Interplay of pituitary adenylate cyclase-activating polypeptide with a silencer element to regulate the upstream promoter of the human gonadotropin-releasing hormone receptor gene

Multiple transcription start sites were identified in the human gonadotropi n releasing hormone receptor (hGnRHR) gene. Recently, an upstream promoter residing at - 1727/ - 1674, in vicinity of a CAP site at - 1673, was charac terized. In this report, we elucidated the underlying mechanisms for the re gulation of this promoter. Functionally, this promoter was constitutively s uppressed by a silencer element ( - 1673/- 1351) situated immediately downs tream to it. On the other hand, pituitary adenylate cyclase-activating poly peptide (PACAP), via the cAMP pathway, was found to be the extracellular cu e to control the upstream promoter. Following PACAP-27, PACAP-38 (30 nM) an d forskolin (25 muM) treatment, there were significant increases in the rep orter gent: activities. By deletion analysis, the region residing at - 1727 to - 1577, containing the distal promoter and 97 bp of the silencer was su bsequently round to be responsible for PACAP/cAMP induction. To localize th e PACAP-dependent cis-acting element(s) within the silencer, block replacem ent scanning mutation was performed and a hGnRHR gene PACAP-responsive clem ent (GPRE) was identified at - 1676/ - 1648. The actions of PACAPs and fors kolin on the GPRE were further evidenced by gel mobility shift assays. Ther e was an increase in protein binding onto this element only after peptide t reatment. As GnRH receptor number on gonadotrope cell surface is a key fact or in regulating gonadotropin release, the present study provides an insigh t into the interplay between PACAP and GnRH receptors on pituitary gonadotr opes to control human reproductive functions. (C) 2001 Elsevier Science Ire land Ltd. All rights reserved.