Cloning and characterization of allograft inflammatory factor-1 (AIF-1) from Manila clam Venerupis philippinarum

Allograft inflammatory factor-1 (AIF-1) is a 17 kDa interferon-γ-inducible Ca(2+)-binding EF-hand protein that plays a significant role not only in different host responses to inflammatory stimuli, but in the whole host immune defense reaction. In the present study, the full-length cDNA of AIF-1 was identified from manila clam Venerupis philippinarum (denoted as VpAIF-1) by cDNA library and RACE approaches. The cDNA of VpAIF-1 consisted of a 5-terminal untranslated region (UTR) of 153 bp, a 3'UTR of 219 bp with a poly (A) tail, and an open reading frame (ORF) of 516 bp encoding a polypeptide of 171 amino acids with the putative molecular mass of 17 kDa. The deduced amino acid of VpAIF-1 shared two EF hand Ca(2+)-binding motifs like other AIF-1s. Phylogenetic analysis further indicated that VpAIF-1 had higher evolutional conservation to invertebrate than vertebrate counterparts and should be a new member of the AIF-1 protein family. Spatial expression analysis indicated that mRNA transcript of VpAIF-1 was found to be most abundantly expressed in the tissues of haemocytes, gills and hepatopancreas, weakly expressed in the tissues of mantle, muscle, and foot. After challenged by Vibrio anguillarum, the mRNA level of VpAIF-1 in overall haemocytes population was recorded by quantitative real-time RT-PCR. VpAIF-1 mRNA was down-regulated in the first 12 h post-infection. Then, the expression level increased to the peak at 72 h and recovered to the 48 h-level at 96 h. All these results indicated that VpAIF-1 was involved in the immune response against microbe infection and might be contributed to the clearance of bacterial pathogens.