Regulation of human CYP2C9 expression by electrophilic stress involves AP-1 activation and DNA looping

Makia, N. L. and Surapureddi, S. and Monostory, K. and Prough, R. A. and Goldstein, J. A. (2014) Regulation of human CYP2C9 expression by electrophilic stress involves AP-1 activation and DNA looping. MOLECULAR PHARMACOLOGY, 86 (2). pp. 125-137. ISSN 0026-895X

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Abstract

CYP2C9 and CYP2C19 are important human enzymes that metabolize therapeutic drugs, environmental chemicals and physiologically important endogenous compounds. Initial studies using primary human hepatocytes showed induction of both the CYP2C9 and CYP2C19 genes by tert-butylhydroquinone (tBHQ). As a pro-oxidant, tBHQ regulates the expression of cytoprotective genes by activation of redox-sensing transcription factors, such as the nuclear factor E2-related factor 2 (Nrf2) and members of the activator protein 1 (AP-1) family of proteins. The promoter region of CYP2C9 contains two putative AP-1 sites (TGAGTCA) at positions -2201 and -1930 which are also highly conserved in CYP2C19. The CYP2C9 promoter is activated by ectopic expression of cFos and JunD while Nrf2 had no effect. Using specific kinase inhibitors for MAPK, we showed that ERK and JNK are essential for tBHQ-induced expression of CYP2C9. EMSA assays demonstrate that cFos distinctly interacts with the distal AP-1 site and JunD with the proximal site. Because cFos regulates target genes as heterodimers with Jun proteins, we hypothesized that DNA looping might be required to bring the distal and proximal AP-1 sites together to activate the CYP2C9 promoter. Chromosome conformation capture (3C) analyses confirmed the formation of a DNA loop in the CYP2C9 promoter possibly allowing interaction between cFos at the distal site and JunD at the proximal site to activate CYP2C9 transcription in response to electrophiles. These results indicate that oxidative stress generated by exposure to electrophilic xenobiotics and metabolites induces the expression of CYP2C9 and CYP2C19 in human hepatocytes.

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