Wnt Pathway Regulation in Embryos and Cells

描述（由申请人提供）： Wnt信号通路在从早期胚胎发育的调节到干细胞生长的控制的多种生物过程中至关重要。这种途径的失调导致多种癌症，包括结肠癌，这是美国癌症相关死亡的第二大原因。调节该途径的中心是“破坏复合物”，其是包括激酶GSK 3的蛋白质的集合，其控制转录激活因子β-连环蛋白（Wnt信号传导的主要效应物）的水平。然而在许多细胞类型中，细胞间Wnt信号通过未知的机制调节破坏复合物的活性，在早期非洲爪蟾发育中，该复合物通过含有GSK 3结合蛋白GBP的易位颗粒在细胞内调节。 这项提案的一个主要目标将建立在我们上一个资助期的结果之上，即GBP与驱动蛋白轻链（KLC）相互作用，我们提出驱动蛋白轻链在易位所需的马达蛋白和Wnt途径的其他调节因子（如Dishevelled）之间提供了一个关键的桥梁。使用各种方法来破坏驱动蛋白的功能，我们将测试驱动蛋白在颗粒运动和轴形成中的作用。此外，为了了解颗粒是如何组装在非洲爪蟾卵母细胞，我们将测试的假设，酪蛋白激酶1 α（CK 1 α），可能激活的卷曲7受体，在颗粒形成中具有重要作用。最后，使用斑马鱼作为模型系统，我们将测试我们的命题，这些背侧颗粒用于其他低等脊椎动物。 研究的第二个主要领域将继续我们的Wnt细胞内通路的结构研究，重点是CK 1 β，一个激酶是必不可少的Wnt信号转导。我们将研究CK 1 β如何抑制其自身的活性，因为最近的研究表明，Wnt信号调节CK 1 β的自抑制。我们将解决CK 1 β与Dishevelled结合的结构，Dishevelled是CK 1 β活性的主要目标之一。最后，使用结构-功能方法，我们将确定Dishevelled是否是调节Wnt通路中CK 1 β活性的重要靶点。由于Wnt通路在脊椎动物发育和肿瘤发生中起着关键作用，这些研究将对提高出生缺陷和癌症的分子基础的认识非常重要。

DESCRIPTION (provided by applicant): The Wnt signaling pathway is of critical importance in a wide variety of biological processes from the regulation of early embryonic development to the control of stem cell growth. Misregulation of this pathway leads to a variety of cancers, including colon cancer, the second major cause of cancer-related death in the United States. Central to the regulation of this pathway is the "Destruction Complex", an assembly of proteins including the kinase GSK3, which controls the levels of the transcriptional activator beta-catenin, a principal effector of Wnt signaling. Whereas in many cell types intercellular Wnt signals regulate the activity of the Destruction Complex through an unknown mechanism, in early Xenopus development, the complex is regulated intracellularly by translocating particles containing a GSK3 binding protein, GBP. One major goal of this proposal will build on our result from the last funding period that GBP interacts with kinesin light chain (KLC), which we have proposed provides a critical bridge between a motor protein required for translocation and other regulators of the Wnt pathway such as Dishevelled. Using a variety of approaches to disrupt kinesin function, we will test the proposed role of kinesin in particle movement and axis formation. Furthermore, in order to understand how the particles are assembled in the Xenopus oocyte, we will test the hypothesis that casein kinase 1epsilon (CK1epsilon), potentially activated by the Frizzled 7 receptor, has an essential role in particle formation. Finally, using zebrafish as a model system, we will test our proposition that these dorsalizing particles are used in other lower vertebrates. The second major area of research will continue our structural studies of the Wnt intracellular pathway, focusing on CK1epsilon, a kinase that is essential for transducing Wnt signals. We will examine how CK1epsilon inhibits its own activity, since recent studies have shown that Wnt signaling regulates the autoinhibition of CK1epsilon. We will solve the structure of CK1epsilon bound to Dishevelled, one of the principle targets of CK1epsilon activity. Finally, using a structure-function approach, we will determine if Dishevelled is the essential target of CK1epsilon activity in regulating the Wnt pathway. Since the Wnt pathway plays key roles in vertebrate development and oncogenesis, these studies will be important in advancing knowledge of the molecular basis of birth defects and cancer.