Role of activator protein-1 in the down-regulation of the human CYP2J2 gene in hypoxia.
The cytochrome P450 (CYP) 2J2 arachidonic acid epoxygenase gene was down-regulated at a pre-translational level in human hepatoma-derived HepG2 cells incubated in a hypoxic environment; under these conditions, the expression of c-Jun and c-Fos mRNA and protein was increased. The 5'-upstream region of the CYP2J2 gene was isolated by amplification of a 2341 bp fragment and putative regulatory elements that resembled activator protein-1 (AP-1)-like sequences were identified. From transient transfection analysis, c-Jun was found to strongly activate a CYP2J2 -luciferase reporter construct, but co-transfection with plasmids encoding c-Fos or c-Fos-related antigens, Fra-1 and -2, abrogated reporter activity. Using a series of deletion-reporter constructs, a c-Jun-responsive module was identified between bp -152 and -50 in CYP2J2 : this region contained an AP-1-like element between bp -56 and -63. The capacity of this element to interact directly with c-Jun, but not c-Fos, was confirmed by electromobility-shift assay analysis. Mutagenesis of the -56/-63 element abolished most, but not all, of the activation of CYP2J2 by c-Jun, thus implicating an additional site within the c-Jun-responsive region. The present results establish an important role for c-Jun in the control of CYP2J2 expression in liver cells. Activation of c-Fos expression by hypoxia promotes the formation of c-Jun/c-Fos heterodimers, which decrease the binding of c-Jun to the CYP2J2 upstream region, leading to gene down-regulation.
Department of Physiology and Pharmacology, School of Medical Sciences, University of New South Wales, Sydney, NSW 2052, Australia.
Nicole Y
NY
Marden
Eva
E
Fiala-Beer
Shi-Hua
SH
Xiang
Michael
M
Murray
D001483:Base Sequence; D001665:Binding Sites; D000090762:Cytochrome P-450 CYP2J2; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA; D015536:Down-Regulation / Q000502:physiology*; D016762:Genes, fos; D016758:Genes, jun; D006801:Humans; D000860:Hypoxia / Q000235:genetics*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010105:Oxygenases / Q000235:genetics*; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018808:Transcription Factor AP-1 / Q000378:metabolism / Q000502:physiology*; D014407:Tumor Cells, Cultured
C000717350:CYP2J2 protein, human; D018808:Transcription Factor AP-1; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; D000090762:Cytochrome P-450 CYP2J2