GPI-anchored influenza hemagglutinin induces hemifusion to both red blood cell and planar bilayer membranes.
GPI-anchored influenza hemagglutinin induces hemifusion to both red blood cell and planar bilayer membranes.
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DOI：
10.1083/jcb.131.3.679
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发表时间：
1995-11
期刊：
影响因子：
--
通讯作者：
Cohen FS
中科院分区：
文献类型：
--
作者：
Melikyan GB;White JM;Cohen FS
在融合条件下，荧光染料从红细胞（RBC）的外单层小叶重新分布到表达糖磷脂酰肌醇锚定的流感病毒血凝素（GPI-HA）的细胞，而无需水性染料转移。这表明发生了半融合，但没有完全融合（Kemble, G. W., T. Danieli, and J. M. White. 1994. Cell. 76:383-391）。我们通过用 FM4-64 标记红细胞的内部单层小叶并观察这些内部小叶没有与 GPI-HA 表达细胞连续，从而扩展了半融合的证据。半融合分离的水内容物区域，即半融合膈肌，似乎被延长并且寿命很长。但是，当与表达 GPI-HA 的细胞半融合的红细胞渗透性膨胀时，一些隔膜被破坏，并且观察到内部小叶和水性染料的扩散。这是完全融合的特征：内部小叶和水性探针扩散至表达野生型HA (wt-HA)的细胞。通过同步视频荧光显微镜和时间分辨电导纳测量，我们严格证明了表达 GPI-HA 的细胞与平面双层膜半融合：建立了脂质连续性，而没有形成融合孔。半融合面积变大。相反，对于表达wt-HA的细胞，在脂质染料扩散之前，总是观察到融合孔，证实发生了完全融合。我们提出了一种弹性耦合模型，其中 wt-HA 的胞外域诱导半融合，而表达 GPI-HA 的细胞中不存在的跨膜域则介导完全融合。
Under fusogenic conditions, fluorescent dye redistributed from the outer monolayer leaflet of red blood cells (RBCs) to cells expressing glycophosphatidylinositol-anchored influenza virus hemagglutinin (GPI- HA) without transfer of aqueous dye. This suggests that hemifusion, but not full fusion, occurred (Kemble, G. W., T. Danieli, and J. M. White. 1994. Cell. 76:383-391). We extended the evidence for hemifusion by labeling the inner monolayer leaflets of RBCs with FM4-64 and observing that these inner leaflets did not become continuous with GPI-HA- expressing cells. The region of hemifusion-separated aqueous contents, the hemifusion diaphragm, appeared to be extended and was long-lived. But when RBCs hemifused to GPI-HA-expressing cells were osmotically swollen, some diaphragms were disrupted, and spread of both inner leaflet and aqueous dyes was observed. This was characteristic of full fusion: inner leaflet and aqueous probes spread to cells expressing wild-type HA (wt-HA). By simultaneous video fluorescence microscopy and time-resolved electrical admittance measurements, we rigorously demonstrated that GPI-HA-expressing cells hemifuse to planar bilayer membranes: lipid continuity was established without formation of fusion pores. The hemifusion area became large. In contrast, for cells expressing wt-HA, before lipid dye spread, fusion pores were always observed, establishing that full fusion occurred. We present an elastic coupling model in which the ectodomain of wt-HA induces hemifusion and the transmembrane domain, absent in the GPI-HA-expressing cells, mediates full fusion.