Regulation of Drosophila Epithelial Polarity

描述(申请人提供)：上皮细胞有一个独特的结构，包括一个根尖偏振的质膜，这对它们在胚胎发育和成年生理中的正常功能至关重要。上皮极性缺陷导致许多人类疾病，包括充血性心力衰竭和囊性纤维化，并与良性肿瘤和恶性肿瘤之间的转化有关。这个项目的长期目标是了解产生上皮极性的细胞和分子机制。虽然细胞培养中的研究已经确定了启动上皮极化的表面信号，但调节细胞内部机制的蛋白质的身份和机制尚不清楚，例如细胞骨架和蛋白质运输途径，以执行极性。为了确定这样的调控因子，我们在果蝇中使用基因筛查来寻找胚胎、幼虫或成虫阶段上皮结构所需的蛋白质。我们的筛查发现了一种名为Scribble(Scrib)的细胞连接定位蛋白，它对所有三个发育阶段的上皮细胞的极性都是必不可少的。Scrib还作为一种肿瘤抑制因子，揭示了Scrib与果蝇的极性和增殖控制有关。Scrib编码一个含有四个PDZ蛋白相互作用结构域的蛋白质，这表明它是一种支架蛋白。在许多与细胞极性有关的蛋白质中都发现了PDZ结构域，但PDZ介导的极化机制尚不清楚。本方案中的实验旨在通过分离与Scrib相互作用调节极性的蛋白质来揭示Scrib极化活性的机制。 其目的是：1)确定极性调节所需的Scrib相互作用结构域；2)生化分离Scrib复合体的新成员，并表征它们在上皮细胞中的功能；以及3)从基因上确定Scrib途径中的其他基因作为Scrib活性的修饰物。鉴于Scrib结构、功能和定位在系统发育过程中的高度保守性，我们的结果很可能为所有动物的上皮极性机制提供新的见解，并有助于我们理解上皮细胞结构丢失如何促进人类肿瘤的进展。

DESCRIPTION (provided by applicant): Epithelial cells have a distinct architecture, including an apico-basally polarized plasma membrane that is critical for their proper function in embryonic development and adult physiology. Defects in epithelial polarity contribute to many human diseases, including congestive heart failure and cystic fibrosis, and have been implicated in the transition between benign and malignant carcinomas. The long-term goal of this project is to understand the cellular and molecular mechanisms that generate epithelial polarity. While studies in cell culture have identified surface cues responsible for initiating epithelial polarization, the identity and mechanism of proteins that regulate the internal cellular machinery, such as the cytoskeleton and protein trafficking pathways, to execute polarity are not known. To identify such regulators we have used genetic screens in Drosophila to search for proteins required for epithelial architecture of the embryonic, larval, or adult stages. Our screens have identified a cell junction-localized protein called Scribble (Scrib) that is essential for the polarity of epithelia at all three stages of development. Scrib also acts as a tumor suppressor, revealing that Scrib links polarity and proliferation control in Drosophila. Scrib encodes a protein with four PDZ protein-protein interaction domains, suggesting that it acts as a scaffolding protein. PDZ domains are found in many proteins implicated in cell polarity, but the mechanism of PDZ-mediated polarization is not known. The experiments in this proposal are designed to uncover the mechanism of Scrib polarizing activity by isolating the proteins with which Scrib interacts to mediate polarity. The aims are: 1) Determine the Scrib interaction domains required for polarity regulation; 2) Biochemically isolate new members of the Scrib complex, and characterize their functions in epithelia; and 3) Genetically identify additional genes in the Scrib pathway as modifiers of Scrib activity. Given the strong conservation of Scrib structure, function, and localization across phylogeny, it is likely that our results will provide new insight into the mechanisms of epithelial polarity in all animals, and inform our understanding of how loss of epithelial cytoarchitecture may contribute to human tumor progression.