Heparin-binding, highly basic regions within the thyroglobulin type-1 repeat of insulin-like growth factor (IGF)-binding proteins (IGFBPs) -3, -5, and -6 inhibit IGFBP-4 degradation.
MC3T3-E1 murine osteoblasts produce insulin-like growth factor (IGF)-binding protein-4 (IGFBP-4)-degrading proteinase activity, which is inhibited by IGFBP-3 and a highly basic, C-terminal domain of IGFBP-3. Of all the other five IGFBPs, IGFBP-5 and -6 share the highest degree of homology with this domain of IGFBP-3; therefore, we investigated whether these two IGFBPs inhibit IGFBP-4 degradation. Both IGFBP-5 and IGFBP-6 inhibit the degradation of 125I-IGFBP-4 by MC3T3-E1-conditioned media, and their inhibitory effects are variably reversed by IGFs. Synthetic peptides containing highly basic, C-terminal regions of IGFBP-5 and IGFBP-6 inhibit 125I-IGFBP-4 degradation, as does an homologous IGFBP-3 peptide, yet each peptide displays a different IC50, with the IGFBP-5 peptide being the most potent and the IGFBP-6 peptide being the least potent. In contrast, a homologous, yet neutral, IGFBP-4 peptide does not inhibit 125I-IGFBP-4 proteolysis, confirming the role of basic residues in the inhibitory process. The IGFBP-3, -5, and -6 peptides, each of which contains the heparin-binding consensus sequence XBBBXXBX, bind heparin, yet the IGFBP-3 and -5 peptides bind heparin with the highest affinities, whereas the IGFBP-6 peptide binds heparin with approximately 10-fold less affinity. Consistent with these regions being involved in proteinase inhibition, heparin completely reverses their inhibitory effects on 125I-IGFBP-4 proteolysis. Together, these data demonstrate that IGFBP-3, -5, and -6 can function as IGF-reversible inhibitors of IGFBP-4 proteolysis, likely through homologous, highly basic, heparin-binding domains contained within the conserved thyroglobulin type-1 motif present in the C-termini of these IGFBPs.
Fowlkes
J L
JL
Department of Pediatrics, University of Kentucky College of Medicine, Lexington 40536, USA.
Thrailkill
K M
KM
George-Nascimento
C
C
Rosenberg
C K
CK
Serra
D M
DM
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D017077:Culture Media, Conditioned; D006493:Heparin / Q000378:metabolism*; D006801:Humans; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000737:chemistry / Q000378:metabolism*; D018974:Insulin-Like Growth Factor Binding Protein 4 / Q000378:metabolism*; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000737:chemistry / Q000378:metabolism*; D018976:Insulin-Like Growth Factor Binding Protein 6 / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors*; D051379:Mice; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology*; D011266:Pregnancy-Associated Plasma Protein-A; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D017386:Sequence Homology, Amino Acid
D017077:Culture Media, Conditioned; D018972:Insulin-Like Growth Factor Binding Protein 3; D018974:Insulin-Like Growth Factor Binding Protein 4; D018975:Insulin-Like Growth Factor Binding Protein 5; D018976:Insulin-Like Growth Factor Binding Protein 6; D010446:Peptide Fragments; D011994:Recombinant Proteins; D006493:Heparin; D008666:Metalloendopeptidases; D011266:Pregnancy-Associated Plasma Protein-A