Elbekai Reem H, El-Kadi Ayman O S
Faculty of Pharmacy and Pharmaceutical Sciences, 3126 Dentistry/Pharmacy Centre, University of Alberta, Edmonton, Alberta, Canada.
Toxicol Lett. 2007 Aug;172(3):106-19. doi: 10.1016/j.toxlet.2007.05.009. Epub 2007 May 24.
Heavy metals alter the carcinogenicity of AhR ligands by modulating the induction of the Cyp1a1 enzyme, but the mechanism(s) remain unresolved. In this study, the effect of the heavy metals, As(3+), Cd(2+), and Cr(6+), on the transcriptional and posttranscriptional regulation of the Cyp1a1 gene was investigated in Hepa 1c1c7 cells. A time-dependent study showed that all metals significantly induced the basal Cyp1a1 mRNA, but only As(3+) and Cd(2+) further potentiated the inducible Cyp1a1 mRNA level. Alternately, Cr(6+) inhibited the TCDD-mediated induction of Cyp1a1 mRNA. As(3+) potentiated the induction of Cyp1a1 mRNA by TCDD only at 3h and 6h after treatment. Cd(2+) on the other hand further potentiated the induction up to 24h after treatment. The metal-mediated induction of Cyp1a1 mRNA was further potentiated by the protein synthesis inhibitor, cycloheximide and the 26S proteasome inhibitor, MG-132, but completely inhibited by the RNA transcription inhibitor, actinomycin-D, implying a transcriptional regulation of the Cyp1a1 gene by the heavy metals. Not surprisingly, Cd(2+) and Cr(6+) activated the DNA-binding capacity of the AhR for the xenobiotic responsive element, as measured by the electrophoretic-mobility shift assay while all three metals induced AhR-dependent luciferase reporter gene expression in transiently transfected Hepa 1c1c7 cells. On the other hand, only As(3+) increased the Cyp1a1 mRNA half-life while Cd(2+) and Cr(6+) increased the Cyp1a1 protein half-life, suggesting the involvement of posttranslational modifications. A significant decrease in TCDD-mediated induction of Cyp1a1 activity was associated with an increase in HO-1 mRNA levels and a concomitant decrease in cellular heme content after all metal treatments. We clearly demonstrated that As(3+), Cd(2+), and Cr(6+) increase Cyp1a1 mRNA levels at the transcriptional and posttranscriptional levels while decreasing Cyp1a1 activity at the posttranslational level.
重金属通过调节Cyp1a1酶的诱导来改变芳烃受体（AhR）配体的致癌性，但其机制仍未明确。在本研究中，我们在Hepa 1c1c7细胞中研究了重金属砷（As(3+)）、镉（Cd(2+)）和铬（Cr(6+)）对Cyp1a1基因转录和转录后调控的影响。一项时间依赖性研究表明，所有金属均显著诱导基础Cyp1a1 mRNA，但只有As(3+)和Cd(2+)进一步增强了可诱导的Cyp1a1 mRNA水平。相反，Cr(6+)抑制了TCDD介导的Cyp1a1 mRNA诱导。As(3+)仅在处理后3小时和6小时增强了TCDD对Cyp1a1 mRNA的诱导。另一方面，Cd(2+)在处理后长达24小时进一步增强了诱导作用。蛋白质合成抑制剂环己酰亚胺和26S蛋白酶体抑制剂MG-132进一步增强了金属介导的Cyp1a1 mRNA诱导，但RNA转录抑制剂放线菌素-D完全抑制了这种诱导，这意味着重金属对Cyp1a1基因有转录调控作用。不出所料，通过电泳迁移率变动分析测定，Cd(2+)和Cr(6+)激活了AhR对异源生物反应元件的DNA结合能力，而所有三种金属在瞬时转染的Hepa 1c1c7细胞中均诱导了AhR依赖性荧光素酶报告基因表达。另一方面，只有As(3+)增加了Cyp1a1 mRNA半衰期，而Cd(2+)和Cr(6+)增加了Cyp1a1蛋白半衰期，这表明存在翻译后修饰。所有金属处理后，TCDD介导的Cyp1a1活性显著降低与HO-1 mRNA水平升高以及细胞血红素含量随之降低有关。我们清楚地证明，As(3+)、Cd(2+)和Cr(6+)在转录和转录后水平上增加Cyp1a1 mRNA水平，而在翻译后水平上降低Cyp1a1活性。
