Dissecting the Molecular Mechanisms of Canonical Wnt Signaling

剖析经典 Wnt 信号转导的分子机制

基本信息

批准号：
8499357
负责人：
Raymond Habas
金额：
$ 27.77万
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：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2014-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8499357
关键词：
Anterior Applications Grants Area Behavior Binding Biochemical Biological Biological Models Cell Culture Techniques Cell Nucleus Cell Proliferation Cell membrane Cells Characteristics Complex Cultured Cells Cytoplasm Cytoplasmic Protein Defect Development Developmental Process Embryo Embryonic Development Embryonic Pattern Specification Family Felis catus Fibrinogen Gene Targeting Glycoproteins Head Human Human Development In Vitro Lead Malignant Neoplasms Mammalian Cell Mammary gland Mediating Membrane Proteins Modeling Molecular Mutation Nature Neoplasm Metastasis Nuclear Nuclear Envelope Nuclear Translocation Pathology Pathway interactions Pattern Formation Phosphoproteins Phosphorylation Site Play Principal Investigator Process Proteins Regulation Role Signal Pathway Signal Transduction Signal Transduction Pathway Signaling Molecule System Testing Transcriptional Regulation Wnt proteins Work Xenopus Xenopus laevis Yeasts base cancer cell casein kinase I cell motility gain of function in vitro Assay in vivo insight loss of function malignant breast neoplasm mutant prevent programs protein function public health relevance receptor research study response trafficking tumor tumor growth tumorigenesis xenopus development yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): A central characteristic of cancer cells is the ability of the cells to become deregulated in their normal function and subsequently undergo rapid division (tumor growth) and changes in their migratory behavior (tumor metastasis). Several signaling molecules along with components of their signaling pathways have been implicated as causative factors for, one of these being Wnt proteins. There is important evidence for a role of Wnt signaling in cancers as Wnt proteins can transform mammary cells and mutations in components of the Wnt signaling pathway such as 2-Catenin have been shown to play causative roles in cancers in humans. In fact, the first Wnt protein, Wnt-1, was identified as a factor when aberrantly expressed in the mammary gland resulted in breast cancer. We therefore propose that identifying the molecular components and understanding the mechanisms of Wnt signaling remains crucial for our understanding of human cancer formation. Studies have shown that Wnt signaling through the cytoplasmic protein Dishevelled (Dvl) induces the stabilization and nuclear translocation of 2-catenin (2-cat), which then regulates transcriptional induction of Wnt-target genes. However, the regulation of 2-cat nuclear translocation remains poorly defined and also the regulation of 2-cat transcriptional activity is not fully understood. This proposal is directed towards the characterization of a new modulator of Wnt signaling. We have identified this molecule termed DBP, for Dishevelled Binding Molecule, via a yeast-two hybrid screen using Dvl as a bait. In mammalian cells, DBP interacts with key components of the Wnt signaling pathway including Dvl and Casein kinase 1 (CK1), and antagonizes Wnt-stimulated transcriptional induction. Cellular localization studies of DBP demonstrate that this protein is normally present in both the cytoplasmic and nuclear compartments of cells. However, in response to Wnt stimulation, DBP becomes predominantly localized to the nucleus. Furthermore, DBP influences the subcellular localization of 2- cat, a crucial factor required for the transcriptional induction of Wnt-target genes. In vivo analysis of DBP during Xenopus development perturbs embryonic pattern formation and depletion of DBP prevents embryonic head formation. Based on our preliminary studies, we hypothesize that DBP is an essential component required for Wnt signaling. We propose two specific aims in this grant proposal and our studies are directed towards a biochemical, cell biological and in-vivo characterization of DBP to delineate its mode of action along with a recently identified effector for DBP. These studies will utilize mammalian culture cells and the Xenopus embryo as model systems. Towards this end, our studies are directed towards uncovering and understanding the role of these two new components of the Wnt signaling pathway. We propose that analyzing these new components will help to define the molecular nature of Wnt signaling from the plasma membrane to the cell nucleus and ultimately lead to a better understanding of how deregulated Wnt signaling results in cancer formation. PUBLIC HEALTH RELEVANCE: Understanding the development of human cancers remains dependent on identifying key signaling molecules and their signal transduction pathways that contribute to this pathology. One key-signaling molecule that has been demonstrated to play causative roles in human cancer is the Wnt protein. Our studies are focused on understanding two new molecules that we have identified that function in the Wnt signaling pathway, and we propose that these studies can provide important new insights into cancer formation.

描述（由申请人提供）：癌细胞的一个核心特征是细胞正常功能失调并随后经历快速分裂（肿瘤生长）和迁移行为改变（肿瘤转移）的能力。一些信号分子及其信号通路的组成部分已被认为是致病因素，其中之一是 Wnt 蛋白。有重要证据表明 Wnt 信号传导在癌症中发挥作用，因为 Wnt 蛋白可以转化乳腺细胞，并且 Wnt 信号传导通路成分（例如 2-连环蛋白）的突变已被证明在人类癌症中发挥致病作用。事实上，第一个 Wnt 蛋白 Wnt-1 被确定为乳腺中异常表达导致乳腺癌的一个因素。因此，我们建议识别 Wnt 信号传导的分子成分并了解其机制对于我们了解人类癌症的形成仍然至关重要。研究表明，Wnt 信号通过细胞质蛋白 Disheveled (Dvl) 诱导 2-catenin (2-cat) 的稳定和核转位，然后调节 Wnt 靶基因的转录诱导。然而，2-cat 核易位的调节仍然不明确，并且 2-cat 转录活性的调节也尚未完全了解。该提案针对 Wnt 信号传导新调制器的表征。我们通过使用 Dvl 作为诱饵的酵母-两种杂交筛选，鉴定出了这种称为 DBP（散乱结合分子）的分子。在哺乳动物细胞中，DBP 与 Wnt 信号通路的关键成分（包括 Dvl 和酪蛋白激酶 1 (CK1)）相互作用，并拮抗 Wnt 刺激的转录诱导。 DBP 的细胞定位研究表明，该蛋白通常存在于细胞的细胞质和核区室中。然而，响应 Wnt 刺激，DBP 主要定位于细胞核。此外，DBP 影响 2-cat 的亚细胞定位，这是 Wnt 靶基因转录诱导所需的关键因素。非洲爪蟾发育过程中 DBP 的体内分析会扰乱胚胎模式的形成，而 DBP 的消耗会阻止胚胎头部的形成。根据我们的初步研究，我们假设 DBP 是 Wnt 信号传导所需的重要组成部分。我们在这项资助提案中提出了两个具体目标，我们的研究针对 DBP 的生化、细胞生物学和体内表征，以描述其作用模式以及最近确定的 DBP 效应子。这些研究将利用哺乳动物培养细胞和非洲爪蟾胚胎作为模型系统。为此，我们的研究旨在揭示和理解 Wnt 信号通路这两个新组件的作用。我们建议，分析这些新成分将有助于定义从质膜到细胞核的 Wnt 信号传导的分子性质，并最终更好地理解 Wnt 信号传导失调如何导致癌症形成。公共卫生相关性：了解人类癌症的发展仍然依赖于识别导致这种病理学的关键信号分子及其信号转导途径。 Wnt 蛋白是一种已被证明在人类癌症中发挥致病作用的关键信号分子。我们的研究重点是了解我们已经确定的在 Wnt 信号通路中发挥作用的两种新分子，我们认为这些研究可以为癌症形成提供重要的新见解。