Molecular cloning, characterization and expression analysis of caspase-6 in puffer fish (Takifugu obscurus)

Caspases are a family of proteases involved in many important biological processes, including inflammation, cell differentiation and apoptosis. Among them, caspase-6 plays an important role in the execution phase of the apoptotic death cascade. In this study, the full-length of puffer fish caspase - 6 ( Pfcasp - 6 ) was cloned and characterized, containing an open reading frame of 912 bp, a 5′-UTR of 168 bp and a 3′-UTR of 632 bp with a poly (A) tail. The Pfcasp - 6 encoded a polypeptide of 303 amino acids with an estimated molecular mass of 34.09 kDa and a predicted isoelectric point of 5.35. Phylogenetic analysis indicated that Pfcasp-6 was highly homology to that of other teleostean species. Spatial mRNA expression analysis revealed that the ubiquitous expression of Pfcasp - 6 in all examined tissues, with high expression in skin, followed by gill, intestine, kidney , blood, heart and spleen. After the challenges of high temperatures and Aeromonas hydrophila , the transcript expressions of Pfcasp - 6 were significantly up-regulated in blood and spleen. In addition, cellular localization analysis indicated that the distribution of Pfcasp-6 was in the cytoplasm and nucleus. Overexpression of Pfcasp-6 was able to trigger a significant level of cell death in HeLa cells, suggesting its putative role in cell apoptosis. Taken together, the results of this study suggested that Pfcasp - 6 might be an important component in the caspases cascade, and was likely to get involved in the response to different stress in puffer fish.