Chromostatin inhibits catecholamine secretion in adrenal chromaffin cells by activating a protein phosphatase.

Abstract

Chromostatin is a 20-residue peptide derived from chromogranin A (CGA), the major soluble component of secretory granules in adrenal meduliary chromaffin cells. One known biological function of chromostatin is to inhibit the secretagogue-evoked catecholamine secretion from chromaffin cells. Putative receptors are present on the chromaffin-cell plasma membrane, and the activation of such receptors leads to the inhibition of L-type voltage-sensitive calcium channels.We report here that exposure of chromaffin cells to chromostatin modifies neither cAMP and cGMP levels nor protein CGA remains the topic of many unanswered questions concerning its possible functions, although recent data suggest a possible prohormonal role in the autocrine and/or paracrine modulation ofregulated secretion. The sequence of pancreastatin, a 49-residue peptide from porcine pancreas inhibiting insulin secretion from endocrine pancreatic islets (2, 3), amylase release from exocrine pancreas (4), and acid secretion from parietal cells (5), is fully contained within the sequence of porcine CGA. CGA is also the precursor of chromostatin, a peptide that exerts a negative-feedback control on the secretory activity of chromaffin cells (6, 7). In addition, an inhibitory autocrine role of CGA on proopiomelanocortin secretion (8) and a regulatory role of aminoterminal CGA-derived peptides on the secretion of calcitonin gene-regulating products (9) and on vascular contractile responses (10) have been reported. Together these observations strongly support the view that CGA is a prohormone.In a previous report, we described the purification of two -CGA-derived peptides active on catecholamine secretion from cultured chromaffin cells (7). A corresponding 20-amino acid peptide named chromostatin was synthesized and found to completely inhibit catecholamine secretion in the nanomolar to micromolar range. Bovine chromaffin cells possess specific receptors for chromostatin the activation of which markedly decreases Ca2+ entry through voltage-gated L-type calcium channels (11). The main purpose ofthe present study was to investigate how the inhibitory effect of chromostatin is achieved. We suggest that a protein phosphatase (PPase) may be responsible, at least partially, for the chromostatininduced inhibition of catecholamine release. 45Ca2-Uptake Experiments. 45Ca2+ (13.5 mCi/mg of Ca; CaCl2 salt; New England Nuclear/DuPont; 1 Ci = 37 GBq) influx into cells was measured as described (14, 15). MATERIALS AND METHODSDetermination of PPase Activity. Histone was phosphorylated with cAMP-dependent protein kinase as described by Zwiller et al. (16), and the 32P-phosphorylated histone was recovered according to the procedure reported by Meisler and Langan (17). Chromaffin cells were grown for 3-6 days at a density of 5 x 106 cells on 35-mm-diameter plastic dishes. After extensive washing, cells were incubated for 10 min in Locke's solution in the presence or absence of 100 nM of the indicated peptide. Cells were subsequently scraped off in 5 mM Tris-HCl, ...

DARPP‐32 (dopamine and cAMP‐regulated phosphoprotein, Mr 32,000) is a membrane protein in the bovine parathyroid

“…The data above demonstrate that at least one isoform, PP-ly, is present in both cytosolic and particulate fractions of the parathyroid; this is consistent with observations of PP-Iy in varying ratios in the soluble and particulate fractions in cortex, cerebellum, testis, spleen and adrenal [50]. Type 1 protein phosphatases have a broad range of physiological functions, including the regulation of exocytosis [51]. Parathyroid PP-ly is concentrated at the cell periphery in the vicinity of the plasma membrane; this is consistent with a role in the modulation of PTH secretion.…”

Chromogranin A: Secretion of Processed Products from the Stimulated Retrogradely Perfused Bovine Adrenal Gland

“…Bioactive chromostatin has previously been shown to derive from proteolytic cleavage of larger CGA products in v i m (4,18). Synthetic CGA,,,~,,3 inhibits CA release in isolated bovine chromaffin cells by inhibiting L-type CaZf channels (24) and appears to activate a type 2A phosphoprotein phosphatase in these cells (25). In this context, it is of interest to note that perfusion with aprotinin protected against extracellular cleavage of the larger CGAFs into the 22 to 24 kD components during basal release.…”

Chromostatin, a chromogranin A-derived bioactive peptide, is present in human pancreatic insulin (beta) cells.

“…Characterization of the bovine and mouse CGA genes revealed that the CGA gene contains eight exons (43,44 (9) by activating a protein phosphatase (45) and by decreasing calcium channel activity (46). In the pancreas, PST has been shown to inhibit glucoseinduced insulin secretion (8); the physiological function of CST in this organ has yet to be elucidated.…”