Kress S, Greenlee W F
Department of Toxicology, University of Tübingen, Germany.
Cancer Res. 1997 Apr 1;57(7):1264-9.
In this report, we present a characterization of the cell-specific expression of two human cytochrome P450 genes, CYP1A1 and CYP1B1, by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The TCDD-dependent induction of CYP1A1 has been studied extensively and serves as the prototype response for a TCDD-signaling pathway initiated by the reversible binding of TCDD to an intracellular receptor [designated the aryl hydrocarbon (Ah) receptor]. CYP1A1 is induced by TCDD to high levels (45-fold increase) in the human hepatoblastoma line HepG2 as compared with the human renal adenocarcinoma line ACHN. In contrast, CYP1B1 is induced selectively in ACHN cells. Cell-specific induction of CYP1A1 and CYP1B1 mRNA correlates with comparable changes in the corresponding proteins and results, at least in part, from transcriptional activation. Characterization of the mechanism(s) for the differential regulation of CYP1A1 was carried out. Nuclear extracts obtained from either cell line following treatment with TCDD displayed equivalent binding to oligonucleotide probes for two dioxin-responsive elements located 5'-ward of the CYP1A1 promoter. This result obtained with broken cell fractions was confirmed by an intact cell DNA protection assay. Possible involvement of negative regulators is suggested by the presence of a negative regulatory element in the 5' flanking region of the CYP1A1 gene and the observed superinduction of CYP1A1 mRNA by cycloheximide in TCDD-treated HepG2 cells. Electromobility shift analysis using negative regulatory element probes, however, did not detect quantitative differences in the binding of nuclear extract proteins obtained from either HepG2 or ACHN cells treated with TCDD. These findings indicate that the ligand-dependent activation and dioxin-responsive element binding of the Ah receptor required for CYP1A1 induction in HepG2 cells also can occur in ACHN cells. We conclude that the repression of TCDD-dependent CYP1A1 induction in ACHN cells occurs at the level of transactivation in the Ah receptor signal transduction pathway.
在本报告中，我们描述了2,3,7,8-四氯二苯并对二噁英（TCDD）对两个人细胞色素P450基因CYP1A1和CYP1B1细胞特异性表达的影响。TCDD对CYP1A1的诱导作用已得到广泛研究，它是由TCDD与细胞内受体（称为芳烃受体，Ah受体）可逆结合引发的TCDD信号通路的典型反应。与人类肾腺癌ACHN细胞系相比，TCDD可将人肝癌细胞系HepG2中的CYP1A1诱导至高水平（增加45倍）。相反，CYP1B1在ACHN细胞中被选择性诱导。CYP1A1和CYP1B1 mRNA的细胞特异性诱导与相应蛋白质的类似变化相关，并且至少部分是由转录激活引起的。我们对CYP1A1差异调节机制进行了研究。用TCDD处理后，从任一细胞系获得的核提取物与位于CYP1A1启动子5'端的两个二噁英反应元件的寡核苷酸探针显示出同等结合。用破碎细胞组分得到的这一结果通过完整细胞DNA保护试验得到了证实。CYP1A1基因5'侧翼区域存在负调控元件以及在TCDD处理的HepG2细胞中观察到环己酰亚胺对CYP1A1 mRNA的超诱导，提示可能存在负调控因子。然而，使用负调控元件探针进行的电泳迁移率变动分析未检测到用TCDD处理的HepG2或ACHN细胞获得的核提取物蛋白结合上的定量差异。这些发现表明，HepG2细胞中CYP1A1诱导所需的Ah受体的配体依赖性激活和二噁英反应元件结合在ACHN细胞中也可发生。我们得出结论，ACHN细胞中TCDD依赖性CYP1A1诱导的抑制发生在Ah受体信号转导途径的反式激活水平。
