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Molecular cloning, characterization and expression of cathepsin D from grass carp (Ctenopharyngodon idella)

Dong, Zhong-dian, Zhang, Jiao, Ji, Xiang-shan, Zhou, Fen-na, Fu, Yong, Chen, Weiyun, Zeng, Yong-qing, Li, Tong-ming, Wang, Hui
Fish & shellfish immunology 2012 v.33 no.5 pp. 1207-1214
Aeromonas, Ctenopharyngodon idella, Danio rerio, amino acids, bacterial infections, bladder, brain, cathepsin D, complementary DNA, embryogenesis, eyes, genes, heart, innate immunity, intestines, kidneys, liver, molecular cloning, muscles, open reading frames, rapid amplification of cDNA ends, reverse transcriptase polymerase chain reaction, spleen, tissues
Cathepsin D is a lysosomal aspartic proteinase which participates in various degradation functions within the cell. In this current study, we cloned and characterized the complete cDNA of grass carp cathepsin D through 5′- and 3′-RACE. The cathepsin D contained a 56 bp 5′ terminal untranslated region (5′-UTR), a 1197 bp open reading frame encoding 398 amino acids, and a 394 bp 3′-UTR. Grass carp cathepsin D shared high similarity with those from other species, and showed the highest amino acid identity of 91% to Danio rerio. Unlike many other organisms, the grass carp cathepsin D contains only one N-glycosylation site closest to the N-terminal. Real-time quantitative RT-PCR demonstrated that Cathepsin D expressed in all twelve tissues (bladder, brain, liver, heart, gill, muscle, fin, eye, intestines, spleen, gonad and head kidney). The relative expression levels of Cathepsin D in gonad and liver were 26.58 and 24.95 times as much as those in fin, respectively. The expression level of Cathepsin D in muscle approximately 16-fold higher, in intestines and spleen were 12-fold higher. The cathepsin D expression showed an upward trend during embryonic development. After challenged with Aeromonas hydrophil, the expression of grass carp cathepsin D gene showed significant changes in the four test tissues (liver, head kidney, spleen and intestines). The fact that the bacterial infection can obviously improve the cathepsin D expression in immune-related organs, may suggest that cathepsin D plays an important role in the innate immune response of grass carp.