Molecular cloning and characterization of the cathepsin L gene in Pelodiscus sinensis and its expression in response to bacterial challenge

Cathepsin L is one of the most important lysosomal cysteine proteases for the initiation of protein degradation, and it is involved in the immune response in many vertebrates. However, its function in the innate immune system of turtles remains poorly understood. Here, we cloned the cathepsin L gene from the Chinese soft‐shelled turtle Pelodiscus sinensis. We then examined the mRNA expression in different tissues and at different time points after infection by Aeromonas hydrophila or Vibrio parahemolyticus. The full‐length cDNA sequence cloned from P. sinensis was 1,805 bp with a 1,071 bp open reading frame, which encodes a 40.39 kDa polypeptide of 356 amino acids. The deduced protein sequence contained an active triad of Cys, His and Asn, and conserved ERWNIN and GNFD motifs. Homology analysis showed that the deduced amino acid sequence shared 77%–96% identity with other known species. Sequence alignment, phylogenetic analysis and structural comparisons revealed that cathepsin L is a member of the cathepsin family. Real‐time PCR analyses showed that turtle cathepsin L mRNA is ubiquitously expressed in various tissues, and the expression level is higher in the liver than in other tissues. The expression level in the liver peaks 24 hr after A. hydrophila infection and at two times (12 and 72 hr) after V. parahemolyticus infection. These results suggest that the cathepsin L gene of P. sinensis is likely involved in the process of the antibacterial response to A. hydrophila and V. parahemolyticus. This study is of great importance for future work exploring the molecular mechanism of antibacterial immune responses in P. sinensis.