Identifying and characterizing new components for Wnt signaling

识别和表征 Wnt 信号传导的新组件

基本信息

批准号：
8868555
负责人：
Raymond Habas
金额：
$ 23.4万
依托单位：
TEMPLE UNIV OF THE COMMONWEALTH
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2017-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8868555
关键词：
Binding Biochemical Biological Biological Assay Biological Models Cell Polarity Cell Proliferation Cell membrane Cells Colon Carcinoma Complementary DNA Complex Congenital Abnormality Cultured Cells Cytoplasmic Protein Data Defect Development Developmental Process Disease Dsh protein Embryo Embryonic Development Family Felis catus Gene Targeting Glycoproteins Head Human Human Development Human Pathology Lead Ligands Malignant Neoplasms Mammalian Cell Mediating Membrane Methodology Molecular Nature Nuclear Translocation Organism Pathology Pathway interactions Pattern Formation Pilot Projects Play Principal Investigator Process Proteins Role Signal Pathway Signal Transduction Signal Transduction Pathway Signaling Molecule Testing Transducers Wnt proteins Xenopus base beta catenin cell motility gastrulation in vivo innovation insight interest malignant breast neoplasm migration novel programs public health relevance receptor screening tumorigenesis zygote

项目摘要

DESCRIPTION (provided by applicant): The development of a fertilized egg into a multicellular organism results from the interplay between several signaling pathways. One of these signaling pathways required for embryogenesis is the Wnt pathway and this pathway regulates cell fate determination, proliferation, migration and polarity. The Wnt ligand and its signaling components, in addition to regulating embryogenesis, are implicated in tumorigenesis and play causative roles in human colon and breast cancers as well as birth defect disorders. The elucidation of the mechanisms of Wnt signaling therefore remains vital for our understanding of the molecular mechanisms regulating embryogenesis and tumorigenesis. Studies have shown that Wnt signaling is initiated at the plasma membrane via the Frizzled (Fz) receptor along with the LRP5/6 co-receptors. Upon Wnt binding, signaling through the cytoplasmic protein Disheveled (Dvl) induces the stabilization and nuclear translocation of beta-catenin (beta-cat), which then regulates transcriptional induction of Wnt-target genes. This pathway is termed the canonical Wnt pathway. Additionally, there exists a beta-cat independent pathway termed the non-canonical pathway that functions to regulate cell polarity and motility. The non-canonical pathway is also mediated by the Fz protein and Dvl. However what determines how the Wnt signaling branches into these distinct pathways remains a poorly understood phenomenon. The role of Fz remains somewhat of a puzzle in terms of its ability to transduce Wnt signaling as only a handful of Fz-interacting proteins have been identified to date. Whether additional proteins that interact with Fz exist and their roles in modulating Wnt signaling remains unclear. We hypothesize that additional unidentified Fz-interacting proteins exist that play functional modulatory roles for transducing Wnt signaling. Towards this end, we have employed a novel membrane-based screening methodology to isolate Fz-interacting proteins and our preliminary studies have already shown excellent feasibility of this approach as we have isolated novel Fz-interacting proteins. We have further begun to functionally characterize these molecules using rapid and facile assays in Xenopus and our preliminary characterization reveals putative modulatory roles for some of these identified proteins. In this application, we propose two specific aims centered on expanding this screen and performing biochemical, cell biological and in-vivo characterization of the identified Fz-interacting proteins to delineate their roles in the Wnt signaling pathway. Our studies will utilize mammalian culture cells and the Xenopus embryo primarily as model systems. Our proposed studies together are directed towards uncovering and understanding the role of the novel Fz-interacting proteins as key transducers of Wnt signaling. Our preliminary data strongly supports that this screening methodology will provide important new components that will ultimately help to define the molecular nature of Wnt signaling from the plasma membrane to the intracellular milieu. Ultimately these studies can lead to a lucid understanding of how deregulated Wnt signaling results in cancer formation and birth defect disorders.

描述（由适用提供）：通过几种信号通路之间的相互作用，将受精卵发育到多细胞生物中。胚胎发生所需的这些信号通路之一是Wnt途径，该途径调节细胞脂肪的测定，增殖，迁移和极性。除调节胚胎发生外，Wnt配体及其信号成分在肿瘤发生中暗示，并在人类结肠和乳腺癌以及出生缺陷障碍中发挥致命作用。因此，阐明Wnt信号传导机制对于我们对调节胚胎发生和肿瘤发生的分子机制的理解仍然至关重要。研究表明，Wnt信号通过卷曲（FZ）受体以及LRP5/6共受体在质膜上启动。 WNT结合后，通过细胞质蛋白质的信号传导诱导β-catenin（beta-cat）的稳定和核易位，然后调节Wnt-Target基因的转录诱导。该途径称为规范的WNT途径。此外，存在一个beta-cat独立途径，该途径称为非规范途径，该途径可调节细胞极性和运动。非经典途径也由FZ蛋白和DVL介导。但是，决定Wnt信号分支如何进入这些不同的途径的原因仍然是一个鲜为人知的现象。 Fz的作用仍然是一个难题，因为它可以转换Wnt信号传导的能力，因为迄今为止仅确定了少数FZ相互作用的蛋白质。是否存在与FZ相互作用的其他蛋白质及其在调节Wnt信号传导中的作用尚不清楚。我们假设存在其他未鉴定的FZ相互作用蛋白，它们在传输WNT信号传导方面起起作用调节作用。为此，我们采用了一种新型的基于膜的筛选方法来分离FZ相互作用的蛋白质，并且我们的初步研究已经显示出这种方法的出色可行性，因为我们已经分离出了新型的FZ相互作用蛋白。我们已经开始在功能上使用快速和轻松的分析在Xenopus中进行功能表征这些分子，而我们的初步表征揭示了这些鉴定出的蛋白质中的一些假定的调节作用。在此应用中，我们提出了两个具体目标，该目标旨在扩展该屏幕并进行生化，细胞生物学和体内表征已鉴定的FZ相互作用蛋白我们的研究将利用哺乳动物培养细胞和爪蟾胚胎作为模型系统。我们提出的研究共同针对发现和理解新型FZ相互作用蛋白作为Wnt信号的关键传感器的作用。我们的初步数据强烈支持这种筛选方法将提供重要的新组件，最终将有助于定义从质膜到细胞内环境的Wnt信号的分子性质。最终，这些研究可能会导致对失控的Wnt信号传导如何导致癌症形成和先天缺陷障碍的清晰理解。