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 and regulation of immune responses. 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 (3D) 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, have 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 human hepatic stellate cells and peripheral blood mononuclear cells. Understanding the identity of the glycoproteins and defining the structures of the glycans will empower efforts to design and develop selective therapeutics to mitigate galectin-3–mediated biological events.

At the surface of many cells is a compendium of glycoconjugates that form an interface between the cell and its surroundings; the glycocalyx. The glycocalyx serves several functions that have captivated the interest of many groups. Given its privileged residence, this meshwork of sugar-rich biomolecules is poised to transmit signals across the cellular membrane, facilitating communication with the extracellular matrix and mediating important signalling cascades. As a product of the glycan biosynthetic machinery, the glycocalyx can serve as a partial mirror that reports on the cell's glycosylation status. The glycocalyx can also serve as an information-rich barrier, withholding the entry of pathogens into the underlying plasma membrane through glycan-rich molecular messages. In this review, we provide an overview of the different approaches devised to engineer glycans at the cell surface, highlighting considerations of each, as well as illuminating the grand challenges that face the next era of ‘glyco-engineers’. While we have learned much from these techniques, it is evident that much is left to be unearthed.

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Galectin-3 is a glycan-binding protein (GBP) that binds b-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 threedimensional 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. SignificanceBecause 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 proteinglycan interactions. Graphical Abstract

Malignant myoepithelioma is a rare salivary gland carcinoma that might be confused with other neoplastic lesions, including carcinoma and sarcoma. A review of 10 previously reported acceptable cases and 2 new ones shows that the tumor affects older individuals with a mean age of 60 years. The parotid gland was the site of tumor in eight cases and the minor salivary glands of the mouth in four. Microscopically, 9 of the 12 tumors were composed either partly or predominantly of spindle-shaped cells. Plasmacytoid cells were the second most common cell type, and round or epithelioid cells were the least common type. Immunohistochemical evidence of myoepithelial differentiation in 10 evaluated cases included positive staining with antibodies against cytokeratin, muscle specific actin, S 100 protein, and vimentin. Ultrastructural features of a myoepithelial phenotype in nine studied cases were cytoplasmic microfilaments with focal densities, desmosomal cell junctions, basal lamina, and hemidesmosomal junctional plaques. Because of a limited number of reported cases and a paucity of follow-up data, it is not possible at present to accurately predict the clinical course of malignant myoepithelioma of salivary glands. However, it appears that local recurrence is an important cause of treatment failure. Metastasis is infrequent. Int J Surg Pathol 2(2): [133][134][135][136][137][138][139][140] 1994