• Login
    View Item 
    •   Home
    • Brock Theses
    • Masters Theses
    • M.Sc. Biological Sciences
    • View Item
    •   Home
    • Brock Theses
    • Masters Theses
    • M.Sc. Biological Sciences
    • View Item
    JavaScript is disabled for your browser. Some features of this site may not work without it.

    Browse

    All of BrockUCommunitiesPublication DateAuthorsTitlesSubjectsThis CollectionPublication DateAuthorsTitlesSubjectsProfilesView

    My Account

    LoginRegister

    Statistics

    Display statistics

    Engineering Optogenetic Control of Endocytic Recycling: Controlling Rab11 Function in Drosophila melanogaster using Engineered Light-Responsive Nanobodies

    • CSV
    • RefMan
    • EndNote
    • BibTex
    • RefWorks
    Thumbnail
    Name:
    Brock_Ward_Devin_2019.pdf
    Size:
    36.11Mb
    Format:
    PDF
    Download
    Author
    Ward, Devin
    Keyword
    Rab11
    Notch
    Optogenetics
    Nanobodies
    
    Metadata
    Show full item record
    URI
    http://hdl.handle.net/10464/14504
    Abstract
    The regulated transport of materials in cells is an essential function of all living organisms. In eukaryotes, one main family of transport regulators is the Rab GTPases. Rab GTPases utilize GTP to move materials throughout the cell by binding to the membrane of vesicles or endosomes, and trafficking distinct, membrane-associated components throughout the cell. One member of this large family of proteins is Rab11. Rab11 is responsible for endosome recycling: returning membrane proteins and receptors from intracellular recycling endosomes to the cell membrane, where these membrane proteins and receptors may be reused. Although the exact mechanism of Rab11 trafficking is not known, Rab11 appears to be critical for the development and survival of many organisms. Drosophila mutants for the Rab11 gene are not viable, where lethality manifests during embryonic development. This early lethality has imposed significant limitations on elucidating the immediate effects of Rab11 inhibition. Thus, the goal was to engineer a novel method of inhibiting Rab11 in vivo in Drosophila melanogaster. Specifically, the goal was to generate a genomically non-invasive construct (Opto-Nanobody) utilizing an optogenetic, light-sensitive Cryptochrome 2 (Cry2) fused to YFP-targeting nanobodies to bind functional, endogenous, YFP-tagged Rab11. This system promises to provide precise light-responsive spatio-temporal control of Rab11 function in response to blue-light exposure through homo-oligomeric clustering, which has been shown to inhibit Rab-dependent trafficking. Using the Drosophila embryo as a model system, these tools were applied to directly determine the effects of Rab11 inhibition on Notch signaling, and to determine the mechanisms that govern Rab11 trafficking. The Opto-Nanobody was tested in vitro in S2 cells, and was shown to form homo-oligomeric clusters in the presence of blue light and demonstrated the ability to bind to YFP-Rab11. This Opto-Nanobody construct has been inserted into a D. melanogaster injection vector, so that the Opto-Nanobody may be inserted into the D. melanogaster genome, and used to control YFP::Rab11 activity in vivo to elucidate the role of Rab11 in Notch signalling.
    Collections
    M.Sc. Biological Sciences

    entitlement

     
    DSpace software (copyright © 2002 - 2022)  DuraSpace
    Quick Guide | Contact Us
    Open Repository is a service operated by 
    Atmire NV
     

    Export search results

    The export option will allow you to export the current search results of the entered query to a file. Different formats are available for download. To export the items, click on the button corresponding with the preferred download format.

    By default, clicking on the export buttons will result in a download of the allowed maximum amount of items.

    To select a subset of the search results, click "Selective Export" button and make a selection of the items you want to export. The amount of items that can be exported at once is similarly restricted as the full export.

    After making a selection, click one of the export format buttons. The amount of items that will be exported is indicated in the bubble next to export format.