Competitive binding of vascular cell adhesion molecule-1 and the HepII/IIICS domain of fibronectin to the integrin alpha 4 beta 1.

Abstract

The integrin receptor alpha 4 beta 1 binds to two different ligands, the extracellular matrix glycoprotein fibronectin and the endothelial cell surface protein vascular cell adhesion molecule-1 (VCAM-1). Using probes derived from each ligand and a variety of cell adhesion and ligand-receptor binding assays, we have investigated the relationship between the mechanisms of fibronectin and VCAM-1 interaction with alpha 4 beta 1. CS1 peptide, which represents the dominant active site from the HepII/IIICS recognition domain in fibronectin, was found to inhibit VCAM-1-dependent adhesion in three different assays: MOLT-4 T lymphoblastic leukaemia cell attachment to immobilized recombinant soluble VCAM-1 (rsVCAM-1), MOLT-4 cell attachment to monolayers of VCAM-1-transfected COS-1 cells, and A375-SM melanoma cell spreading on immobilized rs VCAM-1. Half-maximal inhibition required CS1 concentrations of 1.7-3.0 mg/ml, some 3-7-fold higher than that needed to autoinhibit adhesion to CS1-IgG conjugate. Using a more sensitive solid-phase receptor-ligand binding assay, CS1 was found to be a potent inhibitor of the binding of rsVCAM-1 to alpha 4 beta 1 (half-maximal inhibition at 13 micrograms/ml). In agreement with cell-based assays, severalfold lower concentrations of CS1 were required to inhibit binding of recombinant HepII/IIICS region of fibronectin (half-maximal inhibition at 3 micrograms/ml). VCAM-1-alpha 4 beta 1 binding was blocked not only by CS1 peptide but also by the recombinant HepII/IIICS region of fibronectin. Kinetic analysis of CS1 inhibition of VCAM-1 binding revealed that it was directly competitive in nature, indicating that VCAM-1 and fibronectin recognize either identical or spatially overlapping binding sites on alpha 4 beta 1. The implications of these results for the future design of VCAM-1 antagonists are discussed.

• Authors: R Makarem, P Newham, J A Askari, L J Green, J Clements, M Edwards, M J Humphries, A P Mould
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 269 Issue 6 Pg. 4005-11 (Feb 11 1994) ISSN: 0021-9258 [Print] United States
• PMID: 7508437 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Cell Adhesion Molecules
• Fibronectins
• Integrin alpha4beta1
• Integrins
• Ligands
• Peptides
• Receptors, Fibronectin
• Vascular Cell Adhesion Molecule-1

Topics

• Amino Acid Sequence
• Binding Sites
• Binding, Competitive
• Cell Adhesion
• Cell Adhesion Molecules (metabolism)
• Fibronectins (metabolism)
• Humans
• In Vitro Techniques
• Integrin alpha4beta1
• Integrins (metabolism)
• Kinetics
• Ligands
• Molecular Sequence Data
• Peptides (chemistry, metabolism)
• Protein Binding
• Receptors, Fibronectin (metabolism)
• Vascular Cell Adhesion Molecule-1