Pituitary adenylate cyclase-activating polypeptide stimulates secretoneurin release and secretogranin II gene transcription in bovine adrenochromaffin cells through multiple signaling pathways and increased binding of pre-existing activator protein-1-like transcription factors.

Secretoneurin (SN) is a novel bioactive peptide that derives from the neuroendocrine protein secretogranin II (SgII) by proteolytic processing and participates in neuro-immune communication. The neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP-38) dose-dependently stimulates (EC(50) approximately 3 nM) SN release (up to 4-fold) and SgII gene expression (up to 60-fold) in cultured bovine adrenochromaffin cells. The effect of PACAP on both SN secretion and SgII mRNA levels is rapid and long lasting. We analyzed in this neuroendocrine cell model the transduction pathways involved in both SN secretion and SgII gene transcription in response to PACAP. The cytosolic calcium chelator BAPTA-AM and the nonselective calcium channel antagonist NiCl(2) equally inhibited both secretion of the peptide and transcription of the SgII gene, indicating a major contribution of calcium influx in PACAP-induced SN biosynthesis and release in chromaffin cells. Inhibition of protein kinase A (PKA) or C (PKC) also reduced PACAP-evoked SN release but did not alter the stimulatory effect of PACAP on SgII mRNA levels. Conversely, application of mitogen-activated protein kinase inhibitors suppressed PACAP-induced SgII gene expression. The effect of PACAP on SgII mRNA levels, like the effect of the PKC stimulator 12-O-tetradecanoylphorbol-13-acetate (TPA), was not affected by cycloheximide, whereas the effects of the PKA stimulator forskolin or cell-depolarization by high K(+) were significantly reduced by the protein synthesis inhibitor. PACAP and TPA both increased the binding activity of the SgII cAMP response element to trans-acting factors present in chromaffin cell nuclear extracts, which are recognized by antibodies to activator protein-1-related proteins. These data indicate that SN biosynthesis is regulated by PACAP in chromaffin cells through complex signaling cascades, suggesting that SN may play a function during trans-synaptic stimulation of the adrenal medulla.

Frog chromogranin A messenger ribonucleic acid encodes three highly conserved peptides. Coordinate regulation of proopiomelanocortin and chromogranin A gene expression in the pars intermedia of the pituitary during background color adaptation.

Chromogranin A (CgA) is a neuroendocrine secretory protein that is widely used as a marker for endocrine neoplasms but whose function is not completely understood. In mammals, it is thought that CgA is a precursor for biologically active peptides. Here, we describe the cloning of a complementary DNA encoding CgA from a nonmammalian vertebrate, the frog Rana ridibunda. Sequence analysis revealed that frog CgA exhibits only 40-44% amino acid sequence similarity with its mammalian homologues. The amino acid identity is confined to three regions (70-80% identity) of the protein that are flanked by conserved pairs of basic amino acid residues, suggesting that proteolytic processing at these cleavage sites may give rise to three biologically active peptides whose sequences have been highly preserved during evolution. Tissue distribution analysis by Northern blot and in situ hybridization revealed the widespread expression of frog CgA messenger RNA in the brain and in endocrine tissues, the highest concentration occurring in the distal lobe of the pituitary. Adaptation of frog skin color to a dark background caused a concomitant increase in CgA and POMC messenger RNA levels in the intermediate lobe of the pituitary. Taken together, these data indicate that CgA may function as a precursor to three highly conserved peptides that may exert regulatory functions in the neuroendocrine system.