Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) as a Growth Hormone (GH)-releasing factor in grass carp: II. Solution structure of a brain-specific PACAP by nuclear magnetic resonance spectroscopy and functional studies on GH release and gene expression

Abstract

Pituitary adenylate cyclase-activating polypeptide (PACAP) has been proposed to be the ancestral GHRH. Recently, using grass carp as a model for modern-day bony fish, we demonstrated that PACAP nerve fibers are present in close proximity to carp somatotrophs, and mammalian PACAPs can induce GH secretion in carp pituitary cells. To further examine the role of PACAP as a GH-releasing factor in fish, the structural identity of grass carp PACAP was established by molecular cloning. The newly cloned PACAP was found to be a single-copy gene and expressed in the brain but not other tissues. The mature peptides of PACAP, namely PACAP 27 and PACAP 38, were synthesized. As revealed by nuclear magnetic resonance spectroscopies, carp PACAP 38 is composed of a flexible N terminal from His 1 to Ile 5, an extended central helix from Phe 6 to Val 26, and a short helical tail in the C terminal from Arg 29 to Arg 34. The C-terminal helix is located after a hinge region at Leu 27 to Gly 28 and is absent in the solution structures of PACAP 27. The two forms of PACAPs were effective in elevating GH release and GH transcript expression in grass carp pituitary cells. These stimulatory effects occurred with parallel rises in cAMP and Ca 2+ entry via voltage-sensitive Ca 2+ channels in carp somatotrophs. The present study represents the first report for solution structures of nonmammalian PACAPs and provides evidence that a brain-specific isoform of PACAP in fish can stimulate GH synthesis and release at the pituitary level, presumably by activating the appropriate postreceptor signaling mechanisms. Copyright © 2007 by The Endocrine Society.