Transcriptional regulation of arylhydrocarbon receptor (AhR), Arnt and E2F genes on proliferation process in A549 cells by dioxin and its mechanism.

芳烃受体(AhR)是一种配体激活的转录因子，介导了2，3，7，8-四氯二苯偶氮-对二恶英(TCDD)及其相关化合物的一系列毒理和生物效应。AHR属于含有碱性-螺旋-环-螺旋/PAS结构域的转录因子家族。AHR的表达与肺癌的发生发展密切相关。事实上，我们发现AhR的过表达加速了A549细胞的细胞增殖，A549细胞是从人肺泡细胞癌建立的。我们还证实了A549细胞增殖的增加与A549细胞中与癌变相关的转录因子E2F基因表达的激活有关，其机制是通过AhR和AhR核转运蛋白(ArnT)的异源二聚体复合体的表达。在本研究中，我们探讨了E2F基因激活与AhR/ArnT复合体表达关系的机制，通过S…获得了AhR、ArnT和E2F基因表达缺失的细胞系A549细胞中更多的iRNA转染技术。抑制AhR、Arnt和E2F的表达可降低E2F依赖的转录活性。为了确定与E2F依赖的转录活性有关的AhR和Arnt基因的结构域，将一系列与荧光素酶活性相关的缺失构建体导入A549细胞。AhR和Arnt序列的PAS区是激活E2F转录所必需的。其次，利用含有XRE序列的报告基因作为AhR/Arnt结合区，研究了E2F的表达对AhR和Arnt转录活性的影响。抑制E2F的表达增强了AhR/Arnt复合体对XRE序列的转录活性。AhR/Arnt复合体的相互作用需要E2F序列中的Rb蛋白结合区。Rb蛋白是癌变的抑制因子。进一步，我们用免疫沉淀法证明了AhR/Arnt复合体与E2F蛋白形成了络合物。这些结果表明AhR/Arnt和E2F复合体与E2F结合序列的结合促进了细胞的增殖，而与XRE序列的复合体结合则作为基因转录的抑制因子而受到影响。较少

Arylhydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates a spectrum of toxicological and bio-logical effects of 2,3,7,8-tetrachlorodiben.zo-p-dioxine (TCDD) and related compounds. AhR is included in a family of basic-helix-loop-helix/PAS domain-containing transcription factors. AhR expression is associated with the development of human lung carcinoma. In fact, we found that the overexpression of AhR accelerated the cell proliferation of A549 cells, established from a human alveolar cell carcinoma. We also demonstrated that the increased cell proliferations in A549 cells were associated with the activation of E2F gene expression, a transcription factor related in carcinogenesis, by the expression of heterodimer complex of AhR and AhR nuclear translocator (Arnt). In this study, we investigated the mechanisms of relationship of E2F gene activation and expression of AhR/Arnt complex.We obtained the deleted cell lines of AhR, Arnt and E2F genes expression by s … More iRNA transfection technique in A549 cells. E2F-dependent transcription activity was decreased by the suppression of AhR, Arnt and E2F expression. To identify the domains of AhR and Arnt genes responsible for E2F-dependent transcription activity, a series of deletion constructs linked the luciferase activity were transfected into A549 cells. The PAS regions of AhR and Arnt sequences were required for the activation of E2F transcription. Next, an effect of E2F expression to the transcription activity of AhR and Arnt was determined by using of reporter assay contained XRE sequences, as AhR/Arnt-binding regions. The AhR/Arnt complex-dependent transcription activity on XRE sequences was increased by the suppression of E2F expression. The Rb protein-binding regions in E2F sequences were required on the interaction of AhR/Arnt complex. Rb protein is a repressive factor of carcinogenesis. Furthermore, we demonstrated that AhR/Arnt complex was complexed with E2F protein using a immuno-precipitation assay. These results suggested that AhR/Arnt and E2F complex binding on E2F-binding sequences was accelerated the cell proliferation, while the complex binding on XRE sequences was affected as a repressor in gene transcription. Less