The development of a functional assay system of  Wnt/β-catenin signal pathway

FENG Tao; et al

doi:10.13406/j.cnki.cyxb.2004.04.003

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The development of a functional assay system of Wnt/β-catenin signal pathway

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- The development of a functional assay system of Wnt/β-catenin signal pathway
- Journal of Chongqing Medical University Issue 4, Pages: 413-416(2004)
- 作者机构：
  
  1. 重庆医科大学基础医学院生物化学教研室
  2. 重庆医科大学生物医学工程系
  3. 芝加哥大学医学中心分子肿瘤研究室重庆4000163
  4. ,美国
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2004.04.003
  CLC： R341
- Published：2004，
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[1]冯涛,周兰,何通川.Wnt/β-连环蛋白信号途径活性的分析系统的建立[J].重庆医科大学学报,2004(04):413-416.

FENG Tao, et al. The development of a functional assay system of Wnt/β-catenin signal pathway. [J]. Journal of Chongqing Medical University (4):413-416(2004)
[1]冯涛,周兰,何通川.Wnt/β-连环蛋白信号途径活性的分析系统的建立[J].重庆医科大学学报,2004(04):413-416. DOI： 10.13406/j.cnki.cyxb.2004.04.003.

FENG Tao, et al. The development of a functional assay system of Wnt/β-catenin signal pathway. [J]. Journal of Chongqing Medical University (4):413-416(2004) DOI： 10.13406/j.cnki.cyxb.2004.04.003.

摘要

目的 :建立Wnt/ β -连环蛋白信号途径功能活性的分析系统。方法 :首先构建一个表达 β -连环蛋白 -绿色荧光蛋白的融合蛋白的载体

pCMV -bcGFP ;再用之转染HEK2 93细胞

用G4 18和LiCl筛选出稳定表达该融合蛋白的细胞

克隆建株并鉴定。结果 :成功获得稳定表达该融合蛋白的细胞株

即 2 93-bc -GFP细胞株 ;在荧光显微镜下以及蛋白质印迹分析证实 :所获得的 2 93-bc -GFP细胞株表达该融合蛋白的基础水平极低 ;用Wnt1和LiCl处理后可以明显地使该融合蛋白在细胞内的水平升高。结论 :上述结果提示该分析系统是以Wnt依赖的方式、受 β -连环蛋白调节的。该分析系统可以用作一个功能报告系统

以鉴定肿瘤发生中导致 β-连环蛋白信号途径失调的潜在的上游因子

也同样可以用于筛选那些特异抑制人类肿瘤中 β-连环蛋白信号途径活性的潜在的抗癌分子

Abstract

Objective:To develop a Wnt-dependent β-catenin-mediated functional assay system.Methods:To construct a vector

pCMV-bcGFP

which can express the β-catenin and green fluorescent protein(GFP) fusion protein;transfect HEK293 with the vector

select the cells with G418 and LiCl and establish the stable cell line

293-bc-GFP by cloning.Results:293-bc-GFP stable cell line was developed;the background expression of β-catenin-GFP fusion protein was detectable at a very low level in the cell line under fluorescent microscope and via Western blotting analysis;however

the fusion protein was showed to significantly increase in the presence of Wnt1 and lithium.Conclusion:Our results showed that the system is regulated by β-catenin in a Wnt-responsive fasion.Thus

it can be used as a functional reporter to identify potential upstream factors during tumorigenesis

as well as to screen for the potential anti-cancer agents that specifically inhibit β-catenin signaling in human tumors.

关键词

Wnt/β-连环蛋白信号途径β-连环蛋白-绿色荧光蛋白功能分析系统

Keywords

Wnt/β-catenin signaling pathwayβ-catenin and GFP fusion proteinFunctional assay system

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