Add time:09/10/2019         Source:sciencedirect.com

Recently, we found a region A (a hepatocyte nuclear factor (HNF)-4-binding site from nucleotides −701 to −684) and a region B (an HNF-1-binding site from nucleotides −682 to −666) as cis-acting elements necessary for the transcriptional activation of the human dihydrodiol dehydrogenase (DD)4 gene in human hepatoblastoma HepG2 cells, which express DD4 mRNA [20]. Thus, to investigate the mechanism(s) responsible for the cell-type-specific expression of DD4 mRNA, we constructed a reporter plasmid, pDD4 Foot A + B: −95/+28, in which regions A and B were linked to the human DD4 minimal promoter (−95 to +28) fused to the luciferase gene. The luciferase activity was detectable in HepG2 cells but not in human renal adenocarcinoma ACHN cells transfected with the pDD4 Foot A + B: −95/+28, which do not express DD4 mRNA. A supershift assay using antibodies to HNF-4α, -4γ, -1α, or valiant HNF (vHNF)-1 revealed that HNF-4α, -4γ, and -1α recognized regions A and B in HepG2 cells and that only vHNF-1 bound to regions A and B in ACHN cells. Semiquantitative reverse-transcribed polymerase chain reaction showed that a large amount of vHNF-1-C, which lacked transcriptional activation domain, was expressed in ACHN cells. Transfection of ACHN cells with an expression plasmid of vHNF-1-C did not activate the pDD4 Foot A + B: −95/+28 reporter gene. Taken together, we conclude that the cell-type-specific expression of DD4 mRNA is regulated by vHNF-1-C.

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