Human immunodeficiency virus type 1 Nef protein is incorporated into virus particles and specifically cleaved by the viral proteinase
Human immunodeficiency virus type 1 Nef protein is incorporated into virus particles and specifically cleaved by the viral proteinase
复制标题
DOI：
10.1006/viro.1996.0240
复制
发表时间：
1996-05-01
期刊：
影响因子：
3.7
通讯作者：
Krausslich, HG
中科院分区：
文献类型：
--
作者：
Welker, R;Kotler, H;Krausslich, HG
灵长类免疫缺陷病毒的Nef蛋白对于在体内建立高产致病性感染是必不可少的。在组织培养中，Nef不是感染所必需的，但可以增强病毒的感染性。这种效应在未受刺激的原代淋巴细胞中最明显，并发生在病毒基因表达之前的感染早期。由于Nef表达不会导致病毒组成的明显变化，因此分析Nef是否掺入病毒颗粒是有意义的。在这里，我们表明，Nef是特异性免疫沉淀从放射性标记的人类免疫缺陷病毒1型（HIV-1）感染的细胞和病毒颗粒制剂。定量分析显示Nef以逆转录酶掺入的10%的数量级掺入，这对应于每个病毒体5至10个Nef分子。在感染的细胞中，Nef被检测为全长27-kDa蛋白。相比之下，大约50%的颗粒相关Nef对应于一个18 kDa的物种，其与较大的产品后，在体外裂解纯化的HIV-1 Nef的病毒蛋白酶。Nef裂解颗粒制剂中完全废除了HIV-1蛋白酶的特异性抑制剂。最有可能的是，Nef在病毒颗粒成熟时与病毒结构蛋白同时裂解。这种切割可能在功能上是重要的，因为它将保守的核心结构域从N-末端膜附着区解离。我们的研究结果表明，在组织培养和体内建立未刺激的细胞感染的Nef的深远影响是由病毒相关的Nef介导的，它在病毒基因表达前的早期感染中发挥作用。(C)出版社：Academic Press，Inc.
The Nef protein of primate immunodeficiency viruses is essential for establishing a highly productive pathogenic infection in vivo. In tissue culture, Nef is not required for infection but enhances viral infectivity. This effect is most pronounced in unstimulated primary lymphocytes and occurs in the early phase of infection prior to viral gene expression. Since Nef expression does not lead to obvious changes in virus composition, it was of interest to analyze whether Nef is incorporated into virus particles. Here, we show that Nef is specifically immunoprecipitated from radioactively labeled human immunodeficiency virus type 1 (HIV-1)-infected cells and virus particle preparations. Quantitative analysis revealed Nef to be incorporated on the order of 10% of reverse transcriptase incorporation, which corresponds to 5 to 10 molecules of Nef per virion. In infected cells, Nef was detected as a full-length 27-kDa protein. In contrast, approximately 50% of particle-associated Nef corresponded to an 18-kDa species which comigrated with the larger product after in vitro cleavage of purified HIV-1 Nef by the viral proteinase. Nef cleavage in particle preparations was completely abolished by a specific inhibitor of HIV-1 proteinase. Most likely, Nef is cleaved concomitantly with viral structural proteins on maturation of virus particles. This cleavage is likely to be functionally significant because it dissociates the conserved core domain from the N-terminal membrane attachment region. Our results suggest that the profound influence of Nef on establishing infection of unstimulated cells in tissue culture and in vivo is mediated by virion-associated Nef which functions in early infection before viral gene expression. (C) 1996 Academic Press, Inc.