RETINOIC ACID-INDUCED HEPARIN-BINDING PROTEIN (RIHB) BINDS TO EMBRYONAL CHONDROCYTES AND CARTILAGE PRIMARILY VIA PROTEOGLYCANS

Retinoic acid-induced heparin binding protein (RIHB) is a highly basic , secreted polypeptide expressed during early chick embryogenesis. We have characterized the binding of I-125-labeled RIHB to embryonal chon drocytes in culture. No saturable, high-affinity binding can be observ ed on these cells. Furthermore, no I-125-labeled RIHB was internalized into the chondrocytes at 37 degrees C. The low-affinity binding of I- 125-labeled RIHB observed can be competed with another heparin binding factor, fibroblast growth factor 2, as efficiently as with unlabeled RIHB. The binding can also be almost completely inhibited by preincuba tion of the I-125-labeled RIHB with heparin or with a monoclonal antib ody which recognizes the heparin binding site of both RIHB and HBNF. W hen cross-linking experiments are performed with I-125-labeled RIHB, s pecific RIHB-containing high-molecular-weight complexes are observed; however, these represent only a very small fraction of the bound mater ial. Immunohistochemical analyses of embryonic wing cartilage demonstr ate that a significant fraction of bound RIHB can be removed from unfi xed tissue simply by rinsing with phosphate-buffered saline. The remai ning RIHB can be removed partially by incubation with heparitinase I o r III and completely when the incubation is performed with chondoitina se A, B, C. These results demonstrate that RIHB binds to embryonal. ch ondrocytes and cartilage primarily through proteoglycans of both hepar an sulfate and chondroitin sulfate types. (C) 1995 Academic Press, Inc .