A 66-Base-Pair Enhancer Module Activates the Expression of a Distinct Isoform of UDP-glucuronosyltransferase Family 1 (UGT1A2) in Primary Hepatocytes

Abstract

UGT1A2, an isoform of the UDP-glucuronosyltransferase family 1 (UGT1), is not expressed in the rat liver, but its expression was highly induced in primary cultures of rat hepatocytes. In primary hepatocytes that had been cultured for 70 h, the amount of UGT1A2 mRNA was 100 times higher than that in the rat liver. Deletion analysis of a 4.8-kb promoter region of the UGT1A2 gene revealed that a 66-nucleotide region between −307 and −242 upstream of the transcription start site was required for induction of UGT1A2 expression. The 66-nucleotide region acted on a heterologous promoter in a manner independent of its position and orientation in reporter constructs. Gel mobility shift assay showed that a specific binding protein to this region appeared in the nuclei of cultured hepatocytes, but was not present in the rat liver. DNase I protection analysis revealed the existence of a CTGGCAC core sequence between −274 and −268 of the UGT1A2 promoter. Methylation interference assay showed that the guanine residues at −294 and −287 on the upper strand and the guanine residue at −267 on the lower strand as well as the core sequence were required for the DNA–protein interaction. These results suggest that the 66-nucleotide region, which was designated culture-associated expression responsive enhancer module (CEREM), interacts with a specific nuclear protein and enhances the expression of UGT1A2 in cultured hepatocytes.