Chromogranin A-derived Peptide Research Articles

Distribution and characterisation of the chromogranin A-derived peptide, WE-14, in rat neuroendocrine tissues

Distribution and characterisation of the chromogranin A-derived peptide, WE-14, in rat neuroendocrine tissues

Isolation and primary structure of a novel chromogranin A-derived peptide, WE-14, from a human midgut carcinoid tumour

The primary structure of a novel human chromogranin A-derived tetradecapeptide, WE-14, possessing N-terminal tryptophanyl(W) and C-terminal glutamyl (E) residues was isolated from a hepatic metastasis of an human ileal carcinoid tumour. Human and bovine WE-14 are structurally identical, while rat, mouse and porcine analogues exhibit 93 % homology. WE-14 is flanked by paired basic residues (KR) in all known chromogranin A sequences.

Chromogranin A: localization and stoichiometry in large dense core catecholamine storage vesicles from sympathetic nerve

Chromogranin A is present in both adrenal medullary chromaffin granules and sympathetic nerve large dense core catecholamine storage vesicles (LDVs), yet selective stimulation of sympathetic axons provokes only minor changes in chromogranin A in the circulation. We therefore examined the stoichiometry of chromogranin A storage in purified LDVs as compared to chromaffin granules. Chromogranin A was found in LDVs on immunocytochemical sections of sympathetic axons. Sedimentation of sympathetic axon homogenates on sucrose-D 2O gradients localized chromogranin A, norepinephrine, enkephalins and dopamine-β-hydroxylase to the same gradient particulate fractions, suggesting that they inhabit a particle of the same buoyant density, the LDV. Chromogranin A was identified in LDV by radioimmunoassay, immunoblotting and immunocytochemistry. Purified LDVs contained 17.8 ±4.8% of cell total chromogranin A, at 27.9 ± 3.5-fold enrichment over the original axon homogenate. When LDVs were lysed, all of the chromogranin A immunoreactivity originated from the soluble vesicle core rather than the LDV membrane. Although chromogranin A/catecholamine ratios were similar in LDVs and adrenal chromaffin granules, chromogranin A was a quantitatively minor protein in LDVs, accounting for only 0.16 ± 0.015% of total LDV protein, as compared to 35.2 ± 1.4% of total chromaffin granule protein. Each LDV particle contained approximately 0.94 ± 0.09 chromogranin A molecules. Immunocytochemical data suggested that chromogranin A is costored in large dense core noradrenergic vesicles in subpopulations of sympathetic axons, analogous to enkephalins and neuropeptide Y. Thus, only profound changes in exocytotic catecholamine release from sympathetic axon LDVs would be expected to perturb circulating chromogranin A concentrations. Since chromogranin A-derived peptides also seem to modulate catecholamine release, these results may have functional consequences for secretory control in sympathetic axons.

The tissue distribution of rat chromogranin A-derived peptides: evidence for differential tissue processing from sequence specific antisera

The distribution of chromogranin A and related peptides in rat tissues was investigated using sequence specific antisera. N- and C-terminal antisera and a presumptive C-terminal rat pancreastatin antiserum immunostained an extensive neuroendocrine cell population throughout the gastro-entero-pancreatic tract, anterior pituitary, thyroid and all adrenomedullary cells. However, mid- to C-terminal antisera immunostained a subpopulation of chromogranin A positive cells. Most notable of these was with the KELTAE antiserum (R635.1) which immunostained discrete clusters of adrenomedullary cells and antiserum A87A which immunostained a subpopulation of cells in the anterior pituitary and throughout the gastrointestinal tract. The present study has demonstrated the widespread occurrence of chromogranin A and related peptides in rat neuroendocrine tissues and provides evidence of tissue and cell specific processing.

Heterogeneity of chromogranin A-derived peptides in bovine gut, pancreas and adrenal medulla

Chromogranin A is produced in many endocrine cell types, and is widely used as a marker in endocrine-cell pathology and secretory-cell biology. There is some evidence that it may be proteolytically processed to yield the putative pancreatic regulatory peptide, pancreastatin, and, in order to characterize the relevant pathways in gastrointestinal and pancreatic endocrine cells, we have used, in radioimmunoassay, site-directed antibodies to pancreastatin itself (L331) and to a sequence of chromogranin A immediately C-terminal to pancreastatin (L300). The latter antibody revealed three major forms of immunoreactivity of 8 kDa and five peptides of approx. 3 kDa in bovine pancreas and gut extracts. The 8 kDa peptides were characterized as chromogranin A-(248-313)-peptides, i.e. C-terminally extended forms of pancreastatin; two of the 8 kDa variants differed in two positions, confirming a polymorphism predicted from cDNA sequencing. One of the 3 kDa peptides was characterized as chromogranin A-(297-313)-peptide, i.e. the C-terminal heptadecapeptide of the 8 kDa peptide that would be liberated after cleavage to yield pancreastatin. On the basis of chromatographic studies, immunohistochemistry and the stoichiometry of different immunoreactive peptides, three different pathways of chromogranin A processing were identified: in adrenal chromaffin cells chromogranin A existed mainly as the unmodified intact protein, in pancreatic islet and gastric antral endocrine cells pancreastatin and the 3 kDa peptides were major products, but in small intestine and gastric corpus endocrine cells there was little nor no pancreastatin and the 8 kDa cleavage product predominated. There are therefore important differences in the distribution of chromogranin A-derived peptides between quite closely related populations of endocrine cells that are attributable not only to variable post-translational cleavage but also to the expression of different primary sequences. It seems possible that in different cell types chromogranin A-derived peptides might subserve a variety of different functions.

