Identification of a Glucocorticoid Response Element in the Rat β₂-Adrenergic Receptor Gene

Abstract

Regulation of beta2-adrenergic receptor (beta2AR) levels by glucocorticoids is a physiologically important mechanism for altering beta2AR responsiveness. Glucocorticoids increase beta2AR density by increasing the rate of beta2AR gene transcription, but the cis-elements involved have not been well characterized. We now show that one of six potential glucocorticoid response elements (GREs) in the 5'-flanking region of the rat beta2AR gene is necessary for glucocorticoid-dependent stimulation of receptor gene expression. Using a nested set of deletion fragments of the rat beta2AR gene 5'-flanking region fused to a luciferase reporter gene, glucocorticoid-dependent induction of reporter gene expression in HepG2 cells was localized to a region between positions -643 and -152, relative to the transcription initiation site. In electrophoretic mobility shift assays, a double-stranded oligonucleotide incorporating a near-consensus GRE from this region (positions -379 to -365) formed complexes with the human recombinant glucocorticoid receptor, as well as with nuclear protein from dexamethasone-treated HepG2 cells. Mutation of a single base within this GRE sequence greatly diminished interaction of the mutated oligonucleotide with the human recombinant glucocorticoid receptor. The functional activity of the GRE was characterized using a luciferase reporter construct driven by a minimal thymidine kinase promoter. In HepG2 cells transfected with constructs containing the GRE, dexamethasone increased reporter gene expression approximately 3-fold, whereas a dexamethasone effect was not observed with constructs lacking the GRE. Taken together, these findings show that a GRE located at positions -379 to -365 in the 5'-flanking region of the rat beta2AR gene mediates glucocorticoid stimulation of beta2AR gene transcription.

Disruption of Rat Forebrain Development by Glucocorticoids: Critical Perinatal Periods for Effects on Neural Cell Acquisition and on Cell Signaling Cascades Mediating Noradrenergic and Cholinergic Neurotransmitter/Neurotrophic Responses

“…Dex influences testosterone synthesis and metabolism (Reznikov et al, 2004) and in turn, testosterone regulates the expression of a 2 ARs (Dygalo et al, 2002). Although parallel studies have not been conducted for sex-dependent effects on bAR expression, there are steroid-binding sites in the bAR promoter (Cornett et al, 1998), so that such differences may also exist for this receptor. In either case, the sexselective effects of Dex on adrenergic receptors and corresponding cell signaling during this critical period are likely to contribute to long-term effects.…”

Lasting Effects of Developmental Dexamethasone Treatment on Neural Cell Number and Size, Synaptic Activity, and Cell Signaling: Critical Periods of Vulnerability, Dose–Effect Relationships, Regional Targets, and Sex Selectivity

“…The focused effects on adrenergic receptors may reflect a common origin in long-term hormonal effects of developmental Dex treatment. Both the a 2 AR and the bAR are directly regulated by hormonal input (Cornett et al, 1998;Davies and Lefkowitz, 1984;Dygalo et al, 2002), and it is therefore likely that the shift in hormone synthesis and metabolism that follows Dex treatment (Reznikov et al, 2004) contributes to the changes in receptor expression, in turn providing yet another factor that underlies the sex-selectivity of behavioral outcomes (Bowman et al, 2004;Kreider et al, 2005b).…”

Moderate Global Reduction in Maternal Nutrition Has Differential Stage of Gestation Specific Effects on β1- and β2-Adrenergic Receptors in the Fetal Baboon Liver

“…36 It is suggested that a GC repressor domain in the rat b 1 -AR gene 37 may contribute to the transcriptional suppression of b 1 -ARs by GC. 36 Although b 1 -AR protein levels decline at 38 Although at present it is not known if catecholamine levels change at 0.9G compared to 0.5G in response to MNR, results of our study as well as those of others implicate a role for GC in regulating b-AR gene expression in fetuses of nutrient-restricted baboons.…”