泛素-蛋白酶体系统参与新城疫病毒的早期复制  

THE UBIQUITIN-PROTEASOME SYSTEM PARTICIPATES IN THE EARLY STAGE OF NEWCASTLE DISEASE VIRUS REPLICATION

作　　者：曲昱蓉 詹媛[1] 仇旭升[1] 谭磊[1] 孙英杰[1] 宋翠萍[1] 孟春春[1] 丁铲[1] QU Yu-rong;ZHAN Yuan;QIU Xu-sheng;TAN Lei;SUN Ying-jie;SONG Cui-ping;MENG Chun-chun;DING Chan(Shanghai Veterinary Research Institute, CAAS, Shanghai 200241, China)

机构地区：[1]中国农业科学院上海兽医研究所,上海200241

出　　处：《中国动物传染病学报》2017年第6期1-7,共7页Chinese Journal of Animal Infectious Diseases

基　　金：国家自然科学基金(31372421;31530074);公益性行业(农业)科研专项(201303033)

摘　　要：为了研究泛素-蛋白酶体系统(ubiquitin-proteasome system,UPS)具体参与新城疫病毒(Newcastle disease virus,NDV)复制的哪个阶段,本研究通过在NDV感染不同阶段加入泛素-蛋白酶体系统抑制剂MG-132,结合使用蛋白酶K试验证明泛素-蛋白酶体系统并未参与NDV入侵细胞的过程,而是参与了NDV入侵细胞之后的复制早期阶段(0~6 h)。再利用蔗糖密度梯度离心试验分离细胞内体,使用针对内体标志蛋白Rab5的抗体确定内体位于第8~10层。建立了检测Herts/33毒株P基因的绝对荧光定量方法,绘制了标准曲线。并通过该绝对荧光定量法比较MG-132和DMSO处理的细胞内体中病毒含量,发现使用抑制剂处理的细胞内体所在层中病毒含量较高,提示蛋白酶体抑制剂可以使NDV滞留于细胞内体中。本研究首次证明泛素-蛋白酶体系统参与了NDV进入细胞后从内体释放出来的过程。The proteasome system inhibitor MG-132together with the protease K were used to study the role of ubiquitin-proteasome system(UPS)in the specific replication stage of Newcastle disease virus(NDV)in host cells.The results showed that the ubiquitinproteasome system did not participate in the NDV invasion stage but was involved in the early stage of replication(0-6h)after the invasion of the cells.The intracellular endosomes were isolated at8-10layers using sucrose density gradient centrifugation assay and confirmed using antibodies against the endosome marker protein Rab5.The absolute fluorescence quantitative method for detecting the P gene of Herts/33strain was developed to detect the viruses in endosomes from MG-132treated cells and DMSO treated cells.It concluded that the MG-132treated cells had higher viral content in the endosomes,suggesting that ubiquitin-proteasome system inhibited NDV release from endosomes.

关 键 词：泛素-蛋白酶体系统 新城疫病毒 MG-132 内体 

分 类 号：S852.659.5[农业科学—基础兽医学]