Ability of small animal cells to support the postintegration phase of human immunodeficiency virus type-1 replication
Ability of small animal cells to support the postintegration phase of human immunodeficiency virus type-1 replication
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DOI：
10.1006/viro.2002.1755
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发表时间：
2003-01-05
期刊：
影响因子：
3.7
通讯作者：
Matsushita, S
中科院分区：
文献类型：
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作者：
Koito, A;Shigekane, H;Matsushita, S
我们研究了来自多种组织的广泛的小动物细胞，包括啮齿动物、水貂和鸟类细胞，以支持HIV-1复制的整合后步骤的潜力。这些细胞被工程化以支持人类周期蛋白T1的稳定表达，并进一步被HIV-1 gag和pol基因转导。病毒基因表达被人类周期蛋白T1激活，但除水貂细胞外，在人类细胞中没有见过这种水平。此外，p24 CA释放存在相当大的缺陷，特别是在啮齿动物细胞中。蔗糖浮法分离Gag蛋白发现，人细胞中的Gag转运到膜组分，并被有效地处理为p24 CA和p17 MA。共聚焦成像显示Gag在这些细胞的质膜和细胞膜反式高尔基池中以点状模式定位。相比之下，啮齿动物细胞中的大多数Gag主要存在于细胞质复合物中，并且未被加工。用[9,10(n)-H-3]肉豆蔻酸标记显示，在啮齿动物和人类细胞中，n -肉豆蔻酰化Pr55（gag）的程度相似，这表明尽管gag的n -肉豆蔻酰化是膜结合所必需的，但它不足以赋予膜靶向特异性。值得注意的是，尽管细胞内Gag加工水平降低，水貂Mv.1。在支持病毒粒子组装和释放方面，Lu细胞与人类细胞似乎没有显著差异。对互易异核体的分析表明，在小鼠细胞中缺乏有效组装和释放感染性病毒粒子所需的细胞因子，但在水貂和人类细胞中似乎都存在功能。我们的研究结果证实并扩展了先前关于HIV在非人类细胞中复制的多重阻滞的报道，这些阻滞在小鼠细胞中最为深刻。他们还提出了一种可能性，即其他小动物，如水貂，可以作为研究HIV-1感染和疾病的新模型系统。(C) 2002 Elsevier Science （USA）。
We examine the potential for a broad range of small animal cells, including rodent, mink, and avian cells, from multiple tissues to support postintegration steps of HIV-1 replication. These cells were engineered so as to support a stable expression of human cyclin T1 and were further transduced with HIV-1 gag and pol genes. Viral gene expression was activated by the presence of human cyclin T1, but, with the exception of mink cells, was not at the level seen in human cells. Furthermore, there were considerable defects in p24 CA release, in particular in the case of rodent cells. Fractionation of Gag proteins by sucrose floatation revealed that the Gag in human cells trafficked to membrane fractions and was processed to p24 CA and p17 MA efficiently. Confocal imaging demonstrated that Gag was localized in a punctate pattern at the plasma membrane as well as intracellular membrane trans-Golgi cisternae in these cells. In contrast, the majority of Gag in rodent cells was largely present in cytosolic complexes and remained unprocessed. Labeling with [9,10(n)-H-3]myristic acid showed a similar degree of N-myristoylated Pr55(gag) in rodent and human cells, indicating that while N-myristoylation of Gag was essential for membrane binding, it was not sufficient to confer membrane targeting specificity. Remarkably, despite the reduced level of intracellular Gag processing, mink Mv.1.Lu cells did not appear to differ significantly from human cells in support of virion assembly and release. Analysis of reciprocal heterokaryons suggested that the cellular factor(s) required for efficient assembly and release of infectious virions is lacking in murine cells but appears to be functionally present in mink as well as human cells. Our findings confirm and extend previous reports of multiple blocks to HIV replication in nonhuman cells that are most profound in murine cells. They also raise the possibility that other small animals, such as mink, could serve as novel model systems for studying HIV-1 infection and disease. (C) 2002 Elsevier Science (USA).