LEF1增强Trop2启动子转录活性  

LEF1 Activates Transcription of Trop2 Gene Promoter

作　　者：漆彦斌[1] 刘叙翔 杨丽霞[1] 黄朝峰[1] 

机构地区：[1]中山大学中山医学院人类病毒学研究所,广州市510080

出　　处：《医学分子生物学杂志》2014年第3期155-160,共6页Journal of Medical Molecular Biology

基　　金：国家自然科学基金（No.31170831）,广东省引进创新科研团队计划（No.2009010058）

摘　　要：目的探究Wnt信号通路相关蛋白LEF1、TCF1、野生型β—catenin和激活型β—catenin对肿瘤相关钙信号转导因子Trop2启动子转录活性的调控作用，找出调控Trop2转录活性的转录因子并分析其可能的分子机制。方法通过生物信息学方法预测Trop2启动子上潜在的转录因子，构建小鼠Trop2启动子报告质粒，在293T细胞中，采用荧光素酶双报告系统分析LEF1、TCF1、野生型β-catenin和激活型β—catenin对Trop2启动子的调控作用。找出Trop2启动子上的TCF／LEF结合位点，通过定点突变的方法构建Trop2启动子TCF／LEF结合位点突变体，探究Trop2启动子转录活性调控可能的分子机制。结果经预测，Trop2启动子上存在LEF1、TCF1等众多的潜在转录因子结合位点。荧光素酶双报告实验证明。在293T细胞中。与空载体相比，LEF1、TCF1都不同程度地增强Trop2启动子转录活性。其中LEF1对Trop2启动子转录活性的增强作用最为明显，达108倍。突变Trop2启动子上的TCF／LEF结合位点可以明显降低LEF1对Trop2启动子的转录活性。结论在293T细胞中，LEF1可以极大地增强Trop2启动子的转录活性。LEF1上调Trop2启动子转录活性是通过结合到Trop2启动子上的两个TCF／LEF结合位点而发挥作用的。Objective To examine the role of Wnt pathway-related proteins LEF 1, TCF1, wild type β-catenin and actived β-catenin in the regulation of the transcription activity of tumor-associated calcium signal transducer 2 （ Trop2 ）, identify the transcription factors that can regulate the transcription activity of Trop2 and dissect the possible molecule mechanisms. Methods Bioinformatics methods were used to predict the potential transcription factors of Trop2 promoter. The mouse Trop2 promoter recombinant of pGL4 was constructed as a reporter plasmid. Then, the activation effects of LEF 1, TCF1, wild type β-catenin and actived β-catenin on the Trop2 promoter were analyzed by the dual fluorescent reporter system in 293 T cells. After identification of the TCF/LEF binding sites in the Trop2 promoter, the mutation in the TCF/LEF binding sites of the Trop2 promoter was induced by site-directed mutagenesis in order to examine the molecular mechanism invol- ving the regulation of the transcription activity of the Trop2 promoter. Results The binding sites of LEF1, TCF1 and many other transcription factors were found in the Trop2 promoter. The dual fluo- rescent reporter system showed that compared with the control vector, LEF 1 and TCF1 could upregulate the transcription activity of Trop2 promoter in 293 T cells. Especially, LEF-1 had the highest up-regulated effect on the tre.nscription activity of Trop2 promoter, 108 times that of the control. The mutation in the TCF/LEF binding sites of the Trop2 promoter could significantly abrogate the up-regulated activity of Trop2 promoter by LEF1. Conclusion LEF1 can up-regulate the transcription activity of Trop2 promoter in 293 T cells through the two TCF/LEF binding sites in the Trop2 promoter.

关 键 词：Trop2 LEF1 启动子 转录激活 

分 类 号：Q75[生物学—分子生物学]