TRANSCRIPTIONAL CONTROL OF THE RAT HEPATIC CYP2E1 GENE

The rat hepatic CYP2E1 gene becomes transcriptionally activated within 1 day after birth. This activation can be mimicked by using the 5′ end of the gene in a cell-free nuclear extract prepared from hepatocytes taken from rats at different developmental stages. Deletion analysis revealed that a positive element located between -127 and -89 was responsible for 90% of the in vitro transcription activity of adult liver extracts. Protein binding studies revealed that this region was operationally equivalent to the binding site for the factor HNF-1. Two other protein-binding regions were uncovered, one of which corresponded to the site for a CCAAT-binding factor NFY. The other site was a palindrome sequence unique to the CYP2E1 gene. These latter two factors did not significantly contribute to transcriptional activity in vitro and were not conserved between the rat and human CYP2E1 genes. Extracts prepared from fetal and newborn livers were transcriptionally inactive, whereas extracts from livers of 3-day-old rats were fully active toward the CYP2E1 gene. DNase I footprinting patterns indistinguishable between fetal and adult extracts were obtained for all three factors. However, gel mobility shift assays revealed a second, higher-mobility band produced by fetal and newborn liver extracts bound to the HNF-1 oligomer. UV-cross-linking studies showed that adult and fetal extracts had only a single 98-kilodalton protein that bound to this oligomer. In contrast, adult lung samples, also transcriptionally inactive toward the CYP2E1 gene, contained two proteins of slightly greater than 110 kilodaltons. These results suggest that the CYP2E1 gene is positively regulated in adult rats by HNF-1 or a protein similar in DNA-binding properties to HNF-1. The role of this factor or other protein-protein interactions in the lack of CYP2E1 transcription in fetal and newborn animals remains unclear.