The warm temperature acclimation-related 65-kDa protein, Wap65, in goldfish and its gene expression

Abstract

cDNAs encoding a warm temperature acclimation-related protein (Wap65) were cloned from the muscle and hepatopancreas cDNA libraries of the warm temperature-acclimated goldfish Carassius auratus, and their nucleotide sequences containing 5'- and 3'-noncoding regions together with their polyadenylation signal were determined. The deduced amino acid sequence of Wap65 was 31% homologous to rat hemopexin. However, goldfish Wap65 lacked a few possible glycosylation sites and presumed functional histidine residues, implying that it may have different functions from hemopexin. Wap65 contained a leader peptide of 30 amino acids and a mature protein region of 415 amino acids. Southern blot analysis demonstrated that the protein is expressed by a single copy gene in the goldfish haploid genome. In RNA blot analysis using isolated cDNA clones, a single transcript of about 2.0 kilobases was detected in the hepatopancreas but not in brain, muscle, or hemocytes. The abundancy of this transcript markedly increased in the hepatopancreas as a result of warm temperature acclimation. Electrophoretic analysis of plasma proteins revealed a good correlation of plasma Wap65 levels to those of the corresponding transcript in the hepatopancreas, suggesting that serum Wap65 concentrations are regulated mainly by transcript levels in the hepatopancreas via the secretion process.