The regulation of glucose transporter gene expression by cyclic adenosine monophosphate in NIH3T3 fibroblasts.
The effect of cAMP on glucose transport was studied in fibroblastic cells. Incubation of confluent NIH3T3 cells for 6 h in the presence of cholera toxin (10 ng/ml) and 3-isobutyl-1-methylxanthine [(IBMX) 0.2 mM] or 8-bromo-cAMP (0.3 mM) and IBMX resulted in a 4-fold increase in the rate of deoxyglucose uptake; no change in hexose transport could be detected after treatment for 30 min. Either cholera toxin (0.3 ng/ml-30 ng/ml) or 8-bromo-cAMP (30 microM-3 mM) increased the expression of the mRNA encoding the glucose transporter (GT) protein, as determined by hybridization of size-fractionated total RNA to a rat brain GT cDNA. Activation of adenylate cyclase by forskolin also rapidly induced a 4- to 10-fold increase in GT mRNA. The rise in the level of GT mRNA was maximal 3-4 h after addition of the drug, and returned to basal values by 16 h. The stimulation was concentration dependent, with forskolin producing a maximal effect at 30 microM. The effect of a submaximal concentration (1 microM) of forskolin was greatly enhanced in the presence of IBMX (0.2 mM), which alone had little effect on GT mRNA levels. The forskolin-stimulated increase in GT mRNA was not blocked by inhibition of protein synthesis by cycloheximide (10 micrograms/ml) or anisomycin (100 microM). The involvement of GT gene transcription was assessed by the nuclear run-on assay. Treatment of the cells with 30 microM forskolin increased transcription 10-fold within 30 min; the activation was not blocked by cycloheximide.(ABSTRACT TRUNCATED AT 250 WORDS)
Hiraki
Y
Y
Department of Cellular and Molecular Physiology, Harvard Medical School, Boston, Massachusetts 02115.
McMorrow
I M
IM
Birnbaum
M J
MJ
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D000841:Anisomycin / Q000494:pharmacology; D001693:Biological Transport, Active; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D005347:Fibroblasts; D005786:Gene Expression Regulation; D005947:Glucose / Q000378:metabolism; D066298:In Vitro Techniques; D009004:Monosaccharide Transport Proteins / Q000235:genetics*; D011493:Protein Kinase C / Q000502:physiology; D012313:RNA / Q000032:analysis; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D009004:Monosaccharide Transport Proteins; D005576:Colforsin; D012313:RNA; D000841:Anisomycin; D002772:Cholera Toxin; D000242:Cyclic AMP; D011493:Protein Kinase C; D005947:Glucose; D013755:Tetradecanoylphorbol Acetate; D015056:1-Methyl-3-isobutylxanthine