Characterization of Insulin-Like Growth Factor-I Receptors in PC 12 Pheochromocytoma Cells and Bovine Adrenal Medulla

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Abstract
Competitive binding studies indicated that PC 12 cells have receptors for insulin-like growth factor-I (IGF-I). There are ∼ 11,100 ± 1,500 IGF-I receptors/cell; these receptors have an apparent KD for IGF-I of 7.2 ± 0.6 nM. Covalent cross-linking of l25I-IGF-I to PC 12 cells labeled a 125,000-130,000-Mr protein, presumably the α-subunit of the IGF-I receptor. Although PC 12 cells also have insulin receptors, the 125I-IGF-I appeared to be cross-linked to IGF-I receptors, because 100 nM IGF-I competed for labeling but 100 nM insulin did not. Bovine chromaffin cells also have IGF-I receptors. The protein tyrosyl kinase activity of IGF-I receptors from bovine adrenal medulla and PC 12 cells was examined after purification of the receptors by wheat germ agglutinin-Sepharose chromatography. IGF-I (10 nM) stimulated autophosphorylation of the β-subunits of the IGF-I receptors from both preparations; the β-subunits from both sources had Mr values of ∼97,000. IGF-I also stimulated phosphorylation of the synthetic substrate poly(Glu:Tyr)4:1 by both receptor preparations. IGF-I (IC50 of ∼0.2 nM) was much more potent than insulin at stimulating phosphorylation of poly(Glu:Tyr) by the bovine adrenal medulla preparation. A maximal concentration of IGF-I (10 nM) increased phosphorylation approximately threefold. IGF-I was slightly more effective than insulin at stimulating the phosphorylation of poly(Glu:Tyr) by the PC 12 cell receptor preparation, but neither ligand produced a maximal effect at concentrations up to 100 nM. This result probably reflects the presence of comparable numbers of IGF-I and insulin receptors on PCI 2 cells. Our data indicate that the IGF-I receptors on PC 12 cells are similar to IGF-I receptors in the adrenal medulla and other tissues. Presumably these receptors are responsible for the stimulation of PC 12 cell replication by IGF-I.