REGULATION OF BETA-CATENIN NUCLEARIZATION

β-连环蛋白核化的调控

• 批准号: 7020692
• 负责人: CHARLES A. ETTENSOHN
• 金额: $ 23.98万
• 依托单位: CARNEGIE-MELLON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2008-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7020692
• 关键词: REGULATION; BETA; CATENIN; NUCLEARIZATION

DESCRIPTION (provided by applicant): The accumulation of beta-catenin in the nuclei of specific blastomeres is a critical early step in animal development. In all deuterostomes (echinoderms and chordates/vertebrates) that have been studied, accumulation of beta-catenin in the nuclei of specific blastomeres along the axis of the cleavage stage embryo is one of the first indications of embryonic polarity. This differential "nuclearization" of beta-catenin is essential for proper germ layer formation and the establishment of inductive centers. The sea urchin embryo is a powerful model system for studying beta-catenin nuclearization. The embryo develops externally and is optically transparent, facilitating the application of a wide variety of light optical technologies. The system is unique in that specific cell types can be isolated from early embryos in large quantities. Most of the molecules of the beta-catenin pathway have been cloned from sea urchin and a large number of molecular biological tools are available. Gene expression can be manipulated by expression of dominant negative constructs and by microinjection of "morpholino" antisense oligonucleotides. The proposed work will identify the mechanisms that underlie beta-catenin nuclearization in early embryos. There are two major aims: 1) Using GFP-tagged proteins and time-lapse, 3-D confocal microscopy, the dynamics and turnover of key regulators of beta-catenin degradation will be studied in vivo. We will measure the half-life of beta-catenin in specific blastomeres of the early embryo, which will provide essential information concerning the cellular and molecular mechanisms of differential nuclearization. Through blastomere isolation experiments, we will test the role of cell-cell interactions in regulating beta-catenin nuclearization. 2) We will test three major models of GSK3 regulation along the embryo axis. In addition, using dominant negative constructs and morpholinos, we will examine the developmental function of two regulators of GSK3 activity, disheveled and Akt/PKB.

描述（由申请人提供）：β-catenin在特定胚胎的核中的积累是动物发育的重要早期一步。在已经研究的所有氘核抑制剂（棘皮动物和脊椎动物）中，沿特定胚胎的核中β-catenin的积累沿特异性胚胎的核沿切割阶段胚胎的轴轴是胚胎极性的第一个迹象之一。 β-catenin的这种差异“核化”对于适当的生殖层形成和归纳中心的建立至关重要。海胆胚胎是研究β-catenin核化的强大模型系统。该胚胎在外部发展，并且在光学上是透明的，从而促进了多种轻型光学技术的应用。该系统是独一无二的，因为可以大量从早期胚胎中隔离特定的细胞类型。 β-catenin途径的大多数分子都是从海胆中克隆的，并且提供了大量分子生物学工具。基因表达可以通过显性负构建体的表达和对“形态上”反义寡核苷酸的显微注射来操纵。拟议的工作将确定早期胚胎中β-catenin核化基础的机制。有两个主要目的：1）使用GFP标记的蛋白质和延时，3-D共聚焦显微镜，将在体内研究β-catenin降解的关键调节剂的动力学和周转。我们将在早期胚胎的特定胚泡中测量β-catenin的半衰期，该胚胎将提供有关差异核化细胞和分子机制的重要信息。通过胚泡分离实验，我们将测试细胞 - 细胞相互作用在调节β-连冠核化中的作用。 2）我们将测试沿胚胎轴的三种主要模型GSK3调节。此外，使用主要的负构建体和形态学，我们将研究GSK3活性的两个调节剂，Disheveled和Akt/PKB的发育功能。