Insulin binding to human astrocytoma cells and its effect on uridine incorporation into nucleic acid.
Binding of [125I]monoiodoinsulin to human astrocytoma cells (U-373 MG) was time dependent, reaching equilibrium after 1 h at 22 degrees C with equilibrium binding corresponding to 2.2 fmol/mg protein: this represents approximately 2,000 occupied binding sites per cell. The t1/2 of 125I-insulin dissociation at 22 degrees C was 10 min; the dissociation rate constant of 1.1 X 10(-2) s-1 was unaffected by a high concentration of unlabeled insulin (16.7 microM). Porcine insulin competed for specific 125I-insulin binding in a dose-dependent manner and Scatchard analysis suggested multiple affinity binding sites (higher affinity Ka = 4.4 X 10(8) M-1 and lower affinity Ka = 7.4 X 10(6) M-1). Glucagon and somatostatin did not compete for specific insulin binding. Incubation of cells with insulin (0.5 microM) for 2 h at 37 degrees C increased [2-14C]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 microM), 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.
Kum
W
W
Department of Medicine, Chinese University of Hong Kong, Shatin, NT.
Cockram
C S
CS
Zhu
S Q
SQ
Teoh
R
R
Young
J D
JD
D001254:Astrocytoma / Q000378:metabolism*; D002460:Cell Line; D004176:Dipyridamole / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005934:Glucagon / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D007700:Kinetics; D013004:Somatostatin / Q000378:metabolism; D014529:Uridine / Q000378:metabolism*
D007328:Insulin; D013004:Somatostatin; D004176:Dipyridamole; D005934:Glucagon; D014529:Uridine