A mitogenic peptide amide encoded within the E peptide domain of the insulin-like growth factor IB prohormone.
We have identified an amino acid sequence within the E peptide of the insulin-like growth factor IB (IGF-IB) precursor that is biologically active and designated this peptide insulin-like growth factor IB-(103-124) E1 amide (IBE1). Its existence was predicted by a flanking Gly-Lys-Lys-Lys, a signal sequence for sequential proteolytic cleavage and peptidyl C-terminal amidation. A synthetic analog of the predicted IBE1 peptide, designated Y-23-R-NH2, was generated with tyrosine added at position 0. This peptide at 2-20 nM had growth-promoting effects on both normal and malignant human bronchial epithelial cells. Y-23-R-NH2 bound to specific high-affinity receptors (Kd = 2.8 +/- 1.4 x 10(-11) M) present at 1-2 x 10(4) binding sites per cell. Ligand binding was not inhibited by recombinant insulin or recombinant IGF-I. Furthermore, a monoclonal antibody antagonist to the IGF-I receptor (alpha IR3) did not suppress the proliferative response induced by Y-23-R-NH2. In addition, C-terminal amidation was shown to be important in receptor recognition since the free-acid analog of IBE1 (Y-23-R-OH) did not effectively compete for binding and was not a potent agonist of proliferation. Immunoblot analysis of human lung tumor cell line extracts using an antibody raised against Y-23-R-NH2 detected a low molecular mass band of approximately 5 kDa, implying that a protein product is produced that has immunological similarity to IBE1. Extracts of human, mammalian, and avian livers analyzed on an immunoblot with the anti-Y-23-R-NH2 antibody contained proteins of approximately 21 kDa that were specifically recognized by the antiserum and presumably represent an IGF-I precursor molecule. This implies that in species where an IGF-I mRNA with homology to the human IGF-IB E domain has not yet been described, an alternate mRNA must be produced that contains a sequence similar to that of the midportion of the human IGF-IB E domain. Our findings demonstrate that IBE1 is a growth factor that mediates its effect through a specific high-affinity receptor and is most likely conserved in many species.
Siegfried
J M
JM
Department of Pharmacology, University of Pittsburgh, PA 15261.
Kasprzyk
P G
PG
Treston
A M
AM
Mulshine
J L
JL
Quinn
K A
KA
Cuttitta
F
F
D000577:Amides; D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D006801:Humans; D066298:In Vitro Techniques; D007334:Insulin-Like Growth Factor I / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008934:Mitogens / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011956:Receptors, Cell Surface / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000577:Amides; D008934:Mitogens; D010446:Peptide Fragments; D011498:Protein Precursors; D011956:Receptors, Cell Surface; D007334:Insulin-Like Growth Factor I