Wiseman Steve B, Vijayan Mathilakath M
Department of Biology, University of Waterloo, Waterloo, Ontario, N2L 3G1, Canada.
Comp Biochem Physiol C Toxicol Pharmacol. 2007 Nov;146(4):484-91. doi: 10.1016/j.cbpc.2007.05.006. Epub 2007 Jun 9.
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.
本研究的目的是探讨热休克蛋白90（hsp90）和蛋白酶体在调节虹鳟（Oncorhynchus mykiss）中芳烃受体（AhR）激活及细胞色素P450 1A（Cyp1A）蛋白表达方面的作用。我们将原代培养的鳟鱼肝细胞暴露于AhR激动剂β-萘黄酮（βNF；10⁻⁶ M），并检测AhR和Cyp1A的表达。βNF诱导鳟鱼肝细胞中AhR和Cyp1A1 mRNA丰度出现显著的时间依赖性积累。这种转录本反应之后是AhR和Cyp1A蛋白表达显著升高。暴露于格尔德霉素（GA；1000 ng mL⁻¹），一种用于抑制hsp90功能的苯醌安莎霉素类抗生素，显著降低（约70%）βNF诱导的Cyp1A蛋白表达。此外，暴露于蛋白酶体抑制剂MG - 132（50 μM）完全消除了βNF诱导的鳟鱼肝细胞中Cyp1A蛋白表达。另外，MG - 132处理进一步增强了GA介导的对Cyp1A反应的抑制。MG - 132对Cyp1A反应的影响与BNF介导的AhR mRNA丰度的显著抑制相对应，但对蛋白含量无影响。总之，我们的结果表明βNF介导了鳟鱼肝细胞中AhR含量的自动调节。我们提出hsp90和蛋白酶体在这种配体介导的AhR调节及Cyp1A反应中起关键作用。
