The membrane-proximal domain of vesicular stomatitis virus G protein functions as a membrane fusion potentiator and can induce hemifusion
The membrane-proximal domain of vesicular stomatitis virus G protein functions as a membrane fusion potentiator and can induce hemifusion
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DOI：
10.1128/jvi.76.23.12300-12311.2002
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发表时间：
2002-12-01
影响因子：
5.4
通讯作者：
Whitt, MA
中科院分区：
文献类型：
--
作者：
Jeetendra, E;Robison, CS;Whitt, MA
最近我们发现水疱性口炎病毒（VSV）的G蛋白外域（G stem [GS]）与跨膜和细胞质结构域一起足以介导VSV的有效出芽（C. S. Robison， M. A. Whitt， J.中国生物医学工程学报，74:22 -22,2000）。本研究表明，当GS与异种病毒融合蛋白共表达时，GS也能增强异种病毒融合蛋白的膜融合活性。对于某些融合蛋白，GS共表达时合胞体的形成比单独表达时增加了40倍。GS的融合增强不是蛋白质特异性的，因为它既发生在ph依赖性融合蛋白上，也发生在ph非依赖性融合蛋白上。利用含有n端血凝素（RA）标签的重组水疱性口炎病毒（GS（HA）病毒）编码GS，我们发现GS（HA）病毒在pH 7.0时与野生型病毒一样与细胞结合；而GS（HA）病毒无传染性。对表达GS（HA）的细胞进行三色膜融合分析发现，GS（HA）可诱导脂质混合，但不能诱导胞质混合，表明GS可诱导半灌注。用破坏膜稳定的药物氯丙嗪处理GS（HA）病毒结合的细胞，恢复了半融合阻滞，允许GS（HA）病毒进入并随后复制，表明GS介导的半融合是膜融合途径中的一个功能中间体。使用一系列截断突变体，我们还确定只有14个GS残基，以及VSV G跨膜和细胞质尾部，足以进行融合增强。据我们所知，这是首次报道一种病毒糖蛋白的小结构域可以促进其他不相关的病毒糖蛋白的融合活性。
Recently we showed that the membrane-proximal stem region of the vesicular stomatitis virus (VSV) G protein ectodomain (G stem [GS]), together with the transmembrane and cytoplasmic domains, was sufficient to mediate efficient VSV budding (C. S. Robison and M. A. Whitt, J. Virol. 74:2239-2246, 2000). Here, we show that GS can also potentiate the membrane fusion activity of heterologous viral fusion proteins when GS is coexpressed with those proteins. For some fusion proteins, there was as much as a 40-fold increase in syncytium formation when GS was coexpressed compared to that seen when the fusion protein was expressed alone. Fusion potentiation by GS was not protein specific, since it occurred with both pH-dependent as well as pH-independent fusion proteins. Using a recombinant vesicular stomatitis virus encoding GS that contained an N-terminal hemagglutinin (RA) tag (GS(HA) virus), we found that the GS(HA) virus bound to cells as well as the wild-type virus did at pH 7.0; however, the GS(HA) virus was noninfectious. Analysis of cells expressing GS(HA) in a three-color membrane fusion assay revealed that GS(HA) could induce lipid mixing but not cytoplasmic mixing, indicating that GS can induce hemifusion. Treatment of GS(HA) virus-bound cells with the membrane-destabilizing drug chlorpromazine rescued the hemifusion block and allowed entry and subsequent replication of GS(HA) virus, demonstrating that GS-mediated hemifusion was a functional intermediate in the membrane fusion pathway. Using a series of truncation mutants, we also determined that only 14 residues of GS, together with the VSV G transmembrane and cytoplasmic tail, were sufficient for fusion potentiation. To our knowledge, this is the first report which shows that a small domain of one viral glycoprotein can promote the fusion activity of other, unrelated viral glycoproteins.