Xu C, Pasco D S
Molecular Biology Laboratory, Maharishi University of Management, Fairfield, Iowa, 52557, USA.
Arch Biochem Biophys. 1998 Aug 15;356(2):142-50. doi: 10.1006/abbi.1998.0770.
We have previously demonstrated that H2O2 downregulates CYP1A1 and CYP1A2 transcription in isolated rat hepatocytes (C. W. Barker, et al., 1994, J. Biol. Chem. 269, 3985-3990). In the present study, induction of chloramphenicol acetyltransferase (CAT) expression driven by 3.1 kb of rat CYP1A1 upstream regulatory sequences was suppressed by 56% in Hepa-1 cells treated with H2O2. Similarly, H2O2 inhibited CAT expression from vectors containing two copies of either xenobiotic-response element (XRE) 1 or XRE2. H2O2 did not inhibit basal CAT expression in cells that were not treated with the inducer beta-napthoflavone. Electrophoretic mobility shift assays demonstrated that the suppression of XRE-dependent transcription by H2O2 was not due to changes in nuclear aryl hydrocarbon (Ah) receptor DNA binding activity. Several types of experiments indicated that modulation of XRE enhancer strength by various means could modify H2O2-dependent suppression of CAT expression. Conditions that increased the transactivation potential of the Ah receptor (increase in XRE copy number or shortening of the distance between XREs and the minimal CYP1A1 promoter) attenuated the action of H2O2, while conditions that reduced XRE-mediated transactivation potential (decrease in XRE copy number, increase of the distance between the XRE and the promoter, or reduction of the number of bound Ah receptors by lowering the concentration of inducer) potentiated the inhibitory action of H2O2.
我们之前已经证明，过氧化氢（H2O2）可下调分离的大鼠肝细胞中细胞色素P450 1A1（CYP1A1）和细胞色素P450 1A2（CYP1A2）的转录（C.W.巴克等人，1994年，《生物化学杂志》269卷，3985 - 3990页）。在本研究中，用H2O2处理的Hepa - 1细胞中，由3.1 kb大鼠CYP1A1上游调控序列驱动的氯霉素乙酰转移酶（CAT）表达诱导被抑制了56%。同样，H2O2抑制了含有两个拷贝的异生素反应元件（XRE）1或XRE2的载体的CAT表达。H2O2不抑制未用诱导剂β - 萘黄酮处理的细胞中的基础CAT表达。电泳迁移率变动分析表明，H2O2对XRE依赖性转录的抑制不是由于核芳烃（Ah）受体DNA结合活性的变化。几种类型的实验表明，通过各种方式调节XRE增强子强度可以改变H2O2对CAT表达的依赖性抑制。增加Ah受体反式激活潜能的条件（增加XRE拷贝数或缩短XRE与最小CYP1A1启动子之间的距离）减弱了H2O2的作用，而降低XRE介导的反式激活潜能的条件（减少XRE拷贝数、增加XRE与启动子之间的距离或通过降低诱导剂浓度减少结合的Ah受体数量）增强了H2O2的抑制作用。
