Formation of the catecholamine release-inhibitory peptide catestatin from chromogranin A. Determination of proteolytic cleavage sites in hormone storage granules.
The catestatin fragment of chromogranin A is an inhibitor of catecholamine release, but its occurrence in vivo has not yet been verified, nor have its precise cleavage sites been established. Here we found extensive processing of catestatin in chromogranin A, as judged by catestatin radioimmunoassay of size-fractionated chromaffin granules. On mass spectrometry, a major catestatin form was bovine chromogranin A(332-364); identity of the peptide was confirmed by diagnostic Met(346) oxidation. Further analysis revealed two additional forms: bovine chromogranin A(333-364) and A(343-362). Synthetic longer (chromogranin A(332-364)) and shorter (chromogranin A(344-364)) versions of catestatin each inhibited catecholamine release from chromaffin cells, with superior potency for the shorter version (IC(50) approximately 2.01 versus approximately 0.35 microm). Radioimmunoassay demonstrated catestatin release from the regulated secretory pathway in chromaffin cells. Human catestatin was cleaved in pheochromocytoma chromaffin granules, with the major form, human chromogranin A(340-372), bounded by dibasic sites. We conclude that catestatin is cleaved extensively in vivo, and the peptide is released by exocytosis. In chromaffin granules, the major form of catestatin is cleaved at dibasic sites, while smaller carboxyl-terminal forms also occur. Knowledge of cleavage sites of catestatin from chromogranin A may provide a useful starting point in analysis of the relationship between structure and function for this peptide.
Taylor
C V
CV
Department of Medicine and Center for Molecular Genetics, University of California, and San Diego Veterans Affairs Healthcare System, San Diego, California 92161, USA.
Taupenot
L
L
Mahata
S K
SK
Mahata
M
M
Wu
H
H
Yasothornsrikul
S
S
Toneff
T
T
Caporale
C
C
Jiang
Q
Q
Parmer
R J
RJ
Hook
V Y
VY
O'Connor
D T
DT
D000313:Adrenal Medulla / Q000378:metabolism / Q000648:ultrastructure; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002395:Catecholamines / Q000378:metabolism*; D002417:Cattle; D019439:Chromaffin Cells / Q000378:metabolism; D053379:Chromogranin A; D002864:Chromogranins / Q000096:biosynthesis* / Q000378:metabolism*; D003594:Cytoplasmic Granules / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000096:biosynthesis*; D017386:Sequence Homology, Amino Acid
C506150:CHGA protein, human; D002395:Catecholamines; D053379:Chromogranin A; D002864:Chromogranins; D010446:Peptide Fragments; C109084:chromogranin A (344-364)