Calpain mediates the dioxin-induced activation and down-regulation of the aryl hydrocarbon receptor.
The aryl hydrocarbon receptor (AhR) is a ligand-activated basic-helix-loop-helix transcription factor that binds polyaromatic hydrocarbons (PAH), such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and mediates their toxicity. Binding of PAH to AhR in the cytoplasm triggers a poorly defined transformation step of the receptor into a nuclear transcription factor. In this study, we show that the calcium-dependent cysteine protease calpain plays a major role in the ligand-induced transformation and signaling of AhR. Fluorescence imaging measurements showed that TCDD treatment elevates intracellular calcium, providing the trigger for calpain activation, as measured toward t-butoxycarbonyl-Leu-Met-chloromethylaminocoumarin, a calpain-specific substrate. Inhibition of calpain activity by the N-benzyloxycarbonyl-Val-Phe-aldehyde (MDL28170) blocked the TCDD-induced nuclear translocation of AhR in Hepa1c1c7 mouse hepatoma cell line. Treatment of the human metastatic breast carcinoma cell line MT-2 with MDL28170 and 3-(4-iodophenyl)-2-mercapto-(Z)-2-propenoic acid (PD 150606), two calpain-selective inhibitors, completely abolished the TCDD-induced transactivation of AhR as assessed by transcription of CYP1A1 gene. Previous studies have established that after TCDD-induced transactivation, the AhR undergoes a massive depletion; we show here that selective calpain inhibitors can block this step, which suggests that the ligand-induced down-regulation of the AhR is calpain-dependent. The data presented support a major role for calpain in the AhR transformation, transactivation, and subsequent down-regulation, and provide a possible explanation for many of the reported phenomena of ligand-independent activation of AhR.
Dale
Yolanda R
YR
Graduate Program in Pharmacology, Meharry Medical College, Nashville, Tennessee, USA.
Eltom
Sakina E
SE
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics; D015536:Down-Regulation / Q000187:drug effects*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D051379:Mice; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D018336:Receptors, Aryl Hydrocarbon / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D019363:Cytochrome P-450 CYP1A1; D002154:Calpain; D002118:Calcium