Molecular Characterization of Differentially Expressed ADP Ribosylation Factor from WSSV Infected Black Tiger Shrimp Penaeus monodon

Abstract

Suppression subtractive hybridization (SSH) cDNA library was constructed using hepatopancreas of the WSSV infected black tiger shrimp, Penaeus monodon. The differentially expressed ESTs of SSH cDNA library represented different functional categories belonging to energy and metabolism, osmoregulation, cytoskeletal and structural proteins, membrane traffic and organelle structure, immune genes, protein folding, transport and synthesis, ovarian and oocyte development, nucleic acid processing, cell cycle and heat shock protein. The ADP ribosylation factor (Arf) was one of the differentially expressed gene which was identified from the SSH cDNA library. The full length cDNA of P. monodon Arf (pmArf) was cloned by 3′- and 5′-rapid amplification of cDNA ends (3′/5′ RACE). The complete nucleotide sequence of pmArf encoded protein sequence of 180 amino acids revealed the characteristic conserved features of the Arf family proteins represented by the P-loop, the switch regions and interswitch region. The sequence homology indicated that the pmArf belonged to class II of Arf family. The gene expression analysis of pmArf and other immune related genes (penaeidin, anti-lipopolysaccharide factor and QM) in the gills, hepatopancreas and gut tissues of the WSSV infected shrimp was carried out by Real-Time PCR. The pmArf gene was up-regulated with highest gene expression levels at 24 h (1.33-fold) in the gill tissues and at 48 h in the gut tissues (1.44-fold) and hepatopancreas (1.46-fold) of WSSV infected shrimp, indicating its potential role in immune response against WSSV infection.