Miles, J orcid.org/0000-0001-8839-9201 and van Oosten-Hawle, P orcid.org/0000-0002-5713-6457 (2020) Tissue-Specific RNAi Tools to Identify Components for Systemic Stress Signaling. Journal of Visualized Experiments, 159. e61357. ISSN 1940-087X
Abstract
Over the past decade there has been a transformative increase in knowledge surrounding the regulation of protein quality control processes, unveiling the importance of intercellular signaling processes in the regulation of cell-nonautonomous proteostasis. Recent studies are now beginning to uncover signaling components and pathways that coordinate protein quality control from one tissue to another. It is therefore important to identify mechanisms and components of the cell-nonautonomous proteostasis network (PN) and its relevance for aging, stress responses and protein misfolding diseases. In the laboratory, we use genetic knockdown by tissue-specific RNAi in combination with stress reporters and tissue-specific proteostasis sensors to study this. We describe methodologies to examine and to identify components of the cell-nonautonomous PN that can act in tissues perceiving a stress condition and in responding cells to activate a protective response. We first describe how to generate hairpin RNAi constructs for constitutive genetic knockdown in specific tissues and how to perform tissue-specific genetic knockdown by feeding RNAi at different life stages. Stress reporters and behavioral assays function as valuable readouts that enable the fast screening of genes and conditions modifying systemic stress signaling processes. Finally, proteostasis sensors expressed in different tissues are utilized to determine changes in the tissue-specific capacity of the PN at different stages of development and aging. Thus, these tools should help clarify and allow monitoring the capacity of PN in specific tissues, while helping to identify components that function in different tissues to mediate cell-nonautonomous PN in an organism.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2020 Journal of Visualized Experiments. This is an author produced version of an article published in Journal of Visualized Experiments. Uploaded in accordance with the publisher's self-archiving policy. |
Keywords: | Biology, Issue 159, proteostasis, tissue-specific, chaperone reporters, C. elegans, cell-nonautonomous, stress response, Transcellular chaperone signaling |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 12 Jun 2020 13:41 |
Last Modified: | 16 Nov 2020 01:43 |
Status: | Published |
Publisher: | Journal of Visualized Experiments (JoVE) |
Identification Number: | 10.3791/61357 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:161827 |