Cross-coupling of signal transduction pathways: the dioxin receptor mediates induction of cytochrome P-450IA1 expression via a protein kinase C-dependent mechanism.

Abstract

Signal transduction by dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin) is mediated by the intracellular dioxin receptor which, in its dioxin-activated state, regulates transcription of target genes encoding drug-metabolizing enzymes, such as cytochrome P-45OIA1 and glutathione S-transferase Ya. Exposure of the dioxin receptor to dioxin leads to an apparent translocation of the receptor to the nucleus in vivo and to a rapid conversion of the receptor from a latent, non-DNA-binding form to a species that binds to dioxin-responsive positive control elements in vitro. This DNA-binding form of receptor appears to be a heterodimeric complex with the helix-loop-helix factor Arnt. In this study, we show that activation of the cytochrome P-450IA1 gene and minimal dioxin-responsive reporter constructs by the dioxin receptor was inhibited following prolonged treatment of human keratinocytes with the phorbol ester 12-0-tetradecanoylphorbol-13-acetate. Inhibition of the receptor-mediated activation response was also achieved by treatment of the cells with a number of protein kinase inhibitors, one of which, calphostin C, shows selectivity for protein kinase C. Taken together, these data suggest that protein kinase C-dependent phosphorylation may play an essential role in the dioxin signaling pathway. This hypothesis is supported by the observation that pretreatment of the cells with 12-0-tetradecanoylphorbol-13-acetate inhibited the DNA-binding activity of the dioxin receptor in vivo. In vivo, the dioxin receptor was found to be a phosphoprotein. In vitro, dephosphorylation of the ligand-activated, heteromeric dioxin receptor form or dephosphorylation of the individual ligand-binding and Arnt receptor subunits inhibited the xenobiotic response element-binding activity. Moreover, dephosphorylation experiments with the individual receptor subunits prior to assembly of the xenobiotic response element-binding receptor form indicated that phosphorylation seemed to be important for the DNA-binding activity per se of the receptor, whereas Arnt appeared to require phosphorylation to interact with the receptor. Finally, a protein kinase C inhibitor-sensitive cytosolic catalytic activity that could restore the DNA-binding activity of the dephosphorylated dioxin receptor form was identified.Dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin) and several related halogenated aromatic hydrocarbons are members of a class of environmental pollutants that give rise to a plethora of teratogenic, tumorigenic, and toxic responses in rodents. In humans, a well-documented toxic effect of this class of compounds is a hyperkeratotic and metaplastic response of the hair follicles and interfollicular epidermis, leading to persistent acne-like lesions called chloracne (reviewed in reference 58). On a molecular level, dioxin induces transcription of genes encoding drug-metabolizing enzymes, such as glutathione S-transferase Ya, quinone reductase, and cytochrome P-450IA1, in both rodent and human cells (for a review, see reference 30).Signal transduction by dio...

Phorbol Ester-induced Transcription of a Fibroblast Growth Factor-binding Protein Is Modulated by a Complex Interplay of Positive and Negative Regulatory Promoter Elements

“…In addition, it is known that TPA causes an immediate up-regulation of PKC followed by long term down-regulation of PKC activity. In contrast, although long term calphostin C appears to be able to down-regulate protein kinase C, it does not cause early induction of PKC activation but rather blocks the inductive effects of TPA on PKC (14). Thus the fact that Calphostin C causes no induction of FGF-BP mRNA and blocks the TPA effect argues that induction of FGF-BP mRNA is through an up-regulation of PKC activity rather than a consequence of long term down-regulation.…”

Definition of a Dioxin Receptor Mutant That Is a Constitutive Activator of Transcription

“…To construct the glucocorticoid receptor/ mouse dioxin receptor deletion constructs for expression in yeast, an NcoI fragment was isolated from pDR⌬LBD/Gem and pDR⌬PASB/Gem and subcloned into NcoI-digested pGRDBDmDR/2HG to give pGRDBD/ DR⌬LBD/2HG and pGRDBD/DR⌬PASB/2HG, respectively. The mammalian reporter gene constructs pTX.DIR and p(GRE) 2 T105LUC and the yeast reporter plasmid pUC⌬SS26X have been described previously (27)(28)(29).…”

Role of the aromatic hydrocarbon receptor in the suppression of cytochrome P-450 2C11 by polycyclic aromatic hydrocarbons

“…The process by which ligand binding transfomis the cytosolic AHR to its functional DNA-binding state is complicated and poorly understood. Phosphorylation of both AHR and ARNT appears to be important for generation of the functional DNA-binding corn plex (Perdew, 1991 ;Berghard et al. 1993).…”