Long-term effects of insulin on the enzyme activity and messenger RNA of glycogen synthase in rat hepatoma H4 cells: an effect of insulin on glycogen synthase mRNA stability.
Insulin induced glycogen synthase activity and decreased glycogen synthase mRNA concentrations in rat hepatoma H4 cells. Total enzyme activity measured with glucose 6-phosphate gradually increased during a 24-h insulin incubation. The time course of glycogen synthase activation measured by the activity ratio (low G-6-P/high G-6-P) in response to insulin was biphasic with the first peak at 15 min and the second peak at 4 to 6 h. When cells were incubated with insulin and cycloheximide, the first peak persisted while the second peak was abolished. These data suggest that the first activation peak derives from the classic effect of insulin via dephosphorylation and the second peak from an insulin-induced protein synthesis of a glycogen synthase activator. Ribonuclease protection assays with a cloned rat liver glycogen synthase cDNA were used to quantitate glycogen synthase mRNA. Insulin unexpectedly decreased glycogen synthase mRNA in a time- and a dose-dependent manner. After incubation with the RNA synthesis inhibitor, 5, 6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB) without and with insulin, the half time of glycogen synthase mRNA decreased from 6.0 +/- 0.80 to 3.9 +/- 0.75 h, respectively. Nuclear run-off experiments with isolated nuclei showed no change of transcription of glycogen synthase mRNA. These data suggest that insulin in this system affects glycogen synthase mRNA stability rather than transcription.
Okubo
M
M
Department of Pharmacology, University of Virginia Health Sciences Center, Charlottesville 22908.
Villar-Palasi
C
C
Nagasaka
Y
Y
Larner
J
J
Larner
A C
AC
Bai
G
G
Lee
E Y
EY
D000818:Animals; D006006:Glycogen Synthase / Q000378:metabolism*; D006207:Half-Life; D007328:Insulin / Q000494:pharmacology*; D008114:Liver Neoplasms, Experimental / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism*; D012334:RNA, Neoplasm / Q000378:metabolism; D051381:Rats; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D007328:Insulin; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D006006:Glycogen Synthase