Molecular Cloning and Functional Analysis of Secretory Phospholipase A2 from Apostichopus japonicus.

Abstract

Secretory phospholipase A2 (sPLA2) plays important roles in phospholipid metabolism, skin barrier maintenance, immune response and other processes in organisms. sPLA2 of sea cucumber A. japonicus (AjPLA2) has not yet been reported. This study successfully amplified the AjPLA2 sequence. The total cDNA of AjPLA2 is 931bp, including a 480bp ORF that encodes 159 amino acids. The AjPLA2 protein includes a 16-aa signal peptide, a 5-aa precursor peptide and a 138-aa mature peptide. Homologous alignment showed that AjPLA2 and the sPLA2s from starfish have the typical domains of the Group IB sPLA2. And additional amino acid sequences were found around the β-Wing, which is different from the Group IB sPLA2. These results showed that AjPLA2 and sPLA2s from starfish all belong to a new group in the Group I sPLA2 family. AjPLA2 is widely distributed in sea cucumber tissues. The functional analysis also showed that AjPLA2 was upregulated in the intestine by feeding. Whenthe body wall was damaged, it was significantly upregulated around the wound. And the expression levels of AjPLA2 were significantly increased in V. splendens-infected sea cucumbers. The results indicated that AjPLA2 plays roles in the sea cucumber immunologicprocess. Combined with the upregulation of unsaturated fatty acids (PUFAs) content in A. japonicus, it demonstrated that AjPLA2 could participate in the immune of A. japonicus by hydrolyzing phospholipid and releasing PUFAs. This study had a solid foundation for the further research of AjPLA2 gene function in vivo, development and application of AjPLA2 protein.