Propolypeptide of von Willebrand Factor Is a Novel Ligand for Very Late Antigen-4 Integrin

doi:10.1074/jbc.272.13.8447

Propolypeptide of von Willebrand Factor Is a Novel Ligand for Very Late Antigen-4 Integrin*

From the ^‡ Department of Biological Sciences, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226, Japan, the
^§ Pharmaceutical Research Department, Sumitomo Metal Industries, Ltd., Kyoto 619-02, Japan, the
^¶ Institute for Protein Chemistry, Osaka University, Osaka 565, Japan, and the
^∥ Department of Vascular Biology, The Scripps Research Institute, La Jolla, California 92037

**To whom correspondence should be addressed:
Dept. of Biological Sciences, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226, Japan
. Tel.: 81-45-924-5728; Fax: 81-45-924-5808; E-mail: ysaito{at}bio.titech.ac.jp

Abstract

We have previously reported that propolypeptide of von Willebrand factor (pp-vWF) promotes melanoma cell adhesion in a β1 integrin-dependent manner. In this report, we identified the α subunit of the cell adhesion receptor for pp-vWF as α4. Human leukemia cell lines that express α4β1 integrin (very late antigen-4, VLA-4), but not cell lines which lack VLA-4, attached well to pp-vWF substrate and these adhesions were completely inhibited by anti-α4 integrin monoclonal antibody HP2/1. Adhesion of mouse melanoma expressing α4 integrin was also inhibited by anti-mouse α4 mAb PS/2. Furthermore, transfection of human α4 cDNA into α4⁻ Chinese hamster ovary cells resulted in an acquisition of adhesive activity to pp-vWF, indicating that pp-vWF is a ligand for VLA-4 integrin. Using a recombinant fragment of pp-vWF, the cell attachment site was shown to be located within amino acid residues 376-455 of pp-vWF. A series of synthetic peptides covering this region were tested for the ability to promote cell attachment and a 15-residue peptide designated T2-15 (DCQDHSFSIVIETVQ, residues numbered 395-409) promoted VLA-4 dependent cell adhesion. 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 found that α4β1 integrin complex from extracts of surface iodinated B16 cells specifically bound to the peptide. These results strongly suggest that pp-vWF is a novel physiological ligand for VLA-4.

Footnotes

↵* This work was supported in part by General Scientific Research Grant-in-Aid 06454644 from the Ministry of Education, Science, and Culture of Japan and by grants from the Ito Memorial Foundation, the Nissan Science Foundation, and the Kanagawa Academy of Science and Technology. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

pp-vWF

propolypeptide of von Willebrand factor

CS1

type III connecting segment region 1

mAb

monoclonal antibody

CHO

Chinese hamster ovary

GST

glutathione S-transferase

PAGE

polyacrylamide gel electrophoresis.
↵²F. Shimizu and J. Takagi, unpublished observations.
- Received November 7, 1996.