Adhesion of cultured bovine aortic endothelial cells to laminin-1 mediated by dystroglycan

Hisao Shimizu, Hiroshi Hosokawa, Haruaki Ninomiya, Jeffrey H. Miner, Tomoh Masaki

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Abstract

Expression of dystroglycan (DG) by cultured bovine aortic endothelial (BAE) cells was confirmed by cDNA cloning from a BAE cDNA library, Northern blotting of mRNA, Western blotting of membrane proteins, and double immunostaining with antibodies against βDG and platelet endothelial cell adhesion molecule-1. Immunocytochemical analysis revealed localization of DG in multiple plaques on the basal side of resting cells. This patchy distribution was obscured in migrating cells, in which the most prominent staining was observed in the trailing edge anchoring the cells to the substratum. Biotin-labeled laminin-1 overlay assay of dissociated BAE membrane proteins indicated the interaction of laminin-1 with αDG. The laminin α5 globular domain fragment expressed in bacteria and labeled with biotin could also bind αDG on the membrane blot, and the unlabeled fragment disrupted the binding of biotin-laminin-1 to αDG. The interaction of biotin- laminin-1 with αDG was inhibited by soluble αDG contained in the conditioned medium from DG cDNA-transfected BAE cells and by a series of glycosaminoglycans (heparin, dextran sulfate, and fucoidan). Soluble αDG in the conditioned medium inhibited the adhesion of BAE cells to laminin-1- coated dishes, whereas it had no effect on their adhesion to fibronectin. All three glycosaminoglycans that disrupted the biotin-laminin-1 binding to αDG inhibited BAE cell adhesion to laminin-1, whereas they failed to inhibit the adhesion to fibronectin. These results indicate a role of DG as a non- integrin laminin receptor involved in vascular endothelial cell adhesion to the extracellular matrix.

Original languageEnglish
Pages (from-to)11995-12000
Number of pages6
JournalJournal of Biological Chemistry
Volume274
Issue number17
DOIs
StatePublished - Apr 23 1999

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