Regulation of Chromogranin A and Chromogranin B (Secretogranin I) Synthesis in Bovine Cultured Chromaffin Cells

Abstract

in the present study we investigated the regulation of chromogranin A (CGA) and chromogranin B (CGB) biosynthesis in bovine shromaffin cells maintained in primary culture. Cellular proteins were labelled with [35S]methionine and the incorporated radioactivity was used as an index of the s y n t h e s i s rate. The radioactivity incorporated into CGA was determined by immunoprecipitation, m d that into CGB was quantified by a dot immunobinding assay using specific antibodies. Incubation of cells with carbamylcholine, .iigh K' or histamine, three potent stimulators of catecholamine secretion in chromaffin cells, increased the rate of CGA and CGB synthesis. On the other hand bradykinin, angiotensin II and prostaglandin E, , which cause little secretion, also produced an ,ncrease in both CGA and CGB synthesis. These results suggest that in chromaffin cells, the biosynthesis of chromogranins is not closely linked to the secretory activity. Inhibition of protein kinase C by sphingosine or by long-term treatment with phorbol esters, completely abolished the synthesis of CGA and CGB induced by carbamylcholine, bradykinin and prostaglandin E, but decreased only partially the stimulating effect of histamine. Thus, protein kinase C may not be the sole effector involved in t h e secretagogueinduced modulation of chromogranin synthesis. Forskolin, an activator of adenylate cyclase had no effect on CGA synthesis, but significantly enhanced the incorporation of radioactivity into CGB. The effect of forskolin was not modified by protein kinase C inhibitors and was additive to that induced by phorbol esters indicating that cyclic AMP did not stimulate CGB synthesis through a protein kinase C-dependent pathway. These observations suggest that the biosynthesis of CGA and CGB in chromaffin granules IS independently regulated.The secretory granules of adrenal medullary chromaffin cells contain a high concentration of soluble peptides and proteins, [hat are co-released with catecholamines into the circulation in response to splanchnic nerve stimulation. The soluble acidic proteins of chromaffin granules have been collectively called chromogranins. Chromogranin A (CGA) and chromogranin B (CGB) also known as secretogranin I, together constitute about 50% of the total soluble proteins. A third, highly acidic, though less abundant protein is found in chromafin granules: chromogranin C also named secretogranin 11. Chromogranins are widely distributed throughout tissues of the neuroendocrine system including thyroid and parathyroid, pituitary, pancreatic islets and various cells of the endocrine gut in addition to adrenal medulla and adrenergic neurons (for review see I). Chromogranins are therefore increasingly used as markers of normal and neoplastic neuroendocrine cells (2-4).The different members of the chromogranin family share certain structural features: they are extremely heat stable, highly acidic proteins, capable of binding C a Z f with low affinity but high capacity ( 5 , 6), and are post-translationally modified by ph...

Processing of Chromogranin B in Bovine Adrenal Medulla. Identification of Secretolytin, the Endogenous C-Terminal Fragment of Residues 614-626 with Antibacterial Activity

“…Different results are consistent with the hypothesis that a key step in the sorting of the chromogranins is their selective, calcium-induced and low pH-induced aggregation in the trans-Golgi network, resulting in the formation of dense cores that exclude constitutive secretory proteins (Huttner et al, 1991). Previously, in our laboratory (Galindo et al, 1991), we reported results indicating (a) that in chromaffin cells, the biosynthesis of chromogranins is not closely linked to the secretory activity and (b) that the biosynthesis of CGA and CGB in chromaffin granules is independently regulated. The relative distribution of CGA and CGB in different species and the different degrees of processing suggest that the two proteins are each regulated by specific mechanisms.…”

The N‐ and C‐terminal fragments of ubiquitin are important for the antimicrobial activities

“…Chromaffin cells (2×10 8 ) were isolated from fresh bovine adrenal glands, cultured as previously reported (18), and plated at a density of 10 7 cells/100 mm in plastic Petri dishes. After 3 days in culture, cells were washed four times with Locke's solution (140 mM NaCl, 4.7 mM KCl, 1.2 mM MgSO 4 , 2.5 mM CaCl 2 , 11 mM glucose, 0.5 mM ascorbic acid, 15 mM Hepes, pH 7.4) to clean out dead cells or cell debris (19). The extracellular medium of nonstimulated cells was collected and used as a control.…”

Processing of Chromogranin B in Bovine Adrenal Medulla. Identification of Secretolytin, the Endogenous C-Terminal Fragment of Residues 614-626 with Antibacterial Activity

“…The relative distribution of CGA and CGB in different species and the different degrees of processing suggest that the two proteins are each regulated by specific mechanisms. Previously, in our laboratory (Galindo et al, 1991), we reported results indicating (a) that in chromaffin cells, the biosynthesis of chromogranins is not closely linked to the secretory activity and (b) that the biosynthesis of CGA and CGB in chromaffin granules is independently regulated.…”