Adhesion dynamics in Drosophila border cell migration

描述（由申请人提供）： 细胞迁移是胚胎发育的一个重要特征，细胞迁移调控不当会导致出生缺陷和肿瘤转移。我们已经开发了一个简单的模型系统的一个正向遗传学的方法来研究细胞的运动在体内，一个子集的卵泡细胞，称为边缘细胞，在果蝇卵巢的迁移。我们已经确定，多种细胞外信号调节它们的运动。1)一种全局类固醇激素信号，蜕皮激素，通过蜕皮激素受体和称为Taiman的转录共激活因子起作用; 2）高度定位的细胞因子信号，其激活JAK/STAT途径;和3）几种生长因子，其通过EGFR和PVR受体酪氨酸激酶发出信号以引导细胞到达其目的地。此外，我们还研究了多种与边缘细胞迁移有关的细胞因子相关蛋白和细胞粘附分子。该提案探讨了细胞粘附在迁移边界细胞中动态调节的机制，这是细胞运动的一个重要方面，对于任何细胞类型都没有很好的理解。我们提出三个具体目标。第一个是调查的机制，管理贩运和稳定的E-钙粘蛋白，一个嗜同性细胞-细胞粘附分子，需要在边缘细胞和细胞上，他们迁移。为了测试E-钙粘蛋白在边缘细胞中的翻转速度是否比在非迁移毛囊细胞中更快，我们将采用先前表征的红色荧光蛋白变体，该变体会随着时间的推移而改变颜色，并与E-钙粘蛋白融合。我们将研究EGFR和PVR信号传导是否通过β-连环蛋白/Armadillo上特定酪氨酸残基的磷酸化使细胞粘附不稳定。我们还将确定内吞作用是否是重要的调节细胞表面E-钙粘蛋白在迁移的边缘细胞。我们将测试果蝇膜突蛋白是否有助于E-钙粘蛋白动力学。我们将探讨肌球蛋白VI有助于E-cadherin动态边缘细胞的机制。在第二个具体的目标，我们建议详细研究RhoGAP 93 B有助于边缘细胞迁移的机制，通过研究其生化活性，其表达模式，亚细胞定位，致死表型及其调控。最后，我们建议研究一个假定的下游目标的Rho在边缘细胞，rhophilin的突变体表型，上位性分析与Rho和识别和表征相互作用的蛋白质。

DESCRIPTION (provided by applicant): Cell migration is a fascinating feature of embryonic development, and improperly regulated cell migration contributes to birth defects and tumor metastasis. We have developed a simple model system for a forward genetic approach to the study of cell motility in vivo, the migration of a subset of follicle cells, known as border cells, in the Drosophila ovary. We have established that multiple extracellular signals regulate their movement. 1) a global steroid hormone signal, ecdysone, acting through the ecdysone receptor and a transcriptional coactivator called Taiman; 2) a highly localized cytokine signal, which activates the JAK/STAT pathway; and 3) several growth factors, which signal through the EGFR and PVR receptor tyrosine kinases to guide the cells to their destination. In addition we have studied a variety of cytoskeleton-associated proteins and cell adhesion molecules that function in border cell migration. This proposal explores the mechanisms by which cell adhesion is dynamically regulated in migrating border cells, an important aspect of cell motility that is not well understood for any cell type. We propose three specific aims. The first is to investigate the mechanisms that govern trafficking and stability of E-cadherin, a homophilic cell-cell adhesion molecule that is required in border cells and in the cells upon which they migrate. To test whether E-cadherin is turned over more rapidly in border cells than in non-migrating follicle cells, we will employ a previously characterized variant of the red fluorescent protein that changes color over time, fused to E-cadherin. We will investigate whether EGFR and PVR signaling destabilizes cell adhesion by phosphorylation of specific tyrosine residues on beta-catenin/Armadillo. We will also determine whether endocytosis is important for regulating cell surface E-cadherin in migrating border cells. We will test whether Drosophila moesin contributes to E-cadherin dynamics. And we will investigate the mechanisms by which Myosin VI contributes to E-cadherin dynamics in border cells. In the second specific aim we propose to investigate in detail the mechanisms by which RhoGAP93B contributes to border cell migration by studying its biochemical activity, its expression pattern, subcellular localization, lethal phenotype and its regulation. Finally we propose to study a putative downstream target of Rho in border cells, rhophilin by characterizing the mutant phenotype, epistasis analysis with Rho and by identifying and characterizing interacting proteins.