FUSION OF INFLUENZA HEMAGGLUTININ-EXPRESSING FIBROBLASTS WITH GLYCOPHORIN-BEARING LIPOSOMES - ROLE OF HEMAGGLUTININ SURFACE-DENSITY
FUSION OF INFLUENZA HEMAGGLUTININ-EXPRESSING FIBROBLASTS WITH GLYCOPHORIN-BEARING LIPOSOMES - ROLE OF HEMAGGLUTININ SURFACE-DENSITY
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DOI：
10.1021/bi00493a027
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发表时间：
1990-10-16
期刊：
影响因子：
2.9
通讯作者：
WHITE, JM
中科院分区：
文献类型：
--
作者：
ELLENS, H;BENTZ, J;WHITE, JM
流感病毒通过病毒和宿主细胞膜之间的融合事件进入其宿主细胞的细胞质。融合是由包膜糖蛋白血凝素（HA）介导的，并引发低pH值。要了解有多少血凝素三聚体是必要的，以引起膜融合，我们已经使用了两个NIH 3 T3成纤维细胞系，表达HA蛋白在不同的表面密度。基于每个细胞的HA三聚体的数量和两种细胞系的相对表面积的定量，HAb-2细胞具有比GP 4F细胞高1.9倍的质膜表面密度。两种细胞系的膜侧向扩散系数和HA的移动的部分相同。对带有血型糖蛋白的脂质体与细胞结合的Scatchard分析表明，GP 4F细胞有1700个结合位点，HAb-2细胞有3750个结合位点，脂质体-细胞结合常数实际上相同，约为7 × 10 - 6。1010 M-1结合对脂质体上的血型糖蛋白和细胞上表达的HA具有特异性。采用含毒素和空脂质体的竞争实验使我们能够定量每个细胞融合的脂质体的数量，这是结合脂质体数量的一个小的恒定分数。对于HAb-2细胞，每70个结合的脂质体中约有1个融合，对于GP 4F细胞，每300个结合的脂质体中约有1个融合。因此，尽管HA的表面密度仅为1.9倍，但HAb-2细胞显示出每个结合脂质体的4.4倍多的融合。我们得出以下结论：（i）一个HA三聚体不足以诱导融合。(ii)与血型糖蛋白结合的HA不是诱导融合的HA。也就是说，即使每个HA具有结合和融合功能，这些功能也不由相同的HA三聚体执行。
Influenza virus gains access to the cytoplasm of its host cell by means of a fusion event between viral and host cell membrane. Fusion is mediated by the envelope glycoprotein hemagglutinin (HA) and is triggered by low pH. To learn how many hemagglutinin trimers are necessary to cause membrane fusion, we have used two NIH 3T3 fibroblast cell lines that express HA protein at different surface densities. On the basis of quantitations of the number of HA trimers per cell and the relative surface areas of the two cell lines, the HAb-2 cells have a 1.9-fold higher plasma membrane surface density than the GP4F cells. The membrane lateral diffusion coefficient and the mobile fraction of HA is the same for both cell lines. A Scatchard analysis of the binding of glycophorin-bearing liposomes to the cells showed 1700 bindings sites for the GP4F cells and 3750 binding sites for the HAb-2 cells, with effectively the same liposome-cell binding constant, about 7 .times. 1010 M-1. Binding was specific for glycophorin on the liposomes and HA expressed on the cells. A competition experiment employing toxin-containing and empty liposomes allowed us to quantitate the number of liposomes that fused per cell, which was a small constant fraction of the number of bound liposomes. For the HAb-2 cells, about 1 in every 70 bound liposomes fused and for the GP4F cells about 1 in every 300 bound liposomes fused. Hence, the HAb-2 cells showed 4.4 times more fusion per bound liposome, even though the surface density of HA was only 1.9 times greater. We conclude the following: (i) One HA trimer is not sufficient to induce fusion. (ii) The HA bound to glycophorin is not the HA that induces fusion. That is, even though each HA has a binding and a fusion function, those functions are not performed by the same HA trimer.