TRANSCRIPTIONAL AND POSTTRANSCRIPTIONAL MECHANISMS OF GLUCOCORTICOID-MEDIATED REPRESSION OF PHOSPHOENOLPYRUVATE CARBOXYKINASE GENE-EXPRESSION IN ADIPOCYTES

Glucocorticoids exert pleiotropic effects, among which negative regula tion of transcription has been recognized as of crucial importance. Wh ile glucocorticoids induce phosphoenolpyruvate carboxykinase (PEPCK) g ene expression in liver cells, it represses gene activity in adipose c ells. We used the 3T3-F442A adipocytes to analyze the underlying mecha nisms. in these cells, the synthetic glucocorticoid dexamethasone exer ts a dominant repression either on basal or on beta-agonist stimulatio n of PEPCK gene expression. To determine whether glucocorticoid action required protein synthesis, we employed cycloheximide, anisomycin, an d puramycin, three different translation inhibitors. None of these aff ected induction by isoprenaline or repression by dexamethasone of isop renaline stimulation. In contrast, dexamethasone inhibitory action on basal PEPCK mRNA was totally prevented by the three translation inhibi tors. Time courses of glucocorticoid action on basal and on induction by beta-agonist were similar. Half-maximal effect of dexamethasone on isoprenaline-induced PEPCK mRNA was obtained at about 10 nM, a tenfold higher concentration than that observed for the reduction of basal mR NA. Using the transcription inhibitor DRB, we showed that dexamethason e did not alter mRNA half-life, while isoprenaline strongly stabilized mRNA. in a 3T3-F442A stable transfectant bearing -2,100 base pairs of the PEPCK promoter fused to the chloramphenicol acetyltransferase (CA T) gene, isoprenaline stimulated CAT activity, whereas dexamethasone r educed basal and isoprenaline-induced CAT expression. Hence, beta-agon ists exert both transcriptional and posttranscriptional regulation, wh ile glucocorticoid action is purely transcriptional. However, mechanis ms of glucocorticoid repression of basal and of beta-agonist stimulati on appear different. (C) 1997 Wiley-Liss, Inc.