α(2D/A)-Adrenergic receptor gene induction in the retina by phorbol ester: Involvement of an AP-2 element

Abstract

Background: The α2-adrenergic receptor (α2-AR) expressed in the bovine retina has been demonstrated to be of the α(2D) subtype. The bovine α(2D)-adrenergic receptor (α(2D/A)-AR) gene has been cloned and characterized. This report describes the induction of this gene by phorbol- 12, 13-myristate acetate (PMA), an activator of protein kinase C (PKC). Results: Treatment of the bovine retina for 60 min with PMA (1 μM) resulted in significant and similar increases in α(2D/A)-AR mRNA level and gene transcription. This indicates that PMA causes α(2D/A)-AR gene induction and that this induction takes place directly at the transcriptional level. In C6 cells, treatment with PMA at a concentration which was as low as 0.1 μM induced endogenous α(2D/A)-AR mRNA after 60 min. Luciferase reporter assays in C6 cells mapped the PMA-responsive element to a region between -247 bp and -163 bp on the α(2D/A)-AR promoter. Electrophoretic mobility shift assays showed an increased binding of nuclear factor(s) from PMA-treated bovine retina to this promoter region. Competition assays indicate that an AP-2 element may be involved in the PMA-dependent induction. Conclusion: These findings demonstrate for the first time, the direct induction of the α(2D/A)-AR gene by PMA and support a role for an AP-2 element in the induction mechanism.