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A proximity-labeling proteomic approach to investigate invadopodia molecular landscape in breast cancer cells

Abstract : Metastatic progression is the leading cause of mortality in breast cancer. Invasive tumor cells develop invadopodia to travel through basement membranes and the interstitial matrix. Substantial efforts have been made to characterize invadopodia molecular composition. However, their full molecular identity is still missing due to the difficulty in isolating them. To fill this gap, we developed a non-hypothesis driven proteomic approach based on the BioID proximity biotinylation technology, using the invadopodia-specific protein Tks5α fused to the promiscuous biotin ligase BirA* as bait. In invasive breast cancer cells, Tks5α fusion concentrated to invadopodia and selectively biotinylated invadopodia components, in contrast to a fusion which lacked the membrane-targeting PX domain (Tks5β). Biotinylated proteins were isolated by affinity capture and identified by mass spectrometry. We identified known invadopodia components, revealing the pertinence of our strategy. Furthermore, we observed that Tks5 newly identified close neighbors belonged to a biologically relevant network centered on actin cytoskeleton organization. Analysis of Tks5β interactome demonstrated that some partners bound Tks5 before its recruitment to invadopodia. Thus, the present strategy allowed us to identify novel Tks5 partners that were not identified by traditional approaches and could help get a more comprehensive picture of invadopodia molecular landscape.
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https://www.hal.inserm.fr/inserm-02859860
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Submitted on : Monday, June 8, 2020 - 9:37:23 AM
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Sylvie Thuault, Claire Mamelonet, Joëlle Salameh, Kevin Ostacolo, Brice Chanez, et al.. A proximity-labeling proteomic approach to investigate invadopodia molecular landscape in breast cancer cells. Scientific Reports, Nature Publishing Group, 2020, 10 (1), pp.6787. ⟨10.1038/s41598-020-63926-4⟩. ⟨inserm-02859860⟩

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