Novel Transcriptional Regulation of the Human CYP3A7Gene by Sp1 and Sp3 through Nuclear Factor κB-like Element

Abstract

Human CYP3A7 and CYP3A4 are expressed in fetal and adult livers, respectively, although the 5′-flanking regions of the two genes show 90% homology. The purpose of this study was to clarify the mechanism(s) responsible for the transcriptional regulation of theCYP3A7 gene in human hepatoma HepG2 cells that showed fetal phenotypes. Transfection studies using a series of theCYP3A7 or CYP3A4 promoter-luciferase chimeric genes identified a nuclear factor κB (NF-κB)-like element between nucleotides −2326 and −2297 that conferred the transcriptional activation of the CYP3A7 gene. A 1-base pair mismatch within the corresponding region of the CYP3A4 gene was sufficient for a differential enhancer activity. A gel shift assay using nuclear extracts from HepG2 cells showed that Sp1 and Sp3 bound to the NF-κB-like element of the CYP3A7 but notCYP3A4 gene. Specific activation of the CYP3A7promoter by Sp1 and Sp3 was confirmed by a co-transfection of the p3A7NF-κB or p3A4NF-κB reporter gene with Sp1 or Sp3 expression plasmid into Drosophila cells, which lacked endogenous Sp family. Additionally, introduction of mutations into binding sites for hepatocyte nuclear factor 3β, upstream stimulatory factor 1, and a basic transcription element in the proximal promoter attenuated luciferase activity to 20% of the level seen with the intactCYP3A7 promoter. Thus, we conclude that the expression of the CYP3A7 gene in HepG2 cells is cooperatively regulated by Sp1, Sp3, hepatocyte nuclear factor 3β, and upstream stimulatory factor 1.