolism, and oxidative stress (Mt-1, Fmo3) were significantly (P<0.05) altered in Dnmt1(N/+) mice relative to the wild type mice fed alcohol diet. However, CpG islands encompassing the promoter regions of Agpat9, Lepr, Mt1 and Ppara were methylation-free in both genotypes irrespective of the diet, suggesting that promoter methylation does not regulate their expression. Similarly, 5-mdC content of the liver genome, as measured by LC-MS/MS analysis, was not affected by alcohol diet in the wild type or hypomorphic mice.


CONCLUSIONS
Although feeding alcohol diet reduced Dnmtase activity, the loss of one copy of Dnmt1 protected mice from alcoholic hepatosteatosis by dysregulating genes involved in lipid metabolism and oxidative stress.
Department of Molecular and Cellular Biochemistry, College of Medicine, The Ohio State University, Columbus, Ohio, United States of America.
Huban
H
Kutay
Corie
C
Klepper
Bo
B
Wang
Shu-hao
SH
Hsu
Jharna
J
Datta
Lianbo
L
Yu
Xiaoli
X
Zhang
Sarmila
S
Majumder
Tasneem
T
Motiwala
Nuzhat
N
Khan
Martha
M
Belury
Craig
C
McClain
Samson
S
Jacob
Kalpana
K
Ghoshal
D000438:Alcohols / Q000494:pharmacology; D000821:Animal Feed; D000818:Animals; D002460:Cell Line; D002853:Chromatography, Liquid / Q000379:methods; D018899:CpG Islands; D004247:DNA / Q000235:genetics; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics*; D019175:DNA Methylation; D000090205:DNA Methyltransferase 3A; D004195:Disease Models, Animal; D044127:Epigenesis, Genetic; D000431:Ethanol / Q000494:pharmacology; D005234:Fatty Liver / Q000201:enzymology* / Q000235:genetics*; D016678:Genome; D006801:Humans; D050356:Lipid Metabolism; D008108:Liver Diseases, Alcoholic / Q000235:genetics; D008297:Male; D051379:Mice; D018384:Oxidative Stress; D053719:Tandem Mass Spectrometry / Q000379:methods; D015262:Xenobiotics / Q000378:metabolism; D000097584:DNA Methyltransferase 3B
D000438:Alcohols; D004247:DNA; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D000431:Ethanol; D015262:Xenobiotics; D000097584:DNA Methyltransferase 3B; C000717967:DNMT3A protein, human; C000715535:Dnmt3a protein, mouse; C000619380:DNMT1 protein, human; C000619381:Dnmt1 protein, mouse