Identification of a Negative Glucocorticoid Response Element in the Rat Type 1 Vasoactive Intestinal Polypeptide Receptor Gene

Abstract

Glucocorticoids play important roles in lung development and function by modulating the expression of a variety of genes. The type 1 vasoactive intestinal polypeptide (VIP) receptor gene is highly expressed in the lung where it mediates VIP physiological functions. In this study, the effect of glucocorticoid on VIP receptor gene expression was examined. Dexamethasone (100 n) suppresses endogenous VIP receptor mRNA expression in cultured lung cells. Transient transfection of lung cells with fusion constructs containing various portions of the VIP receptor 5'-flanking sequences linked to the luciferase reporter gene shows that 126 base pairs (bp) of the VIP receptor upstream sequences are sufficient to mediate transcriptional repression by glucocorticoid. DNase I footprinting demonstrates that purified glucocorticoid receptor (GR) binds to the VIP receptor promoter between -21 and -36 bp relative to the transcription start site. Point mutations within this binding site not only abolish GR binding and GR-mediated transcriptional repression of the VIP receptor gene, but basal transcription is also reduced to background levels. Co-transfection of GR expression vector and the VIP receptor GR binding site linked to the thymidine kinase promoter and luciferase shows that this sequence is sufficient to confer glucocorticoid-mediated transcriptional repression to a heterologous promoter. These results indicate that the VIP receptor gene contains a negative glucocorticoid response element between -21 and -36 bp that may act to regulate both basal and glucocorticoid-mediated expressions of the VIP receptor gene in lung cells.