Primary structures and gene organizations of two types of Wap65 from the pufferfish Takifugu rubripes

We isolated two types of cDNA clone encoding hemopexin-like protein, fWap65-1 and fWap65-2, from the pufferfish Takifugu rubripes. The deduced amino acid sequence of fWap65-1 showed 66-67% identity with those of goldfish and carp Wap65s, whereas the sequence of fWap65-2 did 44-46% identity. Both fWap65s showed 32-38% amino acid identity with mammalian hemopexins, in which fWap65-2 was more related than fWap65-1. While hemopexins contain two conserved histidine residues in their heme binding pockets, these residues were also conserved in fWap65-2, but not in fWap65-1. The exon-intron organization was highly conserved between fWap65s and human hemopexin gene, suggesting that Wap65s are fish orthologs of human hemopexin. The 5'-flanking regions of both fWap65s contained various putative transcriptional elements, including Cdx1, GATA-1, C/EBPbeta and LyF-1. The expression patterns of fWap65s in various tissues of Fugu were examined by RT-PCR, demonstrating the dominant expression of both genes in liver followed by brain. In addition, the small quantities of fWap65-1 transcripts were also detected in eye, gill and gonad, whereas the transcripts of fWap65-2 could not be observed except for liver and brain. Although the average values for mRNA levels of both fWap65s in warm-acclimated fish tended to be higher than those of cold-acclimated fish, their differences were not statistically significant.