Chromogranin A: Posttranslational Modifications in Secretory Granules

The primary structure of chromogranin A indicates multiple domains which might be subject to posttranslational modification. We explored chromogranin A's proteolytic cleavage, glycosylation, and possible intermolecular disulfide links, using biochemical and cell biological approaches. Anti-chromogranin A region-specific immunoblots on chromaffin granules suggested bidirectional endoproteolytic cleavage of chromogranin A; control experiments ruled out artifactual cleavage during granule isolation or lysis. Isolation of chromogranin A-derived peptides by gel filtration chromatography or sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), followed by N-terminal amino acid sequencing, established several cleavage sites, including at least two at dibasic sites. Secretion of chromogranin A from bovine chromaffin cells did not initiate further cleavage, nor did prolonged exposure of secreted chromogranins to the secretory cells. The chromogranin A cleavage pattern was qualitatively similar in other neuroendocrine tissues, though cleavage was more complete in adrenal medullary than in anterior pituitary hormone storage vesicles, and N-terminal fragments of 45 and 55 kilodaltons were more prominent in the hypothalamus. A similar cleavage pattern was seen in human pheochromocytoma granules, as judged by chromogranin A region-specific immunoblots, fragment isolation by SDS-PAGE, and microsequencing. The presence of full-length chromogranin A as the core protein of a chromaffin granule soluble proteoglycan was suggested in bovine (but not human) chromaffin granules by glycoprotein staining, chondroitinase ABC digestion, chemical deglycosylation, and region-specific immunoblotting. Human (but not bovine) chromogranin A displayed intermolecular disulfide crosslinks on SDS-PAGE gels and immunoblotting. These results document diverse structural paths that the chromogranin A molecule may take in endocrine secretory cells after its translation.

Rapid publication a chromogranin a-derived peptide differentially regulates the secretion of calcitonin gene products

We studied the regulation of the secretion of CT and CGRP by chromogranin A (CgA)-derived peptides in a human lung tumor cell line. The amino-terminal peptide of CgA, CgA1-40, stimulated the secretion of CGRP and inhibited the secretion of CT; both effects occurred in a dose-dependent manner. These studies demonstrate a differential effect of a CgA-derived peptide on two products of the CT gene, CT itself and CGRP. CgA may be processed at its multiple dibasic sites to peptides that specifically regulate the secretion of its coresident hormones, in this case two calcitonin gene products that are present in the same secretory vesicle. This novel mechanism represents a new pathway for the control of calcium regulating hormones.

Differential accumulation of catecholamines, proenkephalin- and chromogranin A-derived peptides in the medium after chronic nicotine stimulation of cultured bovine adrenal chromaffin cells

We have used an antiserum to a synthetic peptide fragment of bovine chromogranin A (ChrgA)[Tyr o] bovine ChrgA (306–313): YLSKEWEDA, together with antibodies to proenkephalin-derived peptides, to measure the release of immunoreactive peptides from nicotine-stimulated cultured bovine adrenal chromaffin cells. Over a period of 6 hr the accumulation of YLSKEWEDA immunoreactivity and Met-enkephalin Arg 6Gly 7Leu 8 (MERGL) immunoreactivity in the medium of 10 μM nicotine-stimulated cells was shown to be biphasic; the initial phase occurred in the first 15–30 min and the second phase reached a peak after 4 hr. In contrast, catecholamine release occurred monophasically over the initial 15–30 min. Investigation of the second phase of peptide accumulation revealed that it was due in part to nicotine-evoked exocytosis and in part to extracellular processing of high molecular weight precursor proteins.

Differential accumulation of catecholamines and of proenkephalin A- and chromogranin A-derived peptides during chronic nicotine stimulation of cultured bovine adrenal chromaffin cells

Differential accumulation of catecholamines and of proenkephalin A- and chromogranin A-derived peptides during chronic nicotine stimulation of cultured bovine adrenal chromaffin cells

Isolation and characterization from bovine ileum of chromogranin A-derived peptides related to pancreastatin

Isolation and characterization from bovine ileum of chromogranin A-derived peptides related to pancreastatin

Secretion from chromaffin cells is controlled by chromogranin A-derived peptides.

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.