Poon D T, Li G, Aldovini A
Department of Medicine, Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
J Virol. 1998 Mar;72(3):1983-93. doi: 10.1128/JVI.72.3.1983-1993.1998.
The nucleocapsid protein (NC) of retroviruses plays a major role in genomic RNA packaging, and some evidence has implicated the matrix protein (MA) of certain retroviruses in viral RNA binding. To further investigate the role of NC in the selective recognition of genomic viral RNA and to address the potential contribution of MA in this process, we constructed chimeric and deletion human immunodeficiency virus type 1 (HIV-1) mutants that alter the NC or MA protein. Both HIV and mouse mammary tumor virus (MMTV) NC proteins have two zinc-binding domains and similar basic amino acid compositions but differ substantially in total length, amino acid sequence, and spacing of the zinc-binding motifs. When the entire NC coding sequence of HIV was replaced with the MMTV NC coding sequence, we found that the HIV genome was incorporated into virions at 50% of wild-type levels. Viruses produced from chimeric HIV genomes with complete NC replacements, or with the two NC zinc-binding domains replaced with MMTV sequences, preferentially incorporated HIV genomes when both HIV and MMTV genomes were simultaneously present in the cell. Viruses produced from chimeric MMTV genomes in which the MMTV NC had been replaced with HIV NC preferentially incorporated MMTV genomes when both HIV and MMTV genomes were simultaneously present in the cell. In contrast, viruses produced from chimeric HIV genomes containing the Moloney NC, which contains a single zinc-binding motif, were previously shown to preferentially incorporate Moloney genomic RNA. Taken together, these results indicate that an NC protein with two zinc-binding motifs is required for specific HIV RNA packaging and that the amino acid context of these motifs, while contributing to the process, is less crucial for specificity. The data also suggest that HIV NC may not be the exclusive determinant of RNA selectivity. Analysis of an HIV MA mutant revealed that specific RNA packaging does not require MA protein.
逆转录病毒的核衣壳蛋白（NC）在基因组RNA包装中起主要作用，并且一些证据表明某些逆转录病毒的基质蛋白（MA）参与病毒RNA结合。为了进一步研究NC在基因组病毒RNA选择性识别中的作用，并探讨MA在此过程中的潜在贡献，我们构建了改变NC或MA蛋白的嵌合和缺失型1型人类免疫缺陷病毒（HIV-1）突变体。HIV和小鼠乳腺肿瘤病毒（MMTV）的NC蛋白都有两个锌结合结构域和相似的碱性氨基酸组成，但在全长、氨基酸序列和锌结合基序的间距上有很大差异。当HIV的整个NC编码序列被MMTV的NC编码序列取代时，我们发现HIV基因组以野生型水平50%的比例整合到病毒颗粒中。当HIV和MMTV基因组同时存在于细胞中时, 由完全替换NC或用MMTV序列替换两个NC锌结合结构域的嵌合HIV基因组产生的病毒优先整合HIV基因组。当HIV和MMTV基因组同时存在于细胞中时，由MMTV NC被HIV NC取代的嵌合MMTV基因组产生的病毒优先整合MMTV基因组。相比之下，由含有单个锌结合基序的莫洛尼氏鼠白血病病毒（Moloney）NC的嵌合HIV基因组产生的病毒先前已被证明优先整合莫洛尼氏基因组RNA。综上所述，这些结果表明，具有两个锌结合基序的NC蛋白是HIV特异性RNA包装所必需的，并且这些基序的氨基酸背景虽然有助于这一过程，但对特异性的影响较小。数据还表明，HIV NC可能不是RNA选择性的唯一决定因素。对HIV MA突变体的分析表明，特异性RNA包装不需要MA蛋白。
