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    Characterization of Dishevelled Associated Activator of Morphogenesis 2 (Daam2) in Wnt Signaling During Early Embryonic Development

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    Genre
    Thesis/Dissertation
    Date
    2015
    Author
    Nama, Kaushik
    Advisor
    Habas, Raymond
    Committee member
    Gruberg, Edward R.
    Balciunas, Darius
    Runnels, Loren W.
    Department
    Biology
    Subject
    Developmental Biology
    Permanent link to this record
    http://hdl.handle.net/20.500.12613/3326
    
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    DOI
    http://dx.doi.org/10.34944/dspace/3308
    Abstract
    The Wnt signaling pathway is highly conserved in metazoan animals. Wnt signaling regulates an array of cellular processes that include motility, polarity, cell fate determination, primary axis formation and organogenesis and recently have been implicated in stem cell renewal. Deregulated Wnt signaling has tragic consequences for the developing embryo and is a causative factor for a number of pleiotropic human pathologies such as cancers of the breast, colon and skin, skeletal defects and human birth defect disorders including the most common human neural tube closure defect: spina bifida. Modulation of the actin cytoskeleton via the non-canonical Wnt signaling pathway mediate cell polarity and cell migration that are required for proper vertebrate gastrulation and subsequent neurulation. However, the mechanism(s) by how the non-canonical pathway mediates this actin cytoskeleton modulation is not fully understood. Identifying and characterizing novel signaling components of the non-canonical pathway remain essential to understand its role during embryogenesis. The Formin-homology protein Dishevelled associated activator of morphogenesis 1 (Daam1) was previously characterized as an essential component for non-canonical Wnt-dependent regulation of cytoskeletal reorganization and cell migration. Daam1 and Daam2 are members of the Daam family of proteins but the role of Daam2 in early embryonic development however remains conflicting as to whether it functions in the canonical or non-canonical Wnt signaling pathway. In this thesis, I cloned and functionally characterized the role of Xenopus Daam2 in the Wnt signaling pathway. Co-immunoprecipitation assays confirm the binding of Daam2 with Dishevelled (Dvl) as well as the domains within those proteins required for interaction. Interestingly the binding of Daam2 and Dvl was subject to Wnt regulation. Sub-cellular localization immunofluorescence studies using mammalian cell culture system reveal Daam2 is cytoplasmic and regulates the actin cytoskeleton by modulating the actin filament formation. In Xenopus embryos, Daam2 is temporally expressed at very low levels maternally and its expression increases during neurulation and remains subsequently elevated after neurulation. Daam2 expression is spatially refined to areas of highly dynamic actin reorganization such as mesoderm, notochord and neural tube. The knockdown of Daam2 in Xenopus embryos specifically produces neural tube closure defect suggesting a role in non-canonical signaling. Indeed studies examining the role of Daam2 in canonical Wnt signaling found no role for this protein in canonical Wnt signaling. These studies taken together identify Daam2 as an important Wnt signaling component that functions in the non-canonical Wnt signaling pathway and regulates neural tube morphogenesis.
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