Transcriptional regulation of α-fetoprotein expression by dexamethasone in human hepatoma cells

Abstract

The level of alpha-fetoprotein (AFP) mRNA in HuH-7 human hepatoma cells is elevated by the addition of dexamethasone to the culture medium. To locate the DNA region involved in hormonal regulation of the AFP gene, we constructed recombinant plasmids in which various lengths of the 5'-flanking sequence of the human AFP gene were fused to the CAT gene. Various cell lines were transfected with the recombinant plasmids, incubated with or without 3 x 10(-6) M dexamethasone, and then assayed for chloramphenicol acetyltransferase expression. In hepatoma cells that produce AFP, the dexamethasone treatment resulted in the stimulated chloramphenicol acetyltransferase expression when the transfected plasmids contained 169 base pairs (bp) or longer AFP 5'-flanking sequence. No dexamethasone effect was observed when the 5'-flanking sequence was less than 98 bp long. The dexamethasone stimulation was effectively suppressed by the glucocorticoid antagonist RU486, indicating that this effect is mediated by glucocorticoid receptors. The 71-bp region between positions -169 and -98 contains a nucleotide stretch which is similar to the consensus sequence of the glucocorticoid responsive element (GRE). Partial alterations of this sequence resulted in decreased dexamethasone response. The GRE-containing region stimulated heterologous (SV40) promoter activity in response to dexamethasone treatment in an orientation- and position-independent manner. The GRE and the upstream AFP enhancer function independently from each other.