Molecular and functional analyses of growth hormone-releasing hormone (GHRH) from olive flounder (Paralichthys olivaceus)

Abstract

Growth hormone-releasing hormone (GHRH) is an important neuroendocrine factor that stimulates the release of growth hormone (GH) from the anterior pituitary. Several nonmammalian GHRH-like peptides were reported previously to be encoded by PACAP and processed from the same transcript and prepropolypeptide. However, the true nonmammalian GHRHs in amphibian and fishes were only recently discovered. We identified and characterized the primary structure of the GHRH gene and determined its expression profiles under normal and infectious conditions in the teleost fish, Paralichthys olivaceus. The 142 amino acids of the GHRH precursor are encoded by six exons spanning 2290bp. The flounder GHRH precursor mRNA was constitutively expressed in the brain as well as gills and ovary. Inducible expression of GHRH mRNA was observed in the gills of Edwardsiella tarda-challenged fish. Induction of GHRH mRNA was highest at 24h post-bacterial challenge. Subsequently, the biological role of GHRH was investigated by exogenous treatment of flounder embryogenic cells (hirame natural embryonic cells, HINAE cells) and primary cultured pituitary cells with a synthetic GHRH peptide (fGHRH-28). The 10(-6)M concentration of fGHRH-28 produced intracellular cAMP in HINAE cells and induced growth hormone mRNA in both of HINAE and pituitary cells. The profiles of TNF-α mRNA expression differed from HINAE and pituitary cells after fGHRH-28 treatment. TNF-α mRNA levels elevated approximately 3-fold in HINAE cells, but decreased to one-third in pituitary cells stimulated by fGHRH-28. These results suggest that the flounder GHRH plays roles in the bidirectional communication network between growth and immunity in fish.