PROPOLYPEPTIDE OF VON-WILLEBRAND-FACTOR IS A NOVEL LIGAND FOR VERY LATE ANTIGEN-4 INTEGRIN

We have previously reported that propolypeptide of von Willebrand fact or (pp-vWF) promotes melanoma cell adhesion in a beta 1 integrin-depen dent manner. In this report, we identified the alpha subunit of the ce ll adhesion receptor for pp-vWF as alpha 4. Human leukemia cell lines that express alpha 4 beta 1 integrin (very late antigen-4, VLA-4), but not cell lines which lack VLA-4, attached well to pp-vWF substrate an d these adhesions were completely inhibited by anti-alpha 4 integrin m onoclonal antibody HP2/1. Adhesion of mouse melanoma expressing alpha 4 integrin was also inhibited by anti-mouse alpha 4 mAb PS/2. Furtherm ore, transfection of human alpha 4 cDNA into alpha 4(-) Chinese hamste r ovary cells resulted in an acquisition of adhesive activity to pp-vW F, indicating that pp-vWF is a ligand for VLA-4 integrin. Using a reco mbinant fragment of pp-vWF, the cell attachment site was shown to be l ocated within amino acid residues 376-455 of pp-vWF. A series of synth etic peptides covering this region were tested for the ability to prom ote cell attachment and a 15-residue peptide designated T2-15 (DCQDHSF SIVIETVQ, residues numbered 395-409) promoted VLA-4 dependent cell adh esion. The peptide was also capable of inhibiting cell adhesion to pp- vWF, suggesting that this sequence represents the cell attachment site , By affinity chromatography using peptide T2-15-Sepharose, it was fou nd that alpha 4 beta 1 integrin complex from extracts of surface iodin ated B16 cells specifically bound to the peptide. These results strong ly suggest that pp-vWF is a novel physiological ligand for VLA-4.