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Exploring the Potential of β-Catenin O-GlcNAcylation by Using Fluorescence-Based Engineering and Imaging

Abstract : Monitoring glycosylation changes within cells upon response to stimuli remains challenging because of the complexity of this large family of post-translational modifications (PTMs). We developed an original tool, enabling labeling and visualization of the cell cycle key-regulator β-catenin in its O-GlcNAcylated form, based on intramolecular Förster resonance energy transfer (FRET) technology in cells. We opted for a bioorthogonal chemical reporter strategy based on the dual-labeling of β-catenin with a green fluorescent protein (GFP) for protein sequence combined with a chemically-clicked imaging probe for PTM, resulting in a fast and easy to monitor qualitative FRET assay. We validated this technology by imaging the O-GlcNAcylation status of β-catenin in HeLa cells. The changes in O-GlcNAcylation of β-catenin were varied by perturbing global cellular O-GlcNAc levels with the inhibitors of O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). Finally, we provided a flowchart demonstrating how this technology is transposable to any kind of glycosylation.
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https://hal.univ-lille.fr/hal-03028555
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Soumis le : jeudi 28 janvier 2021 - 15:28:59
Dernière modification le : mercredi 3 novembre 2021 - 05:32:39

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Angelina Kasprowicz, Corentin Spriet, Christine Terryn, Vincent Rigolot, Stephan Hardivillé, et al.. Exploring the Potential of β-Catenin O-GlcNAcylation by Using Fluorescence-Based Engineering and Imaging. Molecules, MDPI, 2020, 25 (19), pp.4501. ⟨10.3390/molecules25194501⟩. ⟨hal-03028555v2⟩

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