泛素-蛋白酶体系统参与新城疫病毒的早期复制

发布时间：2018-05-05 02:38

本文选题：泛素-蛋白酶体系统 + 新城疫病毒　；参考：《中国动物传染病学报》2017年06期

【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。
[Abstract]:In order to study which stage of ubiquitin proteasome system UPSs is involved in the replication of Newcastle disease virus (NDV), In this study, Ubiquitin proteasome system inhibitor MG-132 was added at different stages of NDV infection. Combined with protease K test, it was proved that ubiquitin proteasome system was not involved in the process of NDV invasion. It was involved in the early stage of replication after NDV invasion. Then sucrose density gradient centrifugation was used to isolate the intracellular body and the antibody against the inner marker protein Rab5 was used to determine that the inner body was located in layer 810. An absolute fluorescence quantitative method for the detection of P gene in Herts/33 strain was established and the standard curve was drawn. The virus content in the intracellular body treated with MG-132 and DMSO was compared by the absolute fluorescence quantitative method. The results showed that the virus content in the layer of the intracellular body treated with inhibitor was higher, suggesting that proteasome inhibitor could keep NDV in the intracellular body. This study demonstrated for the first time that the ubiquitin-proteasome system was involved in the release of NDV from the inner body after entering the cell.
【作者单位】：中国农业科学院上海兽医研究所;
【基金】：国家自然科学基金(31372421、31530074) 公益性行业(农业)科研专项(201303033)
【分类号】：S852.65

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