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Insulin Binding to Human Astrocytoma Cells and Its Effect on Uridine Incorporation into Nucleic Acid
Author(s) -
Kum W.,
Cockram C. S.,
Zhu S. Q.,
Teoh R.,
Young J. D.
Publication year - 1989
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1989.tb10923.x
Subject(s) - insulin , nucleic acid , uridine , nucleoside , insulin receptor , dissociation constant , biochemistry , binding site , biology , chemistry , receptor , medicine , endocrinology , rna , microbiology and biotechnology , insulin resistance , gene
Binding of [ 125 I]monoiodoinsulin to human astrocytoma cells (U‐373 MG) was time dependent, reaching equilibrium after 1 h at 22°C with equilibrium binding corresponding to 2.2 fmol/mg protein: this represents approximately 2,000 occupied binding sites per cell. The t 1/2 of 125 I‐insulin dissociation at 22°C was 10 min; the dissociation rate constant of 1.1 × 10 −2 s −1 was unaffected by a high concentration of unlabeled insulin (16.7 μ M ). Porcine insulin competed for specific 125 I‐insulin binding in a dose‐dependent manner and Scatchard analysis suggested multiple affinity binding sites (higher affinity K a = 4.4 × 10 8 M −1 and lower affinity K a = 7.4 × 10 6 M −1 ). Glucagon and somatostatin did not compete for specific insulin binding. Incubation of cells with insulin (0.5 μ M ) for 2 h at 37°C increased [2‐ 14 C]uridine incorporation into nucleic acid by 62 ± 2% (n = 3) above basal. Cyclic AMP, in the absence of insulin, also stimulated nucleoside incorporation into nucleic acid [65 ± 1% (n = 3)] above basal. Preincubation with cyclic AMP followed by insulin had an additive effect on nucleoside incorporation [160 ± 4% (n = 3) above basal]. Dipyridamole (50 μ M ), a nucleoside transport inhibitor, blocked both basal and stimulated uridine incorporation. These studies confirm that human astrocytoma cells possess specific insulin receptors with a demonstrable effect of ligand binding on uridine incorporation into nucleic acid.