Structure of the rat methionine adenosyltransferase 2A gene and its promoter

In this study, to understand the regulation of methionine adenosyltransferase (MAT) gene expression, we isolated the rat MAT2A gene encoding MAT alpha 2, the catalytic subunit of non-hepatic-type enzyme MAT II and characterized its structural organization and 5'-flanking region. The gene spans approximately 7 kbp and consists of nine exons interrupted by eight introns. The transcription initiation site, as demonstrated by primer extension analysis, is located 123 bp upstream of the translation start codon. Comparison of the structural organization of the rat MAT2A gene to that of the mouse MAT1 A gene encoding MAT alpha 1, the subunit of liver-type enzymes MAT I and III, shows that the exon structure of two genes is very similar and the insertion sites of all corresponding introns are identical. A canonical TATA box and a GC box, the potential Spl-binding site, are found 32 bp and 70 bp upstream of the transcription initiation site, respectively. The 5'-flanking region also contains potential recognition sites for various transcription factors including AP-I, AP-2 and NF-IL6 (C/EBP beta), and a large G+C-rich domain with the characteris ties of a CpG island. The 5'-flanking sequence of the rat MAT2A gene has no significant similarity with those of the MAT1A genes. Transient transfection experiments using a luciferase reporter gene showed that the first 820-bp sequence of the 5'-flanking region directed high levels of luciferase activity in cultured rat kidney fibroblast (NRK-49F) and hepatocellular carcinoma (FAA-HTC1) cells, but not in primary rat hepatocytes. Deletion analysis suggested that the first 343 bp of the 5'-flanking region contained cell-type-specific promoter elements of this gene.