Non-canonical Wnt/Dishevelled signaling and cancer cell malignancy

描述（由申请人提供）：本研究的总体目标是探讨非规范Wnt信号通过disheveled （Dvl）蛋白作用的机制，并验证该途径调节癌细胞侵袭并与恶性乳腺癌相关的假设。标准的Wnt/？-catenin途径有助于许多癌症的发生，但最近的证据表明？-catenin独立的或非典型的Wnt信号与恶性进展的后期有关。该通路在细胞极性和形态发生中也起着至关重要的作用。Wnt诱导的Dvl磷酸化为非典型Wnt信号提供了可靠的检测方法，并为非典型途径及其在恶性肿瘤中的功能的分子解剖提供了起点。通过测试对Wnt诱导的磷酸化具有抗性的Dvl突变体的特性，将研究Dvl磷酸化在Wnt信号传导中的功能意义。我们还将验证非典型Wnt信号抑制er阴性乳腺癌细胞侵袭的假设。这将通过细胞迁移和侵袭的体外分析，结合信号分析来实现。我们将探索Dvl磷酸化在细胞迁移中的作用，并确定在这种情况下激活的非规范Wnt信号的下游组分。鉴于Dvl蛋白在介导和调节Wnt信号传导中的关键作用，我们将通过共聚焦显微镜全面分析Dvl在非典型Wnt信号下亚细胞分布的变化。将采用蛋白质组学方法来识别与Dvl相互作用的蛋白质中的关键变化，这些变化是非规范Wnt信号的结果。此外，将产生转基因小鼠，其中乳腺中的非规范Wnt信号将以受调节的方式被激活。该信号对乳腺发育的影响将被确定，这些小鼠将与现有的乳腺癌小鼠模型杂交，以直接测试非规范Wnt信号在肿瘤向侵袭性和转移性癌症进展中的影响。由于有大量证据表明Wnt在癌症中的异常表达，并且大多数Wnt蛋白可引发非规范信号，因此这种尚不为人所知的非规范Wnt信号通路可能具有广泛的医学意义。所涉及的分子机制的表征将为操纵治疗目的的途径提供机会。

DESCRIPTION (provided by applicant): The overall objectives of this research are to investigate mechanisms of non-canonical Wnt signaling acting via Dishevelled (Dvl) protein, and to test the hypothesis that this pathway regulates cancer cell invasion and is relevant to malignant breast cancer. The canonical Wnt/?-catenin pathway contributes to the initiation of many cancers but recent evidence indicates that ?-catenin-independent, or non-canonical, Wnt signaling is relevant to later stages of malignant progression. This pathway also plays crucial roles in cell polarity and morphogenesis. Wnt-induced phosphorylation of Dvl provides a reliable assay for non-canonical Wnt signaling and a starting point for molecular dissection of the non-canonical pathway and its functions in malignancy. The functional significance of Dvl phosphorylation in Wnt signaling will be investigated by testing the properties of Dvl mutants that are resistant to Wnt-induced phosphorylation. We will also test the hypothesis that non-canonical Wnt signaling inhibits the invasion of ER-negative breast cancer cells. This will be achieved by in vitro assays of cell migration and invasion, in conjunction with signaling assays. The role of Dvl phosphorylation in cell migration will be explored, and we will identify downstream components of non-canonical Wnt signaling that are activated in this context. In view of the key role of Dvl protein in mediating and modulating Wnt signaling, a comprehensive analysis of changes in the subcellular distribution of Dvl in response to non-canonical Wnt signals will be performed by confocal microscopy. A proteomics approach will be taken to identify key changes in the proteins with which Dvl interacts as a result of non-canonical Wnt signals. In addition, transgenic mice will be generated in which non-canonical Wnt signaling will be activated in the mammary gland in a regulated manner. Effects of this signaling on mammary development will be determined and the mice will be crossed with existing mouse models of breast cancer to test directly the consequences of non-canonical Wnt signaling in tumor progression towards invasive and metastatic cancer. Since there is extensive evidence of aberrant Wnt expression in cancer, and most Wnt proteins can elicit non-canonical signals, it is likely that this poorly understood pathway of non-canonical Wnt signaling may be of widespread medical significance. Characterization of the molecular mechanisms involved will provide opportunities for manipulating the pathway for therapeutic purposes.