Eukaryotic expression and function analysis of Bombyx mori inhibitor of apoptosis protein (BmIAP) gene
(2.State Key Laboratory of Silkworm Genome Biology, Rongchang, Chongqing, China 400715)
【Abstract】[Aim] Bombyx mori inhibitor of apoptosis protein (BmIAP) has been found in the silkworm, B. mori, and characterized as a protein of the IAP (inhibitor of apoptosis protein) family. This study aims to verify its function as a suppressor of apoptosis in B. mori cells, so as to further study the roles of BmIAP in cell apoptosis. [Methods] The transient expression vector named PIZ/V5-BmIAP-dsRed was transfected in the BmN-SWU1 cells of B. mori by liposome, and the mRNA and protein expression levels of BmIAP gene were evaluated by real-time quantitative PCR and Western blot analysis, respectively. The BmN-SWU1 cells were induced by 150 ng/mL actinomycin D for 12, 18 and 24 h, and the relationship between BmIAP protein and cell apoptosis was assayed by immunofluorescence and real-time quantitative PCR methods. [Results] The PIZ/V5-BmIAP-dsRed vector effectively expressed BmIAP fusion protein of about 60 kD, and the mRNA expression level of BmIAP was up-regulated by 45 folds in BmN-SWU1 cells of B. mori. After BmIAP was transiently expressed for 72 h using PIZ/V5-BmIAP-dsRed vector in the BmN-SWU1 cells, BmIAP significantly inhibited the apoptosis in the BmN-SWU1 cells at 12 h after induction by actinomycin D. [Conclusion] BmIAP can inhibit apoptosis in B. mori cells and is one of the inhibitor of apoptosis proteins in B. mori.
【Keywords】 Bombyx mori; apoptosis; inhibitor of apoptosis protein (IAP) ; eukaryotic expression; transient expression;
(Translated by 陈彤)
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