The rat growth hormone-releasing hormone receptor gene: Structure, regulation, and generation of receptor isoforms with different signaling properties

The interaction of GHRH with membrane-bound receptors on somatotroph cells of the anterior pituitary is an important step in the regulation of GK synt hesis and secretion. The identification of a G protein-coupled receptor for GHRH has made it possible to investigate the pathway by which GHRH regulat es pituitary somatotroph cell function. To initiate an analysis of the mech anisms regulating expression and function of the GHRH receptor, the structu re of the gene and its promoter region were analyzed. The coding sequence o f the rat GHRH receptor gene is contained within 14 exons spanning approxim ately 15 kb of genomic DNA. Four transcription start sites are located with in 286 bp upstream of the initiation codon. The 5' flanking region of the G HRH receptor gene acts as a functional promoter in rat pituitary tumor GH3 cells, and basal promoter activity is enhanced in GH3 and COS7 cells by cot ransfection of an expression construct encoding the pituitary-specific tran scription factor Pit-1. The rat GHRH receptor gene is subject to at least 1 alternative RNA processing event that generates 2 receptor isoforms differ ing by 41 amino acids within the third intracellular loop (IL) of the prote in. The short isoform of the GHRH receptor is predominant in pituitary cell s. The MtT/S pituitary tumor cell line was found to express the GHRH recept or, and different populations of these cells produce predominantly the long or short isoforms of the receptor messenger RNA, suggesting that the alter native splicing can be regulated. Functional analysis of the two GHRH recep tor isoforms demonstrates that both bind GHRH, but only the short isoform s ignals through a cAMP-mediated pathway. Neither receptor isoform is able to stimulate calcium mobilization from internal stores after GHRH treatment. Our findings indicate that the pituitary-specific transcription factor Pit- 1 is involved in the somatotroph-specific expression of the GHRH receptor g ene and that functionally distinct receptor proteins are generated by an al ternative RNA processing mechanism.