MOLECULAR MECHANISM OF E2 MODULATION OF CYP1A TRANSCRIPTIONAL REGULATION

Several hypotheses have been proposed explaining CYP1A1 down-regulation by E2 and their mimics. For example, inhibitory action of E2 could be mediated, at least in part, through the hepatic ER where the ER-E2 complex can interfere with the CYP1A1 gene directly or alternatively with the AhR, and indirectly regulate CYP1A1 gene expression through binding the XRE or estrogens and their mimics may control the recruitment of ER and possibly other co-activators, besides activating the detoxification pathway. We have evaluated mechanistically, the effect of E2 on CYP1A1 expression. Primary salmon hepatocytes were exposed to E2 (0.001 μM) singly and in combination with ER-antagonist (ICI182780), AhR inhibitor (3’,4’- dimethoxyflavone; DMF) or protein synthesis inhibitor (cycloheximide; CyH). Hepatocytes were exposed for 6, 12 and 24 hours. The expression of genes and proteins involved in ER (ERα, ERβ and vitellogenin) and AhR (CYP1A1, AhR-repressor, AhR-isotypes and cofactors) pathways were analysed using qPCR and immunochemical methods. Biochemical assay methods were used for CYP1A1 and proteasomal activities. We showed that E2 produced induction of genes in the ER signalling pathway and inhibition of transcriptional responses of mRNA species in the AhR signalling pathway and that these responses were negatively influenced by protein and receptor inhibitors, respectively. Overall, our data showed that E2 regulation of CYP1A1 involves both receptor deactivation and receptor-protein destabilization. Given that the Per–AhR/Arnt–Sim homology sequence of transcription factor usually associate with each other to form heterodimers and bind the XRE sequences in the promoter regions of the target genes to regulate their expression, the complete mechanism by which estrogenic compounds regulate the CYP system may not have been fully elucidated.