IDENTIFICATION OF CYCLIC-AMP RESPONSE ELEMENT LN THE HUMAN RENIN GENE

In order to identify the mechanism by which cyclic AMP stimulates expr ession of the human renin gene (REN), the effect of forskolin was test ed in transient expression analyses of REN 5'-flanking DNA-chloramphen icol acetyltransferase (CAT) reporter gene constructs in secondary cul tures of human chorio-decidual cells, a major site of renin synthesis. Forskolin induced a mean 5-fold stimulation which was localized to DN A in the region -249 to -162 with respect to the transcription start s ite (+1). Such DNA also mediated a response to forskolin in heterologo us (HSV thymidine kinase) promoter constructs. Strong cAMP-response el ement (CRE) homology at -222 to -218 resembled the target for members of the CRE binding protein (CREB) family. Gel shift assays demonstrate d similarly migrating nucleoprotein complexes for oligonucleotides con taining the putative REN CRE as for a canonical CRE, in chorio-decidua l, JEG-3 and HeLa nuclear extracts. Mutation of residues critical for CREB attachment reduced binding. In conclusion, a CRE was identified a t -222 to -218 that appears critical for cAMP-induced human renin gene transcription. (C) 1994 Academic Press, Inc.