Cell Polarity in Epithelial to Mesenchymal Transition

描述（由申请人提供）：上皮向间充质转化（EMT）是一系列以细胞极性丧失、细胞运动性增强和细胞侵袭性增加为特征的事件。虽然这一过程对正常的胚胎发育至关重要，但在成人中，它的失调会促进转移。本研究的长远目标是了解上皮细胞向间充质细胞的转变如何引发细胞极性的丧失，以及这种去极化如何促进癌症的进展。核心假设是，转录抑制因子Snail是EMT的关键调节因子，它会影响顶基极性复合物crums3 - pals1 - patj和Par6-Par3-aPKC，而这些极性复合物的缺失会加速癌症的进展。这一假设是基于长期以来的观察，即上皮在许多癌症中出现去极化，以及来自模式生物的数据表明，正常上皮结构的丧失导致深度过度增生。该实验室和其他人之前的工作表明，碎屑和Par6极性复合物是形成紧密连接和维持细胞极性所必需的。基于这些观察结果，本提案的实验重点是EMT期间新的碎屑和Par6上皮极性复合物。具体目的是：(1)阐明蜗牛在上皮细胞中表达诱导的顶基极性复合物的特异性缺陷。该研究将研究Snail如何影响极性复合物的定位和蛋白质水平，它是否降低了紧密连接成分基因的表达，E-cadherin表达是否可以挽救极化缺陷，以及Snail水平升高是否与人类肿瘤样本中E-cadherin和crumss的抑制相关。(2)分析蜗牛诱导的上皮细胞去极化的生物学效应，并评估CrumbsS的选择性重新表达是否调节了适当的生物学功能。该研究将调查Snail是否会破坏与细胞极性密切相关的其他细胞过程，如有丝分裂纺锤体极排列和细胞迁移，CrumbsS的重新表达是否会逆转任何功能改变的表型，以及CrumbsS的特异性缺失是否会调节癌细胞的生长或迁移。生化，遗传，蛋白质组学和显微镜为基础的方法将用于解决这些目标。该项目与公共卫生的相关性在于，它将进一步加深我们对导致癌细胞迁移、侵袭和转移的精确分子机制的理解，并有可能确定癌症治疗的新靶点。

DESCRIPTION (provided by applicant): The epithelial to mesenchymal transition (EMT) is a program of events characterized by loss of cell polarity, enhanced cell motility, and increased cell invasion. While this process is essential for normal embryonic development, its deregulation in the adult promotes metastasis. The broad, long term objectives of this proposal are to understand how the epithelial to mesenchymal transition initiates the loss of cellular polarity and how this depolarization might contribute to cancer progression. The central hypothesis is that the transcriptional represser Snail, a key regulator of the EMT, affects the apico-basal polarity complexes Crumbs3-PALS1-PATJ and Par6-Par3-aPKC and that loss of these polarity complexes enhances cancer progression. This hypothesis is based on the long-appreciated observation that the epithelium appears depolarized in many carcinomas, and on data from model organisms demonstrating that loss of proper epithelial architecture results in profound hyperproliferation. Previous work from this laboratory and others has revealed that the Crumbs and Par6 polarity complexes are required for the formation of the tight junction and maintenance of cell polarity. Based on these observations, the experimental focus of this proposal is on the novel Crumbs and Par6 epithelial polarity complexes during EMT. The specific aims are to: (1) Elucidate the specific defects in apico-basal polarity complexes induced by Snail expression in epithelial cells. The study will examine how Snail affects the localization and protein levels of the polarity complexes, if it reduces the expression of the tight junction component genes, if E-cadherin expression can rescue the polarization defects, and if elevated levels of Snail correlate with the suppression of E-cadherin and CrumbsS in human tumor samples. (2) Analyze the biological effects of Snail-induced depolarization in epithelial cells and evaluate whether selective re- expression of CrumbsS modulates proper biological function. The study will investigate if Snail disrupts other cellular processes closely associated with cell polarity such as mitotic spindle pole alignment and cell migration, if re-expression of CrumbsS reverses any functionally altered phenotypes, and whether the specific loss of CrumbsS modulates cancer cell growth or migration. Biochemical, genetic, proteomic, and microscopy-based methods will be used to address these aims. The relevance of this project to public health is that it will further our understanding of the precise molecular mechanisms resulting in cancer cell migration, invasion, and metastasis and potentially identify new targets for cancer therapy.