Involvement of CCAAT/enhancer binding protein-beta (C/EBP beta) in epigenetic regulation of mouse methionine adenosyltransferase 1A gene expression

Methionine adenosyltransferase (MAT) catalyzes the synthesis of S-adenosylmethionine, the main methyl donor in cellular transmethylation reactions and the aminopropyl moiety in polyamine biosynthesis. In mammals, two different genes, MATIA and MAT2A, encode catalytic polypeptides of Liver-specific MAT 1/111 and ubiquitous MAT H, respectively. Reverse transcriptionpolymerase chain reaction showed that MATIA gene expression was at a detectable level in embryonic day 14 mouse fetal liver and subsequently increased. Bisulfite genomic sequencing indicated that the methylation status of 10 CpG sites in the MATIA promoter proximal region was appreciably correlated with the gene expression in mouse developing liver and in adult hepatic cells; hepatic stellate cells and hepatocytes. When mouse hepatoma-derived Hepa-I cells showing extremely low expression of MATIA gene were treated with 5-aza-2'-deoxycytidine and trichostatim A, MATIA gene expression was enhanced. In addition, in vitro methylation of the MATIA promoter region suppressed the MATIA promoter activity in reporter assay. Next, we performed electrophoretic mobility shift assay and found that the transcriptional factor CCAAT/enhancer binding protein-P (C/EBPP) specifically binds to a putative binding site of C/EBPP in the MAT IA promoter. Suppression of C/EBPP expression by short hairpin RNA decreased the MATlA promoter activity and MATIA gene expression, and inhibition of C/EBPP binding to MATIA by site-directed mutagenesis also showed similar results. Western blot analysis and chromatin immunoprecipitation assay indicated that C/EBPO binding is dependent on DNA methylation status. Based on these findings, we conclude that C/EBPP plays an important role in epigenetic regulation of the mature hepatic gene MATIA. (c) 2008 Elsevier Ltd. All rights reserved.