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.
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D017077:Culture Media, Conditioned; D004926:Escherichia coli; D006493:Heparin / Q000494:pharmacology; D006706:Homeostasis; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000378:metabolism* / Q000494:pharmacology*; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology*; D007700:Kinetics; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D010006:Osteoblasts; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology
D017077:Culture Media, Conditioned; D007328:Insulin; D018972:Insulin-Like Growth Factor Binding Protein 3; D018974:Insulin-Like Growth Factor Binding Protein 4; D010446:Peptide Fragments; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II; D006493:Heparin