Investigating the Establishment and Propagation of Planar Cell Polarity

项目摘要 平面细胞极性(PCP)是一种高度保守的途径，它调节细胞内极性的协调。 单个组织平面，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局部化，从均匀膜过渡到不对称膜 随着时间的推移进行分配。卷发(FZ)定位于细胞一侧的散乱(DVL)，而Vangl和 刺痛(PK)定位到另一个。跨膜成分Vang1和Fz被认为招募相反的 细胞之间的复合体，而细胞质成分DVL和PK，介导反馈相互作用 放大细胞内的不对称性。因此，细胞间和细胞内的核心PCP蛋白反馈相互作用 是我们认为PCP建立的核心，然而，PCP背后的特定反馈相互作用 建立，以及它们主要发生在细胞内还是细胞间，还不是很清楚。这个 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自组织来 协调极性的建立和传播。调查这些问题需要的是一个 一种还原系统，可以将细胞内的反馈作用与细胞之间发生的反馈作用分开， 并将这些相互作用从组织水平极化线索的影响中分离出来。我们的实验室已经建立了 小鼠皮肤作为研究PCP建立保守机制的有力模型系统。我们有 体外分离培养的原代角质形成细胞表现为自发的、局部排列的平面。 在没有任何全球线索的情况下出现两极。我建议进一步简化这个系统，使用微图案化和细胞- 模仿接口，以从其最小组件重新构建建立和传播极性。 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蛋白相互作用的分子机制驱动极性建立和 传播。这项研究将提供新的创新工具，推动PCP领域向前发展。重要的是 拟议目标的完成将通过提供洞察力来解决一个基本但鲜为人知的问题 探讨PCP形成的分子机制及揭示上皮细胞的基本特征 生物学和组织组织学。

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.