Genetic Analysis of Beta-Catenin Function in Human CA

G-连环蛋白的致癌激活在许多人类癌症的发病机制中是常见的。就像是 对于许多与癌症相关的基因，13-连环蛋白生物学中一些最有价值的研究一直是 在遗传易驯化的模式生物中进行，如果蝇和小鼠。通过 利用基因打靶方面的最新技术进步，现在可以进行类似的研究 人类癌细胞本身的基因功能。为了做到这一点，我们创造了高效的人类 无启动子的13-连环蛋白基因靶向载体，并利用它们创建人类癌症的等基因组 细胞的不同之处仅在于存在或不存在内源性突变体和/或野生型FS-catenin基因。 初步的功能研究揭示了13-连环蛋白在人类癌症中功能的几个方面。在……里面 特别是，我们已经证明了激活的13-连环蛋白是TCF介导的转录激活所必需的。 人类癌细胞。我们还证明了活化的g-连环蛋白的缺失会导致癌细胞 形成穹顶状结构，概括了分化Caco-2细胞单层时的表型。引航员 实验表明，致癌的13-连环蛋白不是人类结肠癌细胞形成所必需的 免疫缺陷小鼠的异种移植瘤。同样令人惊讶的是，我们没有发现明显的差异 野生型和突变型激活的13-连环蛋白的稳态水平和细胞内定位 经Western Blot和免疫细胞化学分析。最后，我们鉴定并鉴定了BMP-4，一种 利用Affymetrix在人类癌症中作为激活13-连环蛋白信号的假定效应物的转化生长因子-f3同源物 基因芯片分析。我们建议进一步利用这些同基因的人类癌细胞系统来研究 激活的13-连环蛋白在人类癌症发病机制中的作用。特别是，我们计划：1)确定 人癌细胞和组织中激活的13-连环蛋白的细胞生物学和病理生物学表型，2) 确定/3-连环蛋白的致癌激活是否是维持人类致瘤性所必需的 含有多个基因命中的癌细胞，3)确定g-连环蛋白的生化机制(S) 在人类癌症中的激活，以及4)识别和表征由激活的B-连环蛋白调节的基因 转录因子。

Oncogenic activation of g-catenin is common to the pathogenesis of a wide range of human cancers. As is the case for many cancer-related genes, some of the most valuable studies in 13-catenin biology have been undertaken in genetically tractable model organisms such as Drosophila melanogaster and Mus musculus. By exploiting recent technological advances in gene targeting it is now possible to undertake similar studies of gene function in human cancer cells themselves. To do this, we have created high-efficiency human promoterless 13-catenin gene-targeting vectors and employed them to create isogenic sets of human cancer cells that differ only in the presence or absence of their endogenous mutant and/or wild-type fS-catenin genes. Initial functional studies have shed light on several aspects of the function of 13-catenin in human cancer. In particular, we have shown that activated 13-catenin is necessary for TCF-mediated transcriptional activation in human cancer cells. We have also demonstrated that deletion of activated g-catenin causes cancer cells to form dome-like structures, recapitulating a phenotype seen in differentiating Caco-2 cell monolayers. Pilot experiments have suggested that oncogenic 13-catenin is not required for the human colon cancer cells to form xenograft tumors in immunodeficient mice. Also surprisingly, we have detected no discernible differences in the steady state levels and intracellular localization of wild-type and mutant, activated 13-catenin, as detected by Western Blot and immunocytochemistry analysis. Finally, we have identified and characterized BMP-4, a TGF-f3 homolog, as a putative effector of activated 13-catenin signaling in human cancer using Affymetrix Genechip analysis. We propose to further utilize these isogenic human cancer cell systems to study the functions of activated 13-catenin in the pathogenesis of human cancer. In particular, we plan to: 1) Determine the cell biological and pathobiologieal phenotypes of activated 13-catenin in human cancer cells and tissues, 2) Determine whether oncogenic activation of/3-catenin is required for maintenance of tumorigenicity in human cancer cells containing multiple genetic hits, 3) Determine the biochemical mechanism(s) of g-catenin activation in human cancer, and 4) Identify and characterize genes regulated by the activated B-catenin transcription factor.