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bioRxiv · 10.64898/2026.02.05.704096

Galectin-3 binds to the RGD-binding site in a glycan-independent manner and to the allosteric site and activates integrins αvβ3, αIIbβ3, and α5β1

Takada, Y. K.·Wan, Y.-J. Y.·takada, y.
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Abstract

Galectin-3 (Gal3) is one of the most pro-inflammatory proteins and a biomarker of inflammatory diseases and cancer. Previous studies showed that Gal3 binds to v and {beta}1 integrins but it is unclear how Gal3 binds to integrins. Here, we show that Gal3 bound to soluble v{beta}3 and IIb{beta}3 integrins in 1 mM Mn2+ in cell-free conditions in a glycan-independent manner. Docking simulation predicts that Gal3 binds to the classical RGD-binding site (site 1) of v{beta}3, but the predicted Gal3-binding site does not include galactose-binding site. RGDfV or eptifibatide inhibited Gal3 binding to v{beta}3 and IIb{beta}3, respectively, but lactose, pan-galectin inhibitor, did not inhibit Gal3 binding to integrins. Point mutations of the predicted site 1 binding interface of Gal3 effectively inhibited Gal3 binding to site 1. Site 2 is involved in pro-inflammatory signaling (e.g., TNF and IL-6 secretion) and we previously showed that pro-inflammatory cytokines (e.g., CCL5 and TNF) bind to site 2 and allosteric integrin activation. Docking simulation predicts that Gal3 binds to site 2 of v{beta}3 and 5{beta}1. We found that Gal3 induced allosteric activation of soluble integrins v{beta}3, IIb{beta}3, and 5{beta}1 in 1 mM Ca2+ in cell-free conditions. Point mutations in the predicted site 2-binding interface inhibited Gal3-induced integrin activation, suggesting that Gal3 binding to site 2 is required for Gal3-induced integrin activation. Known anti-inflammatory agents, Ivermectin, NRG1, and FGF1 inhibited integrin activation induced by Gal3 in v{beta}3 and IIb{beta}3. These findings suggest that Gal3 binding to site 2 may be a potential mechanism of pro-inflammatory and pro-thrombotic action of Gal3.

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Takada, Y. K., Wan, Y.-J. Y., takada, y.. 2026-02-07. Galectin-3 binds to the RGD-binding site in a glycan-independent manner and to the allosteric site and activates integrins αvβ3, αIIbβ3, and α5β1. https://doi.org/10.64898/2026.02.05.704096

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