Proenkephalin gene expression: interaction of glucocorticoid and cAMP regulatory elements

We used gel shift assays to determine the affinities of a 15 base glucocorticoid response element (GRE) derived from the rat proenkephalin (Penk) gene for the glucocorticoid receptor. A DNA binding domain of the glucocorticoid receptor interacted with a rat GRE (RGRE) with an apparent affinity intermediate between that of a consensus positive GRE oligo (GRE+) and a mismatch GRE oligo (GRE-). When inserted in front of a chloramphenicol acyl transferase (CAT) construct that is driven by a Penk promoter containing a cAMP response element, dexamethasone (10 microM) produced a 5-fold increase with GRE+, a small increase with RGRE, and no change with GRE- or the Penk promoter alone. Forskolin (20 microM) stimulated CAT activity 4- to 9-fold with each construct. However, dexamethasone plus forskolin caused a synergistic induction of CAT expression with the GRE+ oligo, no effect with the RGRE and an antagonistic effect with the Penk promoter alone and the GRE- oligo. These results demonstrate that GRE+, and to a lesser degree RGRE, can mimic the response of the endogenous Penk gene to dexamethasone and forskolin. Furthermore, a dexamethasone activated glucocorticoid receptor may inhibit cAMP mediated transcription of the Penk gene when a GRE+ or RGRE is not present.