Differential cellular responses to adhesive interactions with galectin-8 and fibronectin coated substrates

The mechanisms underlying the cellular response to extracellular matrices (ECM), consisting of multiple adhesive ligands, each with distinct properties, are still poorly understood. Here we address this topic by monitoring the cellular responses to two very different extracellular adhesion molecules - fibronectin and galectin-8 - and to mixtures of the two. Fibronectin is one of the major integrin ligands, inducing cell spreading and development of focal adhesions associated with contractile stress fibers. Galectin-8 is a mammalian lectin, which specifically binds to {beta}-galactoside residues present on some integrins, as well as to other cell surface receptors. We found marked differences in HeLa-JW cell spreading, assembly of focal adhesions and actomyosin stress fibers, and formation of adherent filopodia, on rigid flat substrates functionalized by fibronectin or galectin-8 alone, or by mixtures of these two proteins. Spreading on galectin-8 resulted in a larger projected cell area compared to that on fibronectin, by more extensive formation of filopodia, coupled with an inability to activate focal adhesion and stress fiber assembly. These differences could be partially reversed by experimental manipulations of small G-proteins of the Rho family and their downstream targets, such as formins, the Arp2/3 complex, and Rho kinase. Another factor affecting the spreading process was shown to be the enhanced physical adhesion of the cells to galectin-8, as compared to fibronectin. Notably, at least one process, the formation of adherent filopodia, was synergistically upregulated by both ligands, so filopodia development on the substrate coated with a mixture of fibronectin and galectin-8 was far more prominent than on each ligand alone.