Catecholamines and peptides are co-stored and co-secreted physiologically by exocytosis from sympathochromaffin cells in response to a physiologic nicotinic cholinergic stimulus. During and after secretion, how do such cells program the re-synthesis of their secreted components? That is, what is the mechanism of stimulus-transcription (stimulus-secretion-synthesis) coupling in such cells? Does the same physiologic signal which triggers secretion (a nicotinic agonist) also trigger re-synthesis of secreted peptides and catecholamines? If so, is the signal transduction pathway for nicotinic effects on re-synthesis convergent with (similar to) or divergent from that for its effects on secretion? Chromogranin A (CgA) is the major soluble protein in the core of catecholamine storage vesicles. Its adrenergic functions include binding of calcium and catecholamines within the vesicle core, and generation of peptides such as the vasodilator vasostatin, and an autocrine chromostatin-like activity which feedback-inhibits catecholamine release. The mRNAs for chromogranins A and B, as well as tyrosine hydroxylase, are augmented by nicotinic stimulation. Evidence so-far gathered indicates that the CgA gene is transcriptionally activated by a nicotinic stimulus, and that nicotinic-subtype receptor is specifically required. Transfection of a series of CgA promoter deletion/luciferase reporter constructs localized the nicotinic response to positive and negative promoter domains; the proximal domain contains a functional cyclic AMP response element. The studies described here, guided by 9 functional hypotheses, will clarify the CgA promoter's cis-elements which mediate nicotinic induction, as well as convergent or divergent receptor or post-receptor signal transduction pathways whereby nicotinic stimulation activates both exocytotic secretion and CgA biosynthesis. Transgenic CgA promoter/beta-galactoside reporter constructs (differing in CgA promoter length) will allow us to test whether promoter domains identified by transfection in vitro are also trans-activated by nicotine in vivo. Understanding nicotinic cholinergic regulation of this catecholaminergic protein's biosynthesis will clarify how gene expression is activated to replete transmitter stores in response to secretion from chromaffin cells and sympathetic axons, and how other pre-ganglionic transmitters as well as chromaffin cell secretory products further modify the transcriptional response.
