Bovine Parathyroid Glands Secrete a 26-kDa NTerminal Fragment of Chromogranin-A which Inhibits Parathyroid Cell Secretion*

Abstract

Chromogranin-A (CgA) is a ubiquitous protein which colocalizes in secretory granules of multiple endocrine tissues and cosecretes with peptide hormones from these tissues. Although the function of CgA has remained unknown, there has been recent interest in its potential role as a prohormone for smaller, biologically active peptides. We isolated and characterized a 26-kDa N-terminal fragment of CgA which is a natural breakdown product of bovine parathyroid CgA in storage. A similar, if not identical, fragment of CgA is secreted by bovine parathyroid glands. The secreted fragment elutes on HPLC and migrates on both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and acid-urea gels in the same position as the 26-kDa N-terminal fragment. When added to the incubation medium of dispersed bovine parathyroid cells, the 26-kDa N-terminal fragment of CgA inhibits the low calcium-stimulated secretion of both PTH and CgA. This N-terminal fragment is homologous to betagranin, which is a 21-kDa N-terminal fragment of CgA that is generated from CgA in rat insulin granules. Thus, a naturally occurring betagranin-like N-terminal fragment of bovine parathyroid CgA is not only secreted itself, but can inhibit the secretion of PTH and intact CgA by bovine parathyroid cells. The processing of intact CgA to fragments such as the N-terminal fragment that we describe may be important in the autocrine or paracrine regulation of secretion.

Chromogranin A processing and secretion: specific role of endogenous and exogenous prohormone convertases in the regulated secretory pathway.

“…2), a precursor of biologically active peptides (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12), contains multiple pairs of basic amino acids which are potential sites for cleavage by the endoproteases PC1 and PC2.…”

Characterisation of N-terminal chromogranin A and chromogranin B in mammals by region-specific radioimmunoassays and chromatographic separation methods

“…Hence, these data can be interpreted in two ways, either that the serum concentrations of CgA, or the degree of N-terminal processing to yield free vasostatin, differ among these mammals. Synthetic vasostatin peptides, covering the sequences bCgA1-40 and bCgA1-76, have previously been demonstrated to possess inhibitory actions in the 10-100 nM range in human blood vessels (Aardal & Helle 1992) and on bovine parathyroid cells (Fasciotto et al 1990, Drees et al 1991. These concentrations of vasostatin are more likely to arise in the circulation of the sheep, goat, horse and pig than in cattle and man.…”

“…Some specific domains of the molecules have been assigned with different biological activity. First, the N-terminal domains of CgA, also called vasostatin, have suppressive effects on vascular contractility (Aardal & Helle 1992) and on parathormone secretion (Fasciotto et al 1990, Drees et al 1991. Secondly, the central part of CgA, with defined peptide hormones such as pancreastatin, parastatin and catestatin, has paracrine-or autocrine-inhibiting effects on the secretion of other hormones, such as insulin (Tatemoto et al 1986) and catecholamines (Mahata et al 1997).…”

Effects of calcium on recombinant bovine chromogranin A

“…Analysis by SDS-PAGE (data not shown) suggests that both of these peaks are aminoterminal fragments of CGA. These fragments are endogenously processed in chromaffin granules at the first two pairs of basic residues and are the molecular species active in vasodilation assays (Aardal et al, 1992) and in assays for inhibition of the secretion of parathyroid hormone (Drees et al, 1991). Peak 4 also exhibits a single aminoterminus characteristic of CGA and is eluted 2.1 min earlier than the recombinant CGA under the same elution conditions.…”

“…CGA is a precursor for several peptides recently reported to have biological activity: pancreastatin, an amidated peptide that suppresses glucosestimulated insulin secretion in pancreatic islets (Tatemoto et al, 1986;Eiden, 1987;; chromostatin, a peptide reported to arrest catecholamine secretion from adrenal chromaffin cells (Galindo et al, 1991(Galindo et al, , 1992; and vasostatin, the amino-terminal peptide that inhibits the contraction of venous segments stimulated by K+, noradrenalin, or endothelin (Aardal et Abbreviations: CD, circular dichroism; CGA, chromogranin A; HPLC, high pressure liquid chromatography; m/z, mass-to-charge ratio; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TIC, total ion current. 1992), and which has also been reported to inhibit parathyroid hormone secretion stimulated by low calcium (Drees et al, 1991). CGA also binds calcium and catecholamines and self associates in the presence of calcium at high protein concentration (Cohn et al, 1981;Reiffen & Gratzl, 1986a,b;Leiser & Sherwood, 1989;Yo0 & Albanesi, 1990; Videen et al, 1992).…”