PROCESSING OF CHROMOGRANINS IN CHROMAFFIN CELL-CULTURE - EFFECTS OF RESERPINE AND ALPHA-METHYL-P-TYROSINE

Bovine chromaffin cell cultures were treated with either reserpine or alpha-methyl-p-tyrosine for up to 10 days. Afterwards the cells were h arvested and the degree of proteolytic processing of secretogranin II, chromogranin A and chromogranin B was determined by immunoblotting an d HPLC followed by RIA. There was a significant increase in the proteo lysis of all three chromogranins after 4-6 days in the presence of res erpine. The small peptides formed in the presence of reserpine in vitr o are also produced in vivo. A similar effect was observed with alpha- methyl-p-tyrosine, an inhibitor of tyrosine hydroxylase, but the respo nse took up to 10 days to develop. Both drugs decreased catecholamine levels but reserpine was more effective, reaching a high degree of dep letion after 4 days. In addition, experiments in vitro indicate that l ow millimolar amounts of either adrenaline (IC50 5.2 mM) or noradrenal ine (IC50 2.4 mM) can significantly impair the proteolytic activity of recombinant murine prohormone convertase 1 when assayed with syntheti c fluorogenic and/or peptidyl substrates. We conclude that a lowering of catecholamine levels in chromaffin granules leads to a concomitant increase in proteolytic processing of all secretory peptides. Apparent ly within chromaffin granules the endoproteases are inhibited by catec holamines and thus their removal leads to increased proteolysis.