cDNA cloning and expression of a novel group IB phospholipase A2 in the intestine of the red sea bream,Pagrus (Chrysophrys) major

A cDNA encoding a novel phospholipase A₂ (PLA₂), which was named IN PLA₂, was cloned from the intestine of the red sea bream. The amino acid sequence of IN PLA₂ showed 49–75% homology with those of red sea bream group IB sPLA₂, hepatopancreas DE-1 and DE-2 PLA₂, and gill G-3 PLA₂. IN PLA₂ consists of a prepropeptide of 24 amino acid residues, followed by a mature protein. IN PLA₂ contains 14 cysteines, and includes Cys11, the calcium binding loop and the pancreatic loop that are commonly conserved in group IB sPLA₂ enzymes. In addition, IN PLA₂ is a cationic protein with a pI of 8.52. Therefore, IN PLA₂ was identified as a novel group IB sPLA₂ isoform in red sea bream. IN PLA₂ mRNA was found by northern blot analysis to be expressed mainly in the pyloric caeca and the intestine, and was detected in the goblet cells of the intestine by in situ hybridization. The expression level of IN PLA₂ mRNA was elevated by intravenous injection of lipopolysaccharide—the outer-membrane component of Gram-negative bacteria. These results suggest that IN PLA₂ is secreted from the goblet cells of the intestine in response to stimulus such as bacterial infection, and that it contributes to antimicrobial defense in addition to the digestion of dietary phospholipids in the gastrointestinal tract.