Characterization of the receptors for chicken GHRH and GHRH-related peptides: Identification of a novel receptor for GHRH and the receptor for GHRH-LP (PRP)

Abstract 

Growth hormone–releasing hormone and its structurally related peptides, GHRH-like peptide (GHRH-LP) (also called PRP), peptide histidine-isoleucine (PHI), vasoactive intestinal polypeptide (VIP), and pituitary adenylate cyclase-activating polypeptide (PACAP), have been reported to play important physiological roles in pituitary and extrapituitary tissues of vertebrates; however, little is known about the identity of these GHRH-related peptide receptors in birds. In this study, 6 receptors for GHRH and GHRH-related peptides (cGHRHR₁, cGHRHR₂, cGHRH-LPR, cPAC₁, cVPAC₁, and cVPAC₂) were cloned from chicken brain or pituitary, and their functionalities were examined in Chinese hamster ovary (CHO) cells using a pGL3-CRE-luciferase reporter system. Results showed that: (1) all receptors are G protein–coupled receptors functionally coupled to the intracellular PKA signaling pathway; (2) 2 GHRH receptors (cGHRHR₁ and cGHRHR₂) were identified, and both receptors could be potently activated by cGHRH; (3) cGHRH-LP could activate its specific receptor cGHRH-LPR (cPRP-R), and it also activated cGHRHR₁ and cGHRHR₂; and (4) PACAP could potently activate its receptors cPAC₁, cVPAC₁ and cVPAC₂; however, cVPAC₁ and cVPAC₂ could also be effectively activated by cVIP and tPHI, indicating that they can serve as VIP receptors and potential PHI receptors. Using a reverse transcription polymerase chain reaction assay, we further examined the mRNA expression of these receptors in adult chicken tissues. The expressions of cGHRHR₁, cGHRHR₂, and cGHRH-LPR are restricted mainly to the pituitary and/or brain, whereas cPAC₁, cVPAC₁, and cVPAC₂ are expressed in most of the tissues examined. Collectively, our study identified the receptors for chicken GHRH and GHRH-related peptides, including a novel GHRH receptor (cGHRHR₂), and established a basis to elucidate the roles of these peptides in target tissues.

Abbreviations: c, chicken, CHO, Chinese hamster ovary, CRE, cAMP response element, EST, expression sequence tag, GH, growth hormone, GHRH, growth hormone–releasing hormone, GHRH-LP, GHRH-like peptide, GHRHR₁, GHRH type I receptor, GHRHR₂, GHRH type II receptor, o, ovine, PAC₁, PACAP type I receptor, PACAP, pituitary adenylate cyclase-activating polypeptide, PCR, polymerase chain reaction, PHI/PHM, peptide histidine-isoleucine/methionine, PHI-R, PHI receptor, PHV, peptide histidine-valine, PKA, protein kinase A, PRL, prolactin, PRP, PACAP-related peptide, PRP-R, PRP receptor, RAMPs, receptor activity–modifying proteins, RT, reverse transcription, t, turkey, VIP, vasoactive intestinal polypeptide, VPAC₁, PACAP type II/VIP type I receptor, VPAC₂, PACAP type III/VIP type II receptor

Keywords: GHRH, GHRH-related peptide, GHRHR, GHRH-LPR, PHI-R