Insulin-like growth factor receptors and binding protein in rat neuroblastoma cells

M. A. Sturm, Cheryl A Conover, H. Pham, R. G. Rosenfeld

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

B104, an established rat neuroblastoma cell line exhibiting specific neuronal qualities, was chosen as a model to study insulin-like growth factor (IGF) binding and action in the central nervous system. Specific binding of [125I]IGF-II to B104 membranes averaged 12.2 ± 4.0% (mean ± SD)/100 μg/ml protein compared with [125I]IGF-I binding of 10.1 ± 2.9%. In competitive binding studies employing [125I]IGF-II as the radioligand, high affinity for IGF-II was demonstrated (50% displacement at 2.7 ng/ml), with none for IGF-I or insulin. Upon cross-linking [125I]IGF-I to membranes under reducing conditions, two prominent bands were observed, migrating with apparent mol wt (M(r)) of 135,000 and 280,000. Both bands were inhibited by IGFs and insulin, but not by R-II-PABI, a polyclonal antibody to the type 2 receptor. These bands presumably represent the α-subunit and an incompletely reduced α-α-dimer of the type 1 IGF receptor. When cross-linking [125I]IGF-II to membranes under reducing conditions, the primary labeled bands migrated with apparent M(r) of 260,000 and 280,000. These bands were inhibited by IGF-II anbd R-II-PABI, but not by insulin, and probably represent the monomeric type 2 receptor. In addition, we observed a minor band at apparent M(r) 35,000, which was inhibited by IGF but not by insulin. By a modified cross-linking technique, we confirmed the existence of a small IGF-binding protein in the serum-free conditioned medium of B104 cultures, migrating as two bands with apparent M(r) of 33,000-39,000. These proteins demonstrated high affinity for IGF-I and IGF-II, but none for insulin. In summary, this study demonstrates the presence in B104 rat neuroblastoma cells of 1) abundant classical type 1 and type 2 IGF receptors, and 2) a secreted and membrane-associated small IGF-binding protein.

Original languageEnglish (US)
Pages (from-to)388-396
Number of pages9
JournalEndocrinology
Volume124
Issue number1
StatePublished - 1989
Externally publishedYes

Fingerprint

Somatomedin Receptors
Insulin-Like Growth Factor Binding Proteins
Insulin-Like Growth Factor II
Neuroblastoma
Insulin-Like Growth Factor I
Insulin
IGF Type 1 Receptor
Membranes
Somatomedins
IGF Type 2 Receptor
Competitive Binding
Serum-Free Culture Media
Conditioned Culture Medium
Proteins
Central Nervous System
Cell Line
Antibodies

ASJC Scopus subject areas

  • Endocrinology
  • Endocrinology, Diabetes and Metabolism

Cite this

Insulin-like growth factor receptors and binding protein in rat neuroblastoma cells. / Sturm, M. A.; Conover, Cheryl A; Pham, H.; Rosenfeld, R. G.

In: Endocrinology, Vol. 124, No. 1, 1989, p. 388-396.

Research output: Contribution to journalArticle

Sturm, MA, Conover, CA, Pham, H & Rosenfeld, RG 1989, 'Insulin-like growth factor receptors and binding protein in rat neuroblastoma cells', Endocrinology, vol. 124, no. 1, pp. 388-396.
Sturm, M. A. ; Conover, Cheryl A ; Pham, H. ; Rosenfeld, R. G. / Insulin-like growth factor receptors and binding protein in rat neuroblastoma cells. In: Endocrinology. 1989 ; Vol. 124, No. 1. pp. 388-396.
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N2 - B104, an established rat neuroblastoma cell line exhibiting specific neuronal qualities, was chosen as a model to study insulin-like growth factor (IGF) binding and action in the central nervous system. Specific binding of [125I]IGF-II to B104 membranes averaged 12.2 ± 4.0% (mean ± SD)/100 μg/ml protein compared with [125I]IGF-I binding of 10.1 ± 2.9%. In competitive binding studies employing [125I]IGF-II as the radioligand, high affinity for IGF-II was demonstrated (50% displacement at 2.7 ng/ml), with none for IGF-I or insulin. Upon cross-linking [125I]IGF-I to membranes under reducing conditions, two prominent bands were observed, migrating with apparent mol wt (M(r)) of 135,000 and 280,000. Both bands were inhibited by IGFs and insulin, but not by R-II-PABI, a polyclonal antibody to the type 2 receptor. These bands presumably represent the α-subunit and an incompletely reduced α-α-dimer of the type 1 IGF receptor. When cross-linking [125I]IGF-II to membranes under reducing conditions, the primary labeled bands migrated with apparent M(r) of 260,000 and 280,000. These bands were inhibited by IGF-II anbd R-II-PABI, but not by insulin, and probably represent the monomeric type 2 receptor. In addition, we observed a minor band at apparent M(r) 35,000, which was inhibited by IGF but not by insulin. By a modified cross-linking technique, we confirmed the existence of a small IGF-binding protein in the serum-free conditioned medium of B104 cultures, migrating as two bands with apparent M(r) of 33,000-39,000. These proteins demonstrated high affinity for IGF-I and IGF-II, but none for insulin. In summary, this study demonstrates the presence in B104 rat neuroblastoma cells of 1) abundant classical type 1 and type 2 IGF receptors, and 2) a secreted and membrane-associated small IGF-binding protein.

AB - B104, an established rat neuroblastoma cell line exhibiting specific neuronal qualities, was chosen as a model to study insulin-like growth factor (IGF) binding and action in the central nervous system. Specific binding of [125I]IGF-II to B104 membranes averaged 12.2 ± 4.0% (mean ± SD)/100 μg/ml protein compared with [125I]IGF-I binding of 10.1 ± 2.9%. In competitive binding studies employing [125I]IGF-II as the radioligand, high affinity for IGF-II was demonstrated (50% displacement at 2.7 ng/ml), with none for IGF-I or insulin. Upon cross-linking [125I]IGF-I to membranes under reducing conditions, two prominent bands were observed, migrating with apparent mol wt (M(r)) of 135,000 and 280,000. Both bands were inhibited by IGFs and insulin, but not by R-II-PABI, a polyclonal antibody to the type 2 receptor. These bands presumably represent the α-subunit and an incompletely reduced α-α-dimer of the type 1 IGF receptor. When cross-linking [125I]IGF-II to membranes under reducing conditions, the primary labeled bands migrated with apparent M(r) of 260,000 and 280,000. These bands were inhibited by IGF-II anbd R-II-PABI, but not by insulin, and probably represent the monomeric type 2 receptor. In addition, we observed a minor band at apparent M(r) 35,000, which was inhibited by IGF but not by insulin. By a modified cross-linking technique, we confirmed the existence of a small IGF-binding protein in the serum-free conditioned medium of B104 cultures, migrating as two bands with apparent M(r) of 33,000-39,000. These proteins demonstrated high affinity for IGF-I and IGF-II, but none for insulin. In summary, this study demonstrates the presence in B104 rat neuroblastoma cells of 1) abundant classical type 1 and type 2 IGF receptors, and 2) a secreted and membrane-associated small IGF-binding protein.

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