Structural and biological characterization of bovine insulin-like growth factor binding protein-3.
Insulin-like growth factor binding protein-3 (IGFBP-3) purified from bovine serum shares 19 of 25 amino-terminal amino acid residues with IGFBP-3 purified from human, rat, and porcine sources. A newly characterized bovine fibroblast model was used to investigate the biological effects of purified bovine IGFBP-3 (bIGFBP-3). Coincubation of insulin-like growth factor I (IGF-I) with increasing concentrations of bIGFBP-3 produced a dose-dependent inhibition of IGF-I-stimulated [3H]aminoisobutyric acid (AIB) uptake in cultured bovine fibroblasts. Inhibition was complete at equimolar concentrations of IGF-I and bIGFBP-3. Inhibition of IGF-I-stimulated [3H]AIB uptake paralleled the ability of bIGFBP-3 to prevent [125I]IGF-I cell surface binding. In contrast, preincubation with bIGFBP-3 resulted in a dose-dependent enhancement of IGF-I-stimulated [3H]AIB uptake; a 32-86% increase in IGF-I bioactivity was seen after a 24 h preexposure to 10 nM bIGFBP-3, and a 2- to 6-fold potentiation was seen after a 72 h preincubation. Preincubation with bIGFBP-3 increased both the sensitivity and maximal responsiveness of the cells to IGF-I. The potentiating effects of bIGFBP-3 were associated with increased [125I]IGF-I binding to cultured bovine fibroblasts. Affinity cross-linking experiments indicated that the increase in IGF-I binding was due to increased membrane-associated bIGFBP-3 rather than to a bIGFBP-3-induced increase in type I IGF receptors. bIGFBP-3 had no effect on insulin stimulation of [3H]AIB uptake under either experimental condition. These data suggest that soluble bIGFBP-3 inhibits IGF-I action by sequestering and preventing IGF-I receptor binding, whereas surface-associated bIGFBP-3 enhances the growth-promoting effects of IGF-I in bovine fibroblasts. We propose that IGFBP-3 serves a dual function in modulating IGF action in vivo.
Endocrine Research Unit, Mayo Clinic, Rochester, Minnesota 55905.
C A
CA
Conover
M
M
Ronk
F
F
Lombana
D R
DR
Powell
D000595:Amino Acid Sequence; D000621:Aminoisobutyric Acids / Q000378:metabolism; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D004261:DNA Replication / Q000187:drug effects; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D018969:Insulin-Like Growth Factor Binding Proteins; D007334:Insulin-Like Growth Factor I / Q000378:metabolism / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology; D012689:Sequence Homology, Nucleic Acid; D012867:Skin / Q000378:metabolism
D000621:Aminoisobutyric Acids; D002352:Carrier Proteins; D018969:Insulin-Like Growth Factor Binding Proteins; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I