Understanding developmental control of cell polarity using single-cell in vivo biochemistry

使用单细胞体内生物化学了解细胞极性的发育控制

• 批准号: 10665004
• 负责人: Daniel J Dickinson
• 金额: $ 37.18万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10665004
• 关键词: Age; Animals; Anterior; Apical; Biochemical; Biochemistry; Biological Models; Biological Process; Caenorhabditis elegans; Cell Cycle; Cell Polarity; Cell Separation; Cells; Characteristics; Complex; Cues; Data; Defect; Development; Disease; Disseminated Malignant Neoplasm; Embryo; Eukaryotic Cell; Foundations; Genetic; Goals; Homeostasis; In Vitro; Individual; Knowledge; Learning; Link; Malignant Neoplasms; Maps; Measurement; Membrane; Methods; Microscopy; Mission; Molecular; Mutagenesis; National Institute of General Medical Sciences; Outcome; Phosphotransferases; Play; Positioning Attribute; Prevention; Protein Dynamics; Protein Kinase; Proteins; Regulation; Reproducibility; Research; Role; Signal Pathway; Signal Transduction; Somatic Cell; Specific qualifier value; Stereotyping; System; Testing; Tissues; Work; age related; cell behavior; cell type; disease diagnosis; experimental study; extracellular; in vivo; in vivo Model; innovation; novel; polarized cell; prevent; programs; protein complex; protein protein interaction; response; single molecule; sperm cell; tool; zygote

Project Summary / Abstract Cell polarity is a fundamental feature of eukaryotic cells, and must be coordinated between cells and regulated to allow for normal animal development and tissue homeostasis. Despite genetic identification of proteins involved in cell polarity and a large body of knowledge about their interactions in vitro, it remains unclear how polarity proteins are organized into signaling complexes in cells. This lack of knowledge has prevented the field from understanding mechanisms of developmental control of polarity signaling in vivo. The long-term goal of the proposed research is to resolve the network of protein-protein interactions that supports animal cell polarity and to understand how this network can respond to developmental signals. To enable progress towards this goal, the applicants have developed innovative experimental tools that allow single-molecule measurements of native protein complex abundance in single cells. This project focuses on two evolutionarily conserved protein kinases, called aPKC and PAR-1, that play central roles in polarity by localizing to opposite ends of a polarized cell and dictating polarized cell behaviors. The applicants will make use of the C. elegans early embryo, in which cells reproducibly polarize in response to multiple spatial and temporal cues, to discover mechanistic links between developmental signals and the polarity machinery. The central hypothesis of this work is that that developmental signals control cell polarity by altering the molecular complexes in which aPKC and PAR-1 reside. This hypothesis will be explored by identifying dynamic aPKC and PAR-1 complexes that control polarity (Aim 1); by determining how polarity signaling is coordinated with cell cycle cues in the zygote (Aim 2); and by determining how developmental cues re-program polarity signaling in later embryos. The work proposed in this application is significant because it will reveal fundamental mechanisms controlling cell polarity, and because it places these mechanistic studies in a developmental context. The proposed work is innovative, in the applicant’s opinion, because it uses novel experimental methods to perform biochemical, mechanistic studies in vivo. By studying the biochemical control of aPKC and PAR-1 in multiple cellular and developmental contexts in a single experimental system, this work will identify fundamental mechanisms of PAR polarity signaling and to learn how these mechanisms are deployed to achieve different outcomes during development.

项目总结/摘要 细胞极性是真核细胞的一个基本特征，必须在细胞之间进行协调和调控 以允许正常的动物发育和组织稳态。尽管蛋白质的遗传鉴定 尽管它们参与细胞极性，并有大量关于它们在体外相互作用的知识，但仍不清楚它们是如何相互作用的。 极性蛋白在细胞中被组织成信号复合物。这种知识的缺乏阻碍了这一领域的发展。 了解体内极性信号的发育控制机制。 拟议研究的长期目标是解决蛋白质-蛋白质相互作用网络， 支持动物细胞极性，并了解这个网络如何对发育信号做出反应。到 为了实现这一目标，申请人开发了创新的实验工具， 单细胞中天然蛋白质复合物丰度的单分子测量。该项目的重点是 两种进化上保守的蛋白激酶，称为aPKC和PAR-1，它们在极性中发挥核心作用， 定位于极化细胞的相对端并指示极化细胞行为。申请人将 利用C. elegans早期胚胎，其中细胞可重复地响应多个空间和 时间线索，发现发育信号和极性机制之间的机械联系。 这项工作的中心假设是，发育信号通过改变细胞的极性来控制细胞的极性。 aPKC和PAR-1存在的分子复合物。这个假设将通过识别 控制极性的动态aPKC和PAR-1复合物（Aim 1）;通过确定极性信号如何 与受精卵中的细胞周期线索协调（目标2）;并通过确定发育线索如何重新编程 极性信号在后来的胚胎。这项申请中提出的工作是重要的，因为它将揭示 控制细胞极性的基本机制，因为它将这些机制研究放在一个 发展背景。在申请人看来，拟议的工作是创新的，因为它使用了新颖的 实验方法进行生物化学，机制研究在体内。通过研究生物化学控制 aPKC和PAR-1在多个细胞和发育背景下在一个单一的实验系统，这项工作 将确定PAR极性信号的基本机制，并了解这些机制是如何 在开发过程中实现不同的结果。