SECRETORY PROTEIN TRAFFIC - CHROMOGRANIN-A CONTAINS A DOMINANT TARGETING SIGNAL FOR THE REGULATED PATHWAY

Secretory proteins are targeted into either constitutive (secreted upo n synthesis) or regulated (stored in vesicles and released in response to a secretagogue) pathways. To investigate mechanisms of protein tar geting into catecholamine storage vesicles (CSV), we stably expressed human chromogranin A (CgA), the major soluble protein in human CSV, in the rat pheochromocytoma PC-12 cell line. Chromaffin cell secretagogu es (0.1 mM nicotinic cholinergic agonist, 55 mM K+, or 2 mM Ba++) caus ed cosecretion of human CgA and catecholamines from human CgA-expressi ng cells. Sucrose gradients colocalized human CgA and catecholamines t o subcellular particles of the same buoyant density. Chimeric proteins , in which human CgA (either full-length 1457 amino acids] or truncate d [amino-terminal 226 amino acids]) was fused in-frame to the ordinari ly nonsecreted protein chloramphenicol acetyltransferase (CAT), were e xpressed transiently in PC-12 cells. Both constructs directed CAT acti vity into regulated secretory vesicles, as judged by secretagogue-stim ulated release. These data demonstrate that human CgA expressed in PC- 12 cells is targeted to regulated secretory vesicles. In addition, hum an CgA can divert an ordinarily non-secreted protein into the regulate d secretory pathway, consistent with the operation of a dominant targe ting signal for the regulated pathway within the peptide sequence of C gA.