Characterization of cathepsin B gene from orange-spotted grouper, Epinephelus coioides involved in SGIV infection.
The lysosomal cysteine protease cathepsin B of papain family is a key regulator and signaling molecule that involves in various biological processes, such as the regulation of apoptosis and activation of virus. In the present study, cathepsin B gene (Ec-CB) was cloned and characterized from orange-spotted grouper, Epinephelus coioides. The full-length Ec-CB cDNA was composed of 1918 bp and encoded a polypeptide of 330 amino acids with higher identities to cathepsin B of teleosts and mammalians. Ec-CB possessed typical cathepsin B structural features including an N-terminal signal peptide, the propeptide region and the cysteine protease domain which were conserved in other cathepsin B sequences. Phylogenetic analysis revealed that Ec-CB was most closely related to Lutjanus argentimaculatus. RT-PCR analysis showed that Ec-CB transcript was expressed in all the examined tissues which abundant in spleen, kidney and gill. After challenged with Singapore grouper iridovirus (SGIV) stimulation, the mRNA expression of cathepsin B in E. coioides was up-regulated at 24 h post-infection. Subcellular localization analysis revealed that Ec-CB was distributed predominantly in the cytoplasm. When the fish cells (GS or FHM) were treated with the cathepsin B specific inhibitor CA-074Me, the occurrence of CPE induced by SGIV was delayed, and the viral gene transcription was significantly inhibited. Additionally, SGIV-induced typical apoptosis was also inhibited by CA-074Me in FHM cells. Taken together, our results demonstrated that the Ec-CB might play a functional role in SGIV infection.
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Shina
S
Wei
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Youhua
Y
Huang
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Xiaohong
X
Huang
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China.
Jia
J
Cai
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Yang
Y
Yan
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China.
Chuanyu
C
Guo
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China. Electronic address: qinqw@scsio.ac.cn.
Qiwei
Q
Qin
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology*; D003001:Cloning, Molecular; D003588:Cytopathogenic Effect, Viral / Q000276:immunology; D004266:DNA Virus Infections / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D004151:Dipeptides / Q000494:pharmacology; D005393:Fish Diseases / Q000276:immunology / Q000821:virology*; D017994:Iridovirus / Q000276:immunology*; D008969:Molecular Sequence Data; D010473:Perciformes; D010802:Phylogeny; D012313:RNA / Q000737:chemistry / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique / Q000662:veterinary; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic / Q000276:immunology
C400541:CA 074 methyl ester; D004151:Dipeptides; D012313:RNA; D002401:Cathepsin B
Apoptosis; Cathepsin B; Epinephelus coioides; Singapore grouper iridovirus (SGIV); Virus replication