Molecular cloning of ovine and bovine growth hormone-releasing hormone receptors: The ovine receptor is C-terminally truncated

To provide information about species differences in GH-releasing hormone (G HRH) receptors useful for studies of receptor-ligand binding properties and receptor function, we have cloned the ovine and bovine pituitary GHRH rece ptors (GHRHRs). The ovine receptor (oGHRHR) was cloned from a pituitary com plementary DNA library and encodes a protein that is similar to that of por cine, human, rat, and mouse with, respectively, 84.3, 80.7, 75.9, and 74.0% amino acid identity. Surprisingly, oGHRHR has a 16 amino acid truncation a t its carboxyl-terminal end when compared with GHRHRs from other known mamm als. RT-PCR using pooled pituitary RNA from a different population of sheep could detect only truncated receptor. Bovine GHRHR (bGHRHR) was cloned by RT-PCR and shows 92.5% amino acid sequence identity with oGHRHR, but has no truncation. Genomic sequencing of the appropriate region of goat receptor intron 13 showed that the caprine receptor shares the same truncation seen in sheep. Photoaffinity cross-linking of GHRH to ovine and bovine pituitary membranes confirms that the native ovine pituitary GHRHR protein is smalle r by the amount predicted by the cloned sequences. The truncation did not a ffect GHRH binding as oGHRHR, bGHRHR, human GHRHR, and human GHRHR, which w as truncated by site-directed mutagenesis to match the oGHRHR, all showed c omparable GHRH binding affinity when expressed in transfected cell lines. I n contrast, the ovine and truncated human receptors demonstrated enhanced s ensitivity for GHRH stimulation of cAMP (lowered ED,,) relative to hGHRHR a nd bGHRHR. This suggests that this C-terminal domain acts to inhibit cAMP s ignaling possibly through a role in receptor down regulation.