Aryl hydrocarbon receptor signaling in rainbow trout hepatocytes: role of hsp90 and the proteasome.
The objective of this study was to investigate the role of heat shock protein 90 (hsp90) and the proteasome in regulating aryl hydrocarbon receptor (AhR) activation and cytochrome P450 1A (Cyp1A) protein expression in rainbow trout (Oncorhynchus mykiss). We exposed trout hepatocytes in primary culture to the AhR agonist beta-napthoflavone (betaNF; 10(-6) M) and examined AhR and Cyp1A expression. betaNF-induced a significant temporal accumulation of AhR and Cyp1A1 mRNA abundance in trout hepatocytes. This transcript response was followed by a significantly higher AhR and Cyp1A protein expression. Exposure to geldanamycin (GA; 1000 ng mL(-1)), a benzoquinone ansamycin antibiotic used to inhibit hsp90 function, significantly reduced ( approximately 70%) betaNF-induced Cyp1A protein expression. Also, exposure to the proteasomal inhibitor MG-132 (50 microM) completely abolished betaNF-induced Cyp1A protein expression in trout hepatocytes. In addition, MG-132 treatment further enhanced the GA-mediated suppression of the Cyp1A response. The effect of MG-132 on Cyp1A response corresponded with a significant inhibition of BNF-mediated AhR mRNA abundance, but not protein content. Altogether our results suggest a betaNF-mediated autoregulation of AhR content in trout hepatocytes. We propose a key role for hsp90 and the proteasome in this ligand-mediated AhR regulation and Cyp1A response.
Wiseman
Steve B
SB
Department of Biology, University of Waterloo, Waterloo, Ontario, N2L 3G1, Canada.
Vijayan
Mathilakath M
MM
D000818:Animals; D016227:Benzoquinones / Q000494:pharmacology; D002478:Cells, Cultured; D019363:Cytochrome P-450 CYP1A1 / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism*; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D007976:Leupeptins / Q000494:pharmacology; D017686:Oncorhynchus mykiss; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D019324:beta-Naphthoflavone / Q000494:pharmacology
D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D007976:Leupeptins; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon; D019324:beta-Naphthoflavone; D019363:Cytochrome P-450 CYP1A1; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; C001277:geldanamycin