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. ? ? ? ?
