%0 Journal Article %A Eugene Joeh %A Timothy O’Leary %A Weichao Li %A Richard Hawkins %A Jonathan R. Hung %A Christopher G. Parker %A Mia L. Huang %T Mapping glycan-mediated galectin-3 interactions by live cell proximity labeling %D 2020 %R 10.1101/2020.05.09.058586 %J bioRxiv %P 2020.05.09.058586 %X Galectin-3 is a glycan-binding protein (GBP) that binds β-galactoside glycan structures to orchestrate a variety of important biological events, including the activation of hepatic stellate cells to cause hepatic fibrosis. While the requisite glycan epitopes needed to bind galectin-3 have long been elucidated, the cellular glycoproteins that bear these glycan signatures remain unknown. Given the importance of the three-dimensional arrangement of glycans in dictating GBP interactions, strategies that allow the identification of GBP receptors in live cells, where the native glycan presentation and glycoprotein expression are preserved, possess significant advantages over static and artificial systems. Here, we describe the integration of a proximity labeling method and quantitative mass spectrometry to map the glycan and glycoprotein interactors for galectin-3 in live hepatic stellate cells. Understanding the identity of the glycoproteins and defining the structures of the glycans required for galectin-3 mediated hepatic stellate cell activation will empower efforts to design and develop selective therapeutics to mitigate hepatic fibrosis.Significance Because of the weak interactions between individual glycan-binding proteins (GBP), such as galectin-3, and glycans, strategies that allow the direct interrogation of these interactions in living cells remain limited. Thus, the glycan and glycoprotein ligands that are physiologically relevant for galectin-3 binding are insufficiently described. Here, we used a proximity labeling approach that catalytically tags interactors for galectin-3 and identified its pertinent glycan and glycoprotein counter-receptors in live hepatic stellate cells. This study demonstrates that proximity labeling is a powerful tool for mapping GBP complexes in living cells, and when coupled with chemical inhibitors, it can discriminate between protein-protein and protein-glycan interactions.Competing Interest StatementThe authors have declared no competing interest. %U https://www.biorxiv.org/content/biorxiv/early/2020/05/10/2020.05.09.058586.full.pdf