Molecular cloning of chromogranin A from rat pheochromocytoma cells.
Chromogranin A (CgA) is the major soluble protein in catecholamine storage vesicles. To gain insight into its function, we isolated CgA clones from a size-selected lambda gt10 rat pheochromocytoma complementary DNA (cDNA) library. The longest cDNA insert identified was 2.2 kb and encoded the entire 462-amino acid open reading frame of rat CgA including an 18-amino acid hydrophobic signal peptide. Comparison of rat CgA with the recently published sequences of bovine CgA and human CgA revealed regions of strong homology at the N-and COOH-termini as well as variant areas predominantly in the middle portion of the molecule. Regions highly conserved and therefore suggestive of functional importance included 1) multiple paired basic residues, which may serve as proteolytic processing signals; 2) a region homologous to porcine pancreastatin, a putative modulator of peptide hormone release; and 3) a short hydrophobic disulfide loop region near the N-terminus that may have a role in the targeting of CgA to secretory vesicles. On the other hand, lack of conservation of the membrane attachment sequence arginine-glycine-aspartic acid argues against its functional importance in CgA. In addition, the presence of a unique polyglutamine region in rat CgA points to a possible messenger RNA (mRNA) splice junction. Northern blot experiments demonstrated the presence of an approximately 2.2 kb rat CgA mRNA in a neuroendocrine distribution (adrenal, brain, pheochromocytoma cells, but not skeletal muscle, heart, or kidney). Southern blot studies were consistent with the presence of a single CgA gene within the rat pheochromocytoma cell genome. Finally, comparison of the present rat pheochromocytoma cDNA clones with those recently obtained from normal rat adrenal gland reveals minor but apparently real differences that suggest CgA microheterogeneity.
Parmer
R J
RJ
Department of Medicine, University of California, San Diego.
Koop
A H
AH
Handa
M T
MT
O'Connor
D T
DT
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D053379:Chromogranin A; D002864:Chromogranins / Q000235:genetics*; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D015723:Gene Library; D005796:Genes; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics*; D015345:Oligonucleotide Probes; D010673:Pheochromocytoma / Q000235:genetics*; D011110:Polymorphism, Genetic; D011487:Protein Conformation; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D015183:Restriction Mapping; D014407:Tumor Cells, Cultured
C506150:CHGA protein, human; D053379:Chromogranin A; D002864:Chromogranins; D009419:Nerve Tissue Proteins; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; C046560:beta-granins; C506153:chromogranin A, rat; D004247:DNA