Insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) functions as an IGF-reversible inhibitor of IGFBP-4 proteolysis.

Previous studies have shown that insulin-like growth factor (IGF)-binding protein-4 (IGFBP-4) is degraded only in the presence of exogenous IGFs; however, we found that cation-dependent proteinase activity present in conditioned medium of MC3T3-E1 osteoblasts degrades 125I-recombinant human (rh)IGFBP-4 in the absence of IGFs. Addition of IGF-I, IGF-II, or insulin to conditioned medium had little affect on 125I-rhIGFBP-4 proteolysis, while extraction of IGFs resulted in only a approximately 10% reduction in proteinase activity. Since factors other than IGFs appeared to be involved in regulating IGFBP-4 proteolysis, we hypothesized that IGFBP-3, an IGFBP produced by many cell lines, but not MC3T3-E1 cells, might function as an inhibitor of IGFBP-4 proteolysis. Addition of rhIGFBP-3 to conditioned media inhibited 125I-rhIGFBP-4 proteolysis by 90%, while IGF-I and IGF-II reversed the inhibitory effects of rhIGFBP-3 in a dose-dependent manner. 125I-rhIGFBP-4 proteolysis was not inhibited by N-terminal rhIGFBP-3 fragments that bind IGFs, but was inhibited by two synthetic peptides corresponding to sequences contained in the mid-region or C-terminal region of IGFBP-3. Both inhibitory peptides contain highly basic, putative heparin-binding domains and heparin partially reversed the inhibitory effects of rhIGFBP-3 on 125I-rhIGFBP-4 proteolysis. These data demonstrate that rhIGFBP-3 inhibits IGFBP-4-degrading proteinase activity and binding of IGFs or glycosaminoglycans to IGFBP-3 may induce conformational changes in the binding protein, causing disinhibition of the proteinase.