Investigating the Establishment and Propagation of Planar Cell Polarity

项目摘要 平面细胞极性（PCP）是一个高度保守的途径，调节细胞极性的协调，沿着 单个组织平面，并且是发育和组织功能的基础。PCP的一个定义特征是 其核心成分的不对称定位，从均匀膜过渡到不对称膜 随着时间的推移分布。卷曲（Fz）与紊乱（Dvl）一起定位在细胞的一侧，而Vangl和 刺痛（Pk）定位于另一个。据认为跨膜组分Vangl和Fz募集相对的 细胞质组分Dvl和Pk介导反馈相互作用， 放大了细胞内的不对称性。因此，细胞间和细胞内核心PCP蛋白反馈相互作用 是我们认为PCP是如何建立的核心，然而，PCP背后的特定反馈相互作用 建立，以及它们是否主要发生在细胞内或细胞之间，还没有很好的理解。的 这项工作的总体目标是确定核心PCP蛋白如何通过反馈相互作用进行自组织， 协调极性的建立和传播。调查这些问题所需要的是 还原论系统，可以从细胞之间发生的反馈相互作用中分离出细胞内的反馈相互作用， 并将这些相互作用与组织水平极化线索的影响隔离开。我们的实验室已经建立了 小鼠皮肤作为一个强大的模型系统，研究PCP建立的保守机制。我们有 显示体外分离和培养的原代角质形成细胞显示自发的和局部排列的平面 在没有任何全局线索的情况下产生极性我建议进一步简化这个系统，使用微图案和细胞- 模仿接口以从其最小组件重构建立和传播极性。 使用实时成像，超分辨率显微镜和遗传操作的组合，我将确定 PCP蛋白相互作用驱动极性建立的分子机制， 传播这项研究将提供新的和创新的工具，将推动PCP领域向前发展。重要的是， 完成拟议的目标将通过提供洞察力来解决一个基本的、但人们还不太了解的问题 进入分子机制驱动PCP建立和揭示上皮细胞的基本特征， 生物学和组织组织。

Project Summary Planar cell polarity (PCP) is a highly conserved pathway that regulates the coordination of polarity in cells along a single tissue plane, and is fundamental in development and tissue function. A defining feature of PCP is the asymmetric localization of its core components, which transition from uniform to asymmetric membrane distribution over time. Frizzled (Fz) localizes with Dishevelled (Dvl) on one side of the cell, while Vangl and Prickle (Pk) localize to the other. It is thought that transmembrane components Vangl and Fz recruit opposing complexes between cells while the cytoplasmic components, Dvl and Pk, mediate feedback interactions that amplify asymmetry within cells. Thus, both intercellular and intracellular core PCP protein feedback interactions are central to how we think PCP is established, however, the specific feedback interactions underlying PCP establishment, and whether they primarily occur within cells or between cells, are not well understood. The overall goal of this work is to determine how the core PCP proteins self-organize via feedback interactions to coordinate the establishment and propagation of polarity. What is needed to investigate these questions is a reductionist system that can uncouple intracellular feedback interactions from those that occur between cells, and isolate these interactions from the influence of tissue level polarizing cues. Our lab has established the murine skin as a powerful model system to investigate conserved mechanisms of PCP establishment. We have shown that primary keratinocytes isolated and cultured in vitro display spontaneous and locally aligned planar polarity in the absence of any global cue. I propose to further simplify this system using micropatterning and cell- mimicking interfaces to reconstitute the establishment and propagation polarity from its minimal components. Using a combination of live imaging, super-resolution microscopy, and genetic manipulation, I will determine the molecular mechanisms underlying the PCP protein interactions that drive polarity establishment and propagation. This study will provide new and innovative tools that will push the PCP field forward. Importantly, completion of the proposed aims will address an essential, but poorly understood question, by providing insight into the molecular mechanisms driving PCP establishment and revealing fundamental features of epithelial biology and tissue organization.