Molecular cloning, functional characterization and phylogenetic analysis of TRAIL in Japanese pufferfish Takifugu rubripes

• Published in: Journal of fish biology 2011-09, Vol.79 (3), p.747-760
• Main Authors: Li, J. F., Ai, H. X., Zhang, J., Du, M. X., Zhang, Z., Zhang, J. X., Zhang, S. Q.
• Format: Article
• Language: English
• Subjects: 3D structure; Amino Acid Sequence; Animals; apoptosis; Base Sequence; cDNA; Cloning, Molecular; Escherichia coli; Fish Proteins - genetics; Fish Proteins - isolation & purification; Fish Proteins - metabolism; Gene Components; HeLa Cells; Humans; Marine; Molecular Sequence Data; Phylogeny; Protein Structure, Tertiary; RNA, Messenger - metabolism; Sequence Alignment; Sequence Analysis, DNA; Sequence Homology, Amino Acid; SUMO; Takifugu - genetics; Takifugu - metabolism; Takifugu rubripes; Teleostei; TNF-Related Apoptosis-Inducing Ligand - genetics; TNF-Related Apoptosis-Inducing Ligand - isolation & purification; TNF-Related Apoptosis-Inducing Ligand - metabolism
• ISSN: 0022-1112; 1095-8649
• DOI: 10.1111/j.1095-8649.2011.03058.x

In this study, the complementary DNA (cDNA) of Japanese pufferfish Takifugu rubripes tumour necrosis factor‐related apoptosis‐inducing ligand (TRAIL) was cloned by reverse‐transcription PCR. The open reading frame of the TRAIL consisted of 870 bases. The deduced amino‐acid sequence of the TRAIL showed a high homology with the sequences of other teleosts. Recombinant soluble TRAIL was fused with a small ubiquitin‐related modifier gene to enhance the soluble expression level in Escherichia coli BL21 (DE3). In vitro, the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrasodium bromide (MTT) assay indicated that the purified soluble TRAIL was able to induce apoptosis of Jurkat and HeLa cells in a dose‐dependent manner.