RGD-CAP (βig-h3) enhances the spreading of chondrocytes and fibroblasts via integrin α 1β 1

Abstract

In previous studies, RGD-CAP (collagen-associated protein containing the RGD sequence) isolated from a collagen fiber-rich fraction of pig cartilage was found to be orthologous to human βig-h3, which is synthesized by lung adenocarcinoma cells in response to transforming growth factor-β. In the present study, we examined the effect of recombinant chick RGD-CAP on the spreading of chondrocytes and fibroblasts using RGD-CAP-coated dishes. When rabbit articular chondrocytes, chick embryonic sternal chondrocytes, rabbit peritoneal fibroblasts or human MRC5 fibroblasts were seeded on plastic dishes coated with RGD-CAP, cell spreading was enhanced compared with that on control dishes (bovine serum albumin- or β-galactosidase-coated dishes). The effect of RGD-CAP on the cell spreading required divalent cations (Mg 2+ or Mn 2+), and was reduced by EDTA. Monoclonal antibodies (mAbs) to the human integrin α 1 or β 1 subunit, but not to the α 2, α 3, α 5 or β 2 subunits, suppressed the RGD-CAP-induced spreading of human MRC5 fibroblasts. In a parallel experiment, the mAb to the α 5 subunit, but not the mAb to the α 1 subunit, suppressed fibronectin-induced spreading of these cells. These findings suggest that RGD-CAP is a novel ligand for integrin α 1β 1 that dose not bind to the RGD motif. Accordingly, an RGD-CAP fragment, which carries a deletion in the C-terminal region containing the RGD motif, was still capable of stimulating cell spreading.