Faculty Bibliography

Chromogranin A is a major component of storage granules in many different secretory cell types. After [35S]methionine labelling of proteins from cultured bovine chromaffin cells, chromogranin A was immunoprecipitated with specific antibodies, and the radioactivity incorporated into chromogranin A was determined and used as an index of its synthesis rate. Depolarization of cells with nicotine or high K+ evoked a Ca2+-dependent increase in chromogranin A synthesis, whereas muscarine, which does not evoke significant Ca2+ influx from bovine chromaffin cells, had no effect on chromogranin A synthesis. Forskolin, an activator of adenylate cyclase, affected neither the basal nor the nicotine-stimulated rate of chromogranin A synthesis. In contrast, 12-O-tetradecanoylphorbol 13-acetate (TPA), an activator of protein kinase C, significantly enhanced the incorporation of radioactivity into chromogranin A. Sphingosine, an inhibitor of protein kinase C, abolished both nicotine-stimulated and TPA-induced chromogranin A synthesis. In addition, long-term treatment of chromaffin cells with TPA decreased protein kinase C activity and inhibited the nicotine-stimulated chromogranin A synthesis. These results suggest that protein kinase C may play an important role in the control of chromogranin A synthesis.

Chromogranin A (CGA) is the major protein of the secretory granule from chromaffin cells and also is found in a variety of endocrine cells. Although the sequence of this acidic glycoprotein has been elucidated recently, its biological function is unknown. Here we have purified CGA from chromaffin granules; the final preparation contained the 74-kDa native CGA together with two degradation products--three bands near 60 kDa and a single band of 43 kDa. This preparation was found to inhibit (a maximum inhibition of 60% at 1 microM) the nicotine-induced, but not the high K+-evoked, catecholamine secretion from bovine chromaffin cells maintained in primary culture. Spontaneous release was also affected in the nanomolar CGA protein concentration range. The observation that the inhibitory effect is strictly dependent on a preincubation step together with the modification of the CGA protein profile during this preincubation step suggests that the degradation peptide(s) rather than the 74-kDa native CGA--the approximately equal to 60-kDa bands or the 43-kDa singlet band--is actually involved in secretory cell activity. This was demonstrated by using trypsin-generated peptides that were inhibitory without the preincubation period. The finding that unprocessed CGA is not active on chromaffin cell secretion suggests that this molecule is a precursor of a peptide(s) that is able to regulate catecholamine secretion. Thus, the present data suggest that a CGA-derived peptide(s) could exert a feedback control on chromaffin cell secretory activity--a mechanism that might be of importance during stress situations.