Cloning and expression of group IB phospholipase A(2) isoforms in the red sea bream, Pagrus major

Two cDNA encoding red sea bream DE-1 and DE-2 phospholipases A(2) (PLA(2)) were cloned from the hepatopancreas of red sea bream, Pagrus (Chrysophrys) major. The cDNA of DE-1 PLA(2) encoded a mature protein of 125 amino acid r esidues with an apparent signal peptide of 20 residues and propeptide of 5 residues, and that of DE-2 PLA(2), a mature protein of 126 amino acid resid ues with an apparent signal peptide of 17 residues and propeptide of 6 resi dues. Comparison of the predicted amino acid sequences for mature DE-1 and DE-2 PLA(2) showed that both proteins contain 14 cysteines including Cys 11 and 77 and a pancreatic loop, which are commonly conserved in group IB PLA (2); however, the identity in amino acid sequence between DE-1 and DE-2 PLA (2) was low (47%). A previous report concerning the cDNA cloning of red sea bream gill G-3 PLA(2) and the present results represent the first cloning and sequencing of three distinct isoforms of group IB PLA(2) in a single fi sh species, red sea bream. Reverse transcription-polymerase chain reaction analysis showed that DE-1 PLA(2) mRNA was expressed in the hepatopancreas, pyloric ceca, intestine, spleen, gonad, stomach, and kidney, whereas gill G -3 PLA(2) mRNA was expressed only in the gills and gonad. The expression of DE-2 PLA(2) mRNA was detected in all of the tissues analyzed. These result s indicate that three distinct isoforms of group IB PLA(2), DE-1 and DE-2 P LA(2) in hepatopanceas and gill G-3 PLA(2), are expressed in a tissue-speci fic manner in red sea bream.