Study of gene expression on toxicity of dioxin in cultured cells

二恶英在培养细胞中毒性的基因表达研究

• 批准号: 11672231
• 负责人: TEZUKA Masakatsu
• 金额: $ 1.73万
• 依托单位: Nihon University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11672231/
• 关键词: TCDD; uPA mRNA; 3T3-L1 cells; adipocyte differentiation; clonal expansion; C; EBPα; PPARγ2; p42; p44; Rbタンパク質; p107; Ahレセプター

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a highly toxic compound that has recently attracted much attention as an environmental contaminant, elicits a variety of toxic responses. Most of the toxic effects of TCDD are thought to result from alteration of gene expression. In this study, we investigated the trans-acting factors involved in TCDD-dependent mRNA stabilization in a rat liver cytoplasm after a single dose administration of TCDD.UV-crosslinking study showed that the cytoplasmic protein of 50 kDa (p50) selectively recognized the 3' untranslated region of the urokinase-type plasminogen activator (uPA) mRNA.We also showed that the activation of p50 by TCDD is mediated through a protein phosphorylation cascade but not via de novo protein synthesis.We previously reported that a level of arylhydrocarbon receptor (AhR) protein decreased with ongoing adipose differentiation in 3T3-L1 cells. The AhR is the receptor for TCDD and related compounds. Studies using a TCDD-resistant clone … More of 3T3-L1 cells suggested that the AhR may be involved in the negative regulation of adipose differentiation. To confirm this hypothesis, 3T3-L1 fibroblast cells were stably transfected with a vector expressing high levels of full length sense AhR mRNA, antisense AhR mRNA, or a control vector. Comparison of the differentiation potency of these clones with that of control cells showed that overexpression of the AhR suppressed morphological differentiation as well as inductionof adipocyte-related genes, whereas decreased expression of the AhR induced much greater morphological differentiation and expression of adipocyte-related genes. Activation of C/EBPα and PPARγ2 restored the ability of the AhR-overexpressing cells to differentiate. The cells overexpressing the AhR exhibited the higher p42/p44 MAPkinase activity compared with the control cells. We also showed that activation of the AhR slowed clonal expansion. These results strongly suggest that AhR is a negative regulator of adipose differentiation in 3T3-L1 cells. Less

2,3,7,8-四氯二苯并-对二恶英（TCDD）是一种高毒性化合物，近年来作为一种环境污染物引起了广泛关注。TCDD的大多数毒性作用被认为是由基因表达的改变引起的。在这项研究中，我们研究了单剂量TCDD后大鼠肝细胞质中参与TCDD依赖性mRNA稳定的反式作用因子。紫外交联研究表明，50 kDa的细胞质蛋白（p50）选择性识别尿激酶型纤溶酶原激活物（uPA） mRNA的3'非翻译区。我们还发现TCDD对p50的激活是通过蛋白磷酸化级联介导的，而不是通过从头合成蛋白介导的。我们之前报道过，在3T3-L1细胞中，芳烃受体（AhR）蛋白水平随着脂肪分化而下降。AhR是TCDD及相关化合物的受体。更多3T3-L1细胞的研究表明，AhR可能参与脂肪分化的负调控。为了证实这一假设，3T3-L1成纤维细胞稳定地转染了表达高水平全长有义AhR mRNA、反义AhR mRNA的载体或对照载体。将这些克隆与对照细胞的分化能力进行比较发现，过表达AhR可抑制细胞形态分化和脂肪细胞相关基因的诱导，而过表达AhR可诱导细胞形态分化和脂肪细胞相关基因的表达。激活C/EBPα和PPARγ2可以恢复ahr过表达细胞的分化能力。与对照细胞相比，过表达AhR的细胞表现出更高的p42/p44 MAPkinase活性。我们还发现AhR的激活减缓了克隆的扩增。这些结果强烈提示AhR是3T3-L1细胞脂肪分化的负调节因子。少

项目成果

S.Shimba, M.Tezuka et al.: "2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) Induces Binding of a 50 kDa Protein on the 3' Untranslated Region of Urokinase-Type Plasminogen Activator mRNA"Biochem.Biophys.Res.Commun.. 272. 441-448 (2000)

S.Shimba、M.Tezuka 等人：“2,3,7,8-四氯二苯并-对-二恶英 (TCDD) 诱导 50 kDa 蛋白与尿激酶型纤溶酶原激活剂 mRNA 3 非翻译区结合”Biochem

S.Shimba,M.Tezuka et al.: "2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) Induces Binding of a 50 kDa Protein on the 3'Untranslated Region of Urokinase-Type Plasminogen Activator mRNA"Biochem.Biophys.Res.Commun.. 272. 441-448 (2000)

S.Shimba、M.Tezuka 等人：“2,3,7,8-四氯二苯并-对-二恶英 (TCDD) 诱导 50 kDa 蛋白与尿激酶型纤溶酶原激活剂 mRNA 的 3 非翻译区结合”Biochem