Wu X, Liu H, Xiao H, Conway J A, Hunter E, Kappes J C
Department of Medicine, University of Alabama at Birmingham, 35294, USA.
EMBO J. 1997 Aug 15;16(16):5113-22. doi: 10.1093/emboj/16.16.5113.
The expression and incorporation of retroviral enzymes into virions in the form of Gag/Pol precursor polyproteins is believed to be important for the assembly of infectious viral particles. HIV-1 encodes a 160 kDa Gag/Pol precursor that includes Gag, protease (PR), reverse transcriptase (RT) and integrase (IN). We have developed the use of HIV accessory proteins (Vpr and Vpx) as vehicles to incorporate protein of both viral and non-viral origin into virions by expression in trans as heterologous fusion proteins (Wu et al., 1995, 1996a). To analyze the role of Gag/Pol in the formation of infectious virions, we incorporated RT and IN into HIV-1 particles in trans, as fusion partners of viral protein R (Vpr). Virions derived from an RT and IN minus proviral clone were infectious and replicated through a complete cycle of infection when RT and IN proteins were provided in trans. These results demonstrate that functional RT and IN proteins can be provided in trans, and that their expression and incorporation into virions as components of Gag/Pol are not required for the formation of infectious virions. Thus, for the first time, we have demonstrated for a human pathogenic retrovirus that processes of assembly and the function of critical viral enzymes can be unlinked. This finding will provide unique opportunities to explore retroviral RT/IN function and the role of Gag/Pol in the formation of infectious virions in the context of a replicating virus (in vivo).
逆转录病毒酶以Gag/Pol前体多聚蛋白的形式表达并整合到病毒粒子中，这被认为对传染性病毒颗粒的组装很重要。HIV-1编码一种160 kDa的Gag/Pol前体，其中包括Gag、蛋白酶（PR）、逆转录酶（RT）和整合酶（IN）。我们已开发利用HIV辅助蛋白（Vpr和Vpx）作为载体，通过作为异源融合蛋白的反式表达将病毒和非病毒来源的蛋白整合到病毒粒子中（Wu等人，1995年，1996a）。为了分析Gag/Pol在传染性病毒粒子形成中的作用，我们将RT和IN作为病毒蛋白R（Vpr）的融合伙伴反式整合到HIV-1颗粒中。当反式提供RT和IN蛋白时，源自RT和IN缺失前病毒克隆的病毒粒子具有传染性，并能通过完整的感染周期进行复制。这些结果表明，功能性RT和IN蛋白可以反式提供，并且它们作为Gag/Pol的组成部分表达并整合到病毒粒子中对于传染性病毒粒子的形成并非必需。因此，我们首次证明了对于一种人类致病性逆转录病毒，组装过程和关键病毒酶的功能可以分开。这一发现将为在复制病毒（体内）的背景下探索逆转录病毒RT/IN功能以及Gag/Pol在传染性病毒粒子形成中的作用提供独特的机会。
