Retinoic Acid-mediated Activation of the MouseRenin Enhancer

Abstract

Previous studies demonstrate that the mouserenin gene is regulated by a complex enhancer of transcription located 2.6 kilobases upstream of the transcription start site which is under both positive and negative influence. We demonstrate herein that a positive regulatory element (Eb) is repeated 10 bp upstream (Ec), and both are required for baseline activity of the enhancer. The Eb and Ec core sequences are identical to the consensus sequence for the nuclear hormone receptor superfamily of transcription factors, and transcriptional activity of constructs containing the enhancer is increased after treatment with retinoic acid. Maximal induction requires both Eb and Ec. Expression of endogenousrenin and a renin-promoter controlled transgene in As4.1 cells, and kidney renin mRNA in C57BL/6J mice was induced after retinoid treatment. Gel mobility supershift analysis revealed the binding of RARα and RXRα to oligonucleotides containing both Eb and Ec. Reverse transcriptase-polymerase chain reaction analysis revealed that As4.1 cells express both receptor isoforms, along with RARγ, but do not express RARβ, RXRβ, or RXRγ. Co-transfection of an expression vector encoding wild-type RARα increased enhancer activity, whereas a dominant negative mutant of RARα significantly attenuated retinoic acid-induced activity of the enhancer. These results demonstrate the importance of the Eb and Ec motifs in controlling baseline activity of the reninenhancer, and suggest the potential importance of retinoids in regulating renin expression.