Cell Polarity in Epithelial to Mesenchymal Transition

描述（由申请人提供）：上皮细胞向间质细胞转化（EMT）是一系列事件，其特征在于细胞极性丧失、细胞运动性增强和细胞侵袭增加。虽然这一过程对正常胚胎发育至关重要，但其在成人中的失调会促进转移。该提案的广泛，长期目标是了解上皮细胞向间充质细胞的转化如何引发细胞极性的丧失，以及这种去极化如何有助于癌症进展。核心假设是转录抑制因子Snail（EMT的关键调节因子）影响顶端-基底极性复合物Crumbs 3-PALS 1-PATJ和Par 6-Par 3-aPKC，并且这些极性复合物的丧失增强癌症进展。这一假说是基于长期以来的观察，即在许多癌中上皮出现去极化，以及来自模型生物的数据表明，适当的上皮结构的损失导致严重的过度增殖。该实验室和其他实验室的先前工作已经揭示，Crumbs和Par 6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，该提案的实验重点是EMT期间的新型Crumbs和Par 6上皮极性复合物。具体的目的是：（1）阐明上皮细胞中Snail表达诱导的顶端-基底极性复合体的特异性缺陷。该研究将检查Snail如何影响极性复合物的定位和蛋白水平，如果它减少了紧密连接组分基因的表达，如果E-cadherin表达可以挽救极化缺陷，以及如果Snail水平升高与人类肿瘤样本中E-cadherin和CrumbsS的抑制相关。(2)分析上皮细胞中Snail诱导的去极化的生物效应，并评估CrumbsS的选择性再表达是否调节适当的生物功能。该研究将调查Snail是否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的重新表达是否逆转任何功能改变的表型，以及CrumbsS的特异性丢失是否调节癌细胞生长或迁移。生物化学，遗传学，蛋白质组学和显微镜为基础的方法将被用来解决这些目标。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.