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Engineering of TurboID-Wingless for the identification of Wingless interactors through in vivo proximity labelling
Ana-Miruna Androniciuc1,2, Edward W Tate2,1, Jean-Paul Vincent1
1The Francis Crick Institute, London, England, United Kingdom.
Abstract:
Wnt signalling coordinates growth and cell fate decisions during development and mis-regulation of Wnt signalling in adults is associated with a range of conditions, including cancer and neurodegenerative diseases. Therefore, means of modulating Wnt proteins and/or cofactors could have significant therapeutic potential. As a first step towards enumerating the Wnt interactome, we devised an in vivo proximity labelling strategy to identify proteins that interact with Wingless (Wg), the main Drosophila Wnt. We engineered the wingless locus to express a functional TurboID-Wg fusion at endogenous levels and identified in vivo interactors by streptavidin pull-down from embryos, followed by mass spectrometry. Further analysis may in future extend the screen coverage and deliver functional validation of the newly identified interactors.
Insights
Researchers identified proteins interacting with Wingless (Wg), a key Wnt protein, using an in vivo proximity labeling method in Drosophila. This study is a crucial first step toward understanding the Wnt signaling interactome for potential therapeutic applications.
Area of Science:
- Developmental Biology
- Molecular Biology
- Biochemistry
Background:
- Wnt signaling is crucial for development and its dysregulation is linked to diseases like cancer.
- Modulating Wnt proteins offers therapeutic potential.
- Understanding the Wnt interactome is essential for targeted therapies.
Purpose of the Study:
- To identify proteins that interact with Wingless (Wg) in vivo.
- To establish a foundational Wnt interactome map.
- To explore novel therapeutic targets by understanding Wnt interactions.
Main Methods:
- Engineered the Drosophila wingless locus to express a functional TurboID-Wg fusion protein at endogenous levels.
- Utilized an in vivo proximity labeling strategy.
- Performed streptavidin pull-down assays followed by mass spectrometry on Drosophila embryos.
Main Results:
- Successfully identified in vivo protein interactors of Wingless (Wg).
- Established a novel method for Wnt interactome mapping.
- Generated a preliminary list of Wnt-interacting proteins.
Conclusions:
- The study provides a first step towards a comprehensive Wnt interactome.
- The employed proximity labeling strategy is effective for identifying in vivo protein interactions.
- Future work will focus on expanding screen coverage and functional validation of identified interactors.
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