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: 10455085
• 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/10455085
• 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 Kinase; Proteins; Regulation; Research; Role; Signal Pathway; Signal Transduction; Somatic Cell; Specific qualifier value; Stereotyping; System; Testing; Tissues; Work; age related; base; 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.

项目摘要/摘要 细胞极性是真核细胞的一个基本特征，必须在细胞间相互协调和调控 以保证动物的正常发育和组织的动态平衡。尽管蛋白质的基因鉴定 参与了细胞的极性和关于它们在体外相互作用的大量知识，目前尚不清楚如何 在细胞中，极性蛋白质被组织成信号复合体。这种知识的缺乏阻碍了这个领域的发展 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-蛋白质相互作用的网络， 支持动物细胞的极性，并了解这个网络如何对发育信号做出反应。至 为了使这一目标取得进展，申请者开发了创新的实验工具，使 单细胞天然蛋白质复合体丰度的单分子测量。这个项目的重点是 两个进化上保守的蛋白激酶，称为aPKC和PAR-1，通过 定位到极化细胞的两端，并决定极化细胞的行为。申请者将使 线虫早期胚胎的使用，在该胚胎中，细胞可复制地极化以响应多个空间和 时间线索，以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 APKC和PAR-1所在的分子复合体。这一假说将通过确定 控制极性的动态aPKC和PAR-1复合体(目标1)；通过确定极性信号的强弱 与受精卵中的细胞周期信号协调(目标2)；以及通过确定发育信号如何重新编程 后期胚胎中的极性信号。本申请中提出的工作具有重要意义，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。申请人认为，拟议的工作是创新的，因为它使用了新颖的 实验方法：在活体内进行生化、力学研究。通过对生化控制的研究 在一个单一的实验系统中，在多个细胞和发育环境中研究aPKC和PAR-1，这项工作 将识别PAR极性信号的基本机制，并了解这些机制是如何 为在发展过程中实现不同的结果而部署。