Cloning, eukaryotic expressing and function analysis of Schistosoma japonicum apoptosis gene Sjcaspase3.

Tao Wang; Yang Hong; Hongxiao Han; Chao LV; Bingguang Jia; Xiaodan Cao; Qian Han; Ke LU; Hao LI; Zhiqiang FU; Jiaojiao Lin

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Cloning, eukaryotic expressing and function analysis of Schistosoma japonicum apoptosis gene Sjcaspase3.

Author: Tao WANG ¹ ; Yang HONG ¹ ; Hongxiao HAN ¹ ; Chao LV ¹ ; Bingguang JIA ¹ ; Xiaodan CAO ¹ ; Qian HAN ¹ ; Ke LU ¹ ; Hao LI ¹ ; Zhiqiang FU ¹ ; Jiaojiao LIN ¹
Author Information

1. Key Laboratory of Animal Parasitology, Ministry of Agriculture, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Science, Shanghai 200241, China.
Publication Type:Journal Article
Keywords: Schistosoma japonicum; apoptosis; caspase3; enzyme activity; flow cytometry
MeSH: Animals; Apoptosis; Blotting, Western; Caspase 3; genetics; metabolism; Cloning, Molecular; DNA, Complementary; Female; HeLa Cells; Helminth Proteins; genetics; metabolism; Humans; Male; Real-Time Polymerase Chain Reaction; Recombinant Proteins; Schistosoma japonicum
From: Chinese Journal of Biotechnology 2016;32(7):889-900
CountryChina
Language:Chinese
Abstract: For further research of the apoptosis mechanism of Schistosoma japonicum (S. japonicum). The cDNA encoding Sjcaspase3 of Schistosoma japonicum was amplified by polymerase chain reaction (PCR) technique, which contained 900 nucleotides and encoded 299 amino acids. The theory molecular weight and isoelectric point (PI) of the deduced protein is 33.5 kDa and 6.39, respectively. Real-time PCR was used to analyze the transcription profiles of Sjcaspase3 at different development stages of S. japonicum. The results showed that this gene was expressed in all stages of S. japonicum with the highest expression in 21d worms, and the level of gene transcription in 42 d female worms was higher than that of male worms. The recombinant plasmid pXJ40-FLAG-Sjcaspase3 was constructed and transfection into Hela cells successfully. Real-time PCR and Western blotting analysis showed Sjcaspase3 was successfully expressed in Hela cells. Enzyme activity analysis revealed that recombinant Sjcaspase3 possessed the activity to cut substrate DEVD. Flow cytometry proved that Sjcaspase3 could induce early apoptosis of Hela cells. The results provide the basis for proceeding further study on the biological function of Sjcaspase3 and better understand the apoptosis mechanism of S. japonicum.