Cloning and expression of phospholipase A2 from guinea pig gastric mucosa, its induction by carbachol and secretion in vivo.
A cDNA encoding phospholipase A2 (PLA2) was cloned from guinea pig gastric mucosa using a rat pancreatic-PLA2-cDNA fragment as a probe. The cDNA contains an open reading frame sufficient to encode the entire amino-acid sequence of a PLA2-precursor protein consisting of 146 amino acids, including a putative 16-residue signal peptide and a 6-residue activation peptide at the NH2-terminus. Its nucleotide sequence exhibits 70% similarity to that of rat pancreatic PLA2 cDNA. The deduced amino-acid sequence has all the typical pancreatic PLA2 characteristics, with the exception of the substitution of Phe for Tyr at position 28 in the calcium-binding loop of the mature enzyme. When an expression vector containing the PLA2 cDNA was transfected into COS-7 cells, a major portion of the proenzyme was secreted into the culture medium. Northern-blot analysis showed the mRNA was present in guinea pig lung and pancreas at much lower levels than in the stomach. The effect of carbachol, a muscarinic acetylcholine agonist, on the secretion of gastric PLA2 and on its mRNA level in the gastric mucosa were examined. PLA2 secretion into the gastric juice was maximal 30 min after the subcutaneous administration of carbachol (0.4 mg/kg). It also increased the PLA2-mRNA level in the tissue, the maximal mRNA level being delayed about 15 min compared with that in PLA2 secretion. These results suggest that vagal stimuli may contribute to PLA2 secretion and its compensatory synthesis, and that the secreted PLA2 may participate in the digestion of dietary and biliary phospholipids in the small intestine of guinea pig.
Ying
Z
Z
Department of Molecular Physiological Chemistry, Osaka University Medical School, Japan.
Tojo
H
H
Nonaka
Y
Y
Okamoto
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002217:Carbachol / Q000494:pharmacology*; D002460:Cell Line; D003001:Cloning, Molecular; D004790:Enzyme Induction / Q000187:drug effects; D005753:Gastric Mucosa / Q000201:enzymology*; D006168:Guinea Pigs; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D054467:Phospholipases A2; D012333:RNA, Messenger / Q000032:analysis
D012333:RNA, Messenger; D002217:Carbachol; D010741:Phospholipases A; D054467:Phospholipases A2