FUSION OF INTRACELLULAR AND EXTRACELLULAR FORMS OF VACCINIA VIRUS WITH THE CELL-MEMBRANE
FUSION OF INTRACELLULAR AND EXTRACELLULAR FORMS OF VACCINIA VIRUS WITH THE CELL-MEMBRANE
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DOI：
10.1128/jvi.64.10.4884-4892.1990
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发表时间：
1990-10-01
影响因子：
5.4
通讯作者：
MOSS, B
中科院分区：
文献类型：
--
作者：
DOMS, RW;BLUMENTHAL, R;MOSS, B
采用基于荧光探针稀释的脂质混合实验研究了分离的单包膜牛痘病毒（INV）细胞内型和双包膜细胞外型（EEV）的膜融合活性。荧光标记的牛痘病毒IHD-J和WR株的INV和EEV在中性pH下与HeLa细胞融合，表明病毒进入时与质膜发生融合。与INV相比，EEV的熔合效率更高，动力学速度更快：在1小时内，约50%的结合EEV颗粒融合，而INV颗粒仅为25%。INV和EEV的融合强烈依赖于温度，在34度时降低50%。在28度时减少90%。C.与融合反应有关的牛痘病毒14千道尔顿包膜蛋白的单克隆抗体（J. F. Rodriguez， E. Paez, and M. Esteban， J. Virol. 61:395- 404,1987）完全抑制了INV的初始融合率，但对EEV的融合活性没有影响，这表明牛痘病毒编码两种或更多的膜融合蛋白。最后，感染牛痘病毒WR株的细胞在pH 6.4或以下短暂孵育后形成合胞体，表明酸激活的病毒融合蛋白在细胞表面表达。然而，WR INV和EEV在酸性pH下没有显示出增加的融合活性，这表明酸依赖性融合因子没有被纳入病毒粒子或其在酸性pH下的活性被掩盖。
The membrane fusion activities of the isolated single-enveloped intracellular form of vaccinia virus (INV) and the double-enveloped extracellular (EEV) form were studied by using a lipid-mixing assay based on the dilution of a fluorescent probe. Fluorescently labeled INV and EEV from both the IHD-J and WR strains of vaccinia virus fused with HeLa cells at neutral pH, suggesting that fusion occurs with the plasma membrane during virus entry. EEV fused more efficiently and with faster kinetics than INV: approximately 50% of bound EEV particles fused over the course of 1 h, compared with only 25% of the INV particles. Fusion of INV and EEV was strongly temperature dependent, being decreased by 50% at 34.degree. C and by 90% at 28.degree. C. A monoclonal antibody to a 14-kilodalton envelope protein of INV that has been implicated in the fusion reaction (J. F. Rodriguez, E. Paez, and M. Esteban, J. Virol. 61:395-404, 1987) completely suppressed the initial rate of fusion of INV but had no effect on the fusion activity of EEV, suggesting that vaccinia virus encodes two or more membrane fusion proteins. Finally, cells infected with the WR strain of vaccinia virus formed syncytia when briefly incubated at pH 6.4 or below, indicating that an acid-activated viral fusion protein is expressed on the cell surface. However, WR INV and EEV did not display increased fusion activity at acid pH, suggesting that the acid-dependent fusion factor is not incorporated into virions or that its activity there is masked.