Dissecting the Molecular Mechanisms of Canonical Wnt Signaling

剖析经典 Wnt 信号转导的分子机制

• 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
• 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

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）：癌细胞的主要特征是细胞在其正常功能中不受管制的能力，随后经过快速分裂（肿瘤生长）和迁移行为的变化（肿瘤转移）。几种信号分子以及其信号通路的组成部分已被视为其中一个是Wnt蛋白的原因。有重要的证据表明Wnt信号在癌症中的作用是Wnt蛋白可以在Wnt信号传导途径（例如2-catenin）（例如2-catenin）中转化乳腺细胞和突变。实际上，当在乳腺中异常表达导致乳腺癌时，第一种Wnt蛋白Wnt-1被确定为一个因素。因此，我们建议鉴定分子成分并了解Wnt信号传导的机制对于我们对人类癌症形成的理解至关重要。 研究表明，通过细胞质蛋白质染色（DVL）的Wnt信号传导诱导2-catenin（2-cat）的稳定和核转运，然后调节Wnt-Target基因的转录诱导。但是，2-CAT核易位的调节仍然很差，并且对2-CAT转录活性的调节尚不完全理解。该建议针对WNT信号的新调制器的表征。我们已经通过使用DVL作为诱饵的酵母 - 两种杂交筛网确定了该分子称为DBP，用于染色的结合分子。在哺乳动物细胞中，DBP与Wnt信号通路的关键成分相互作用，包括DVL和酪蛋白激酶1（CK1），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，为了响应Wnt刺激，DBP主要位于核。此外，DBP影响2-CAT的亚细胞定位，这是Wnt-target基因转录诱导所需的关键因素。爪蟾发育过程中DBP的体内分析胚胎模式形成和DBP的耗竭可防止胚胎头部形成。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。我们提出了这项赠款建议中的两个具体目标，我们的研究针对DBP的生化，细胞生物学和体内表征，以描绘其作用方式以及最近确定的DBP效应子。这些研究将利用哺乳动物培养细胞和爪蟾胚胎作为模型系统。 为此，我们的研究旨在发现和理解Wnt信号通路的这两个新组成部分的作用。我们建议分析这些新组件将有助于定义从质膜到细胞核的Wnt信号的分子性质，并最终可以更好地理解对失控的Wnt信号如何导致癌症形成。 公共卫生相关性：了解人类癌症的发展仍然取决于识别有助于这种病理学的关键信号分子及其信号转导途径。 Wnt蛋白质是在人类癌症中起因果作用的一种键信号分子。我们的研究集中在理解我们已经在Wnt信号通路中确定功能的两个新分子，我们建议这些研究可以为癌症形成提供重要的新见解。