家蚕凋亡蛋白抑制因子(BmIAP)基因真核表达与功能分析  

Eukaryotic expression and function analysis of Bombyx mori inhibitor of apoptosis protein(BmIAP) gene

作　　者：易华山[1,2] 马鲜平[1] 潘敏慧 鲁成 

机构地区：[1]西南大学荣昌校区动物医学系,重庆402460 [2]家蚕基因组生物学国家重点实验室,重庆400715

出　　处：《昆虫学报》2017年第1期18-23,共6页Acta Entomologica Sinica

基　　金：西南大学博士基金(SWU114062);中央高校基本业务费专项资金(XDJK2015C032)

摘　　要：【目的】家蚕Bombyx mori凋亡蛋白抑制因子(BmIAP)是在家蚕中发现的一个凋亡蛋白抑制因子(IAP)。本研究旨在验证家蚕BmIAP蛋白在家蚕细胞内的功能特征,以进一步研究BmIAP在家蚕细胞凋亡中的作用。【方法】对构建的瞬时表达载体PIZ/V5-BmIAP-dsRed,应用脂质体转染家蚕Bm N-SWU1细胞;应用实时荧光定量PCR和Western blot分析BmIAP mRNA及蛋白表达水平;通过150 ng/m L放线菌素D诱导家蚕Bm N-SWU1细胞12,18和24 h,应用免疫荧光及实时荧光定量PCR方法,分析BmIAP瞬时表达与细胞凋亡的关系。【结果】构建的瞬时表达载体PIZ/V5-BmIAP-dsRed能在家蚕Bm N-SWU1细胞表达BmIAP蛋白,其BmIAP mRNA表达水平上调近45倍,融合蛋白约为60 k D;BmIAP基因瞬时表达72 h后,BmIAP能显著抑制放线菌素D诱导12 h时的家蚕Bm N-SWU1细胞凋亡。【结论】BmIAP蛋白在家蚕细胞中能抑制家蚕细胞凋亡,是家蚕的一种凋亡蛋白抑制因子。【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 Bm N-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 Bm N-SWU1 cells were induced by 150 ng / m L 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 k D,and the mRNA expression level of BmIAP was up-regulated 45-fold in Bm N-SWU1 cells of B. mori. After BmIAP was transiently expressed for 72 h using PIZ / V5-BmIAPdsRed vector in the Bm N-SWU1 cells,BmIAP significantly inhibited the apoptosis in the Bm N-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.

关 键 词：家蚕 细胞凋亡 凋亡蛋白抑制因子 真核表达 瞬时表达 

分 类 号：Q966[生物学—昆虫学]