Identification of cell-binding site of angiomodulin (AGM/TAF/Mac25) that interacts with heparan sulfates on cell surface.
Angiomodulin (AGM/TAF/mac25) is a 30-kDa glycoprotein that was identified as an integrin-independent cell adhesion protein secreted by human bladder carcinoma cells. AGM is highly accumulated in small blood vessels of tumor tissues. In the present study, we attempted to identify the cell surface receptor and the cell-binding site of AGM using ECV-304 human vascular endothelial cells and BALB/c3T3 mouse fibroblasts. Heparin, heparan sulfate, and dextran sulfate, but not chondroitin sulfate, inhibited both adhesion of the two cell lines to AGM-coated plates and binding of AGM to these cells. Treatment of cells with heparinase, but not chondroitinase, inhibited both cell adhesion to AGM and AGM binding to cells. These results strongly suggested that heparan sulfates are the major receptor for AGM. Furthermore, we determined a 20-amino acid sequence within AGM molecule as its major cell-binding site. The synthetic peptide for the cell-binding sequence showed cell adhesion activity comparable to that of AGM, and the activity was inhibited by heparin and heparan sulfate. The peptide competitively inhibited cell adhesion to AGM and the binding of AGM to cells. These results indicated that AGM binds to cells through interaction of the identified cell-binding sequence with heparan sulfates on cell surface. It was also found that the heparan sulfate-binding peptide inhibited the formation of capillary tube-like structures of vascular endothelial cells in culture.
Sato
J
J
Division of Cell Biology, Kihara Institute for Biological Research, Yokohama City University, Yokohama 244-0813, Japan.
Hasegawa
S
S
Akaogi
K
K
Yasumitsu
H
H
Yamada
S
S
Sugahara
K
K
Miyazaki
K
K
D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line; D016264:Dextran Sulfate / Q000494:pharmacology; D003911:Dextrans / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D005347:Fibroblasts / Q000737:chemistry / Q000187:drug effects; D006025:Glycosaminoglycans / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006497:Heparan Sulfate / Q000737:chemistry* / Q000494:pharmacology; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D009363:Neoplasm Proteins / Q000737:chemistry*; D018919:Neovascularization, Physiologic / Q000502:physiology; D010455:Peptides / Q000494:pharmacology; D011485:Protein Binding
D016264:Dextran Sulfate; D003911:Dextrans; D006025:Glycosaminoglycans; D006026:Glycoside Hydrolases; D006497:Heparan Sulfate; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D010455:Peptides; C086009:tumor-derived adhesion factor