A Novel Epigenetic Landscape in the Biology of Nonalcoholic Fatty Liver Disease: The role of 5-Hydroxymethylcytosine and Sequence Variation in Ten-Eleven Translocation Family of Proteins.

CARLOS J. PIROLA, TOMAS FERNÁNDEZ GIANOTTI, HERNÁN DOPAZO, CRISTIAN ROHR, GUSTAVO O. CASTAÑO, SILVIA SOOKOIAN.

Boston

Congreso; Annual Meeting of the American Association for the Study of Liver Diseases; 2014

AASLD. American Association for the Study of Liver Diseases

Background: 5-hydroxymethylcytosine (5-hmC) is a new epigenetic mark that modulates gene transcription by influencing DNA demethylation and chromatin structure remodeling; non-CG hydroxymethylation appears to be prevalent in the mitochondrial genome. The Ten-Eleven-Translocation (TET 1, 2 and 3) family of proteins is responsible for catalyzing the conversion of 5-methylcytosine (5-mC) to 5-hmC. Because nonalcoholic steatohepatitis (NASH) is associated with altered DNA methylation profiles, changes in mitochondrial DNA (mtDNA) content and PPARGC1A expression, we hypothesized that epigenetic editing by 5-hmC might be a novel mechanism to explain the NAFLD phenotype. Hence, we explored the global levels of 5-hmC in the liver of NAFLD patients at different stages of disease severity (n=67) and controls (n=23). In addition, we screened for genetic variation in TET 1-3 loci by targeted deep sequencing to explore its contribution to the disease biology; for this purpose, we included simple steatosis (SS) n=32, NASH n=32 and controls n=32. Methods: Global 5-hmC quantification in liver genomic DNA was performed by Quest 5-hmC DNA ELISA. Sequencing of TET1 (9.7 kb, 12 exons), TET2 (19 kb, 13 exons) and TET3 (11 kb, 9 exons) was performed by next generation sequencing (Ion Personal Genome Machine) using 316 chips and barcoding (>100 x coverage); liver mtDNA was evaluated by real-time PCR. Results: Examination of liver 5-hmC levels showed significant and positive correlation with liver mtDNA content (Spearman R: 0.50, p