Mutations in the GM1 Binding Site of Simian Virus 40 VP1 Alter Receptor Usage and Cell Tropism
Mutations in the GM1 Binding Site of Simian Virus 40 VP1 Alter Receptor Usage and Cell Tropism
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DOI：
10.1128/jvi.00371-12
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发表时间：
2012-07-01
影响因子：
5.4
通讯作者：
DiMaio, Daniel
中科院分区：
文献类型：
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作者：
Magaldi, Thomas G.;Buch, Michael H. C.;DiMaio, Daniel
多瘤病毒是无包膜病毒，衣壳主要由病毒 VP1 蛋白的 72 个五聚体组成，形成衣壳的外壳并与细胞表面寡糖受体结合。来自密切相关的多瘤病毒的高度保守的 VP1 蛋白可识别不同的寡糖。为了确定仅限于寡糖结合位点的氨基酸变化是否足以确定受体特异性以及受体使用的变化如何影响向性，我们研究了灵长类多瘤病毒猿病毒 40 (SV40)，它使用神经节苷脂 GM1 作为介导细胞结合和进入的受体。在这里，我们使用两次连续的遗传筛选来分离和表征在 VP1 GM1 结合位点中具有突变的可行 SV40 突变体。其中两个突变体完全抵抗 GM1 中和，不再受到 GM1 掺入细胞膜的刺激，并且无法与细胞表面的 GM1 结合。此外，这些突变病毒在细胞表面GM1水平较高的猴细胞中表现出感染缺陷。有趣的是，一种突变体感染具有低细胞表面GM的细胞比野生型病毒更有效，显然是通过利用不同的神经节苷脂受体。我们的结果表明，GM1 结合位点的少量突变足以改变神经节苷脂的使用并改变向性，并且他们表明 VP1 分歧主要是由适应特定受体的要求驱动的。此外，我们的结果表明，GM1 结合是允许猴 CV-1 细胞中液泡形成所必需的。对这些突变体的进一步研究将为多瘤病毒的进入、发病机制和进化提供新的见解。
Polyomaviruses are nonenveloped viruses with capsids composed primarily of 72 pentamers of the viral VP1 protein, which forms the outer shell of the capsid and binds to cell surface oligosaccharide receptors. Highly conserved VP1 proteins from closely related polyomaviruses recognize different oligosaccharides. To determine whether amino acid changes restricted to the oligosaccharide binding site are sufficient to determine receptor specificity and how changes in receptor usage affect tropism, we studied the primate polyomavirus simian virus 40 (SV40), which uses the ganglioside GM1 as a receptor that mediates cell binding and entry. Here, we used two sequential genetic screens to isolate and characterize viable SV40 mutants with mutations in the VP1 GM1 binding site. Two of these mutants were completely resistant to GM1 neutralization, were no longer stimulated by incorporation of GM1 into cell membranes, and were unable to bind to GM1 on the cell surface. In addition, these mutant viruses displayed an infection defect in monkey cells with high levels of cell surface GM1. Interestingly, one mutant infected cells with low cell surface GM] more efficiently than wild-type virus, apparently by utilizing a different ganglioside receptor. Our results indicate that a small number of mutations in the GM1 binding site are sufficient to alter ganglioside usage and change tropism, and they suggest that VP1 divergence is driven primarily by a requirement to accommodate specific receptors. In addition, our results suggest that GM1 binding is required for vacuole formation in permissive monkey CV-1 cells. Further study of these mutants will provide new insight into polyomavirus entry, pathogenesis, and evolution.