Distribution of a major surface-associated glycoprotein,fibronectin, in cultures of adherent cells

Fibronectin was present in media and cell layers of cultures of adherent cells from human skin, kidney, lung, chest wall, liver, and heart. Cell-surface fibronectin, visualized by immunofluorescence, was in dense fibrillar (cultures from lung), discrete fibrillar (e.g., cultures from skin), or punctate (some cultures from kidney) structures. The subunit sizes of cell-surface fibronectin and fibronectin soluble in medium appeared identical in sodium dodecyl sulfate-polyacrylamide gels. To explain the polymorphism of cell-surface fibronectin, there must be chemical differences among the fibronectins synthesized by different cell strains or factors in the cell layer which influence fibronectin binding and aggregation.     

Calcium indirectly regulates immunochemical reactivity and functional activities of the N-domain of thrombospondin-1.

Conformational changes induced in thrombospondin-1 by removal of calcium regulate interactions with some ligands of its N-modules. Because calcium binds primarily to elements of the C-terminal signature domain of thrombospondin-1, which are distant from the N-modules, such regulation was unexpected. To clarify the mechanism for this regulation, we compared ligand binding to the N-modules of thrombospondin-1 in the full-length protein and recombinant trimeric thrombospondin-1 truncated prior to the signature domain. Three monoclonal antibodies were identified that recognize the N-modules, two of which exhibit calcium-dependent binding to native thrombospondin-1 but not to the truncated trimeric protein. These antibodies or calcium selectively modulate interactions of fibronectin, heparin, sulfatide, alpha3beta1 integrin, tumor necrosis factor-alpha-stimulated gene-6 protein, and, to a lesser extent, alpha4beta1 integrin with native thrombospondin-1 but not with the truncated protein. These results indicate connectivity between calcium binding sites in the C-terminal signature domain and the N-modules of thrombospondin-1 that regulates ligand binding and functional activities of the N-modules.     

Assembly of extracellular matrix.

A great challenge in understanding how different extracellular matrices assemble is to sort through the vast number of possible interactions between and among matrix molecules. The most profound insights are likely to come from patients with defined defects of matrix molecules and the use of transgenic mice or other experimental technologies that mimic the complexity of the human system.