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    The Role of SRGAP2 in Vertebrate Gastrulation

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    Genre
    Thesis/Dissertation
    Date
    2016
    Author
    Wadsworth, Richard Lee
    Advisor
    Habas, Raymond
    Committee member
    Sheffield, Joel B.
    Balciunas, Darius
    O'Reilly, Alana
    Department
    Biology
    Subject
    Biology
    Biochemistry
    Biology, Molecular
    Daam1
    Srgap2
    Wnt
    Xenopus
    Permanent link to this record
    http://hdl.handle.net/20.500.12613/4003
    
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    DOI
    http://dx.doi.org/10.34944/dspace/3985
    Abstract
    During vertebrate gastrulation, an elaborate series of cellular motility events occur that define the three germ layers, and establishes the primary body axis. While it is known that non-canonical Wnt signaling plays a crucial role during this period of development, a complete picture of the molecular mechanisms controlling this pathway has yet to be established. Previous studies has shown that one essential component of the pathway linking the Wnt ligands to the cytoskeletal changes that occur during vertebrate development is Dishevelled-associated activator of morphogenesis (Daam1). Daam1 bridges the gap between Dishevelled (Dvl) and the small GTPase RhoA and is required for Wnt-dependent RhoA activation, but the biochemical details of this process are to date still undefined. To identify additional factors that might be involved in this process, a yeast two-hybrid screen using a C-terminal region of Daam1 was performed. During this screen SLIT-ROBO Rho GTPase-activating protein 2 (SRGAP2), was identified. The studies presented here were designed to establish a functional interaction between SRGAP2 and Daam1 during non-canonical Wnt signaling and to characterize the function of SRGAP2 during early vertebrate development. My studies uncover that SRGAP2 and Daam1 do indeed interact and that this interaction is positively influenced by Wnt stimulation. I also uncover via immunocytochemistry, that these two proteins share common sub-cellular localization patterns in HeLa cells that is responsive to Wnt stimulation. Further, I show that ectopic expression of SRGAP2 in HeLa cells has adverse effects on stress fiber formation in Wnt5a treated cells. And lastly, micro-injection experiments in Xenopus laevis show that over- or under-expression of SRGAP2 produces severe gastrulation defects in the developing embryos. These studies together demonstrate that SRGAP2 plays a critical role in regulating non-canonical Wnt signaling through its interaction with Daam1 and through regulation of the monomeric GTPases Rho. It also shows that SRGAP2 plays a functional role for gastrulation during early vertebrate development.
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