State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, School of Basic Medicine, Beijing, China.
Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, School of Basic Medicine, Beijing, China.
Sci Rep. 2018 Jul 13;8(1):10669. doi: 10.1038/s41598-018-28868-y.
Viral fusion proteins are essential for enveloped virus infection. These proteins mediate fusion between the virus envelope and host cellular membrane to release the viral genome into cells. Vesicular stomatitis virus G (VSV G) protein is a typical type III viral fusion protein. To study the mechanism of VSV G protein mediated membrane fusion, we set up a cell-cell fusion system in which cells are marked by different fluorescent proteins. Taking advantage of this system, we performed real-time recording and quantitative analysis of the cell fusion mediated by VSV G. We found that the time scale required for VSV G mediated cell-cell fusion was approximately 1-2 minutes. Next, we specifically examined the function of the transmembrane (TM) region of VSV G protein in membrane fusion by replacing the TM region with those of other fusion proteins. The TM region replacements dramatically impaired VSV G protein function in the cell-cell fusion assay and diminished VSV G mediated lentivirus and recombinant VSV infection efficiency. Further experiments implied that the TM region played a role in the transition from hemi-fusion to full fusion. Several residues within the TM region were identified as important for membrane fusion. Overall, our findings unraveled the important function of the TM region in VSV G mediated viral fusion.
病毒融合蛋白对于包膜病毒的感染至关重要。这些蛋白介导病毒包膜与宿主细胞膜之间的融合，将病毒基因组释放到细胞中。水疱性口炎病毒 G（VSV G）蛋白是一种典型的 III 型病毒融合蛋白。为了研究 VSV G 蛋白介导的膜融合机制，我们建立了一个细胞-细胞融合系统，其中细胞被不同的荧光蛋白标记。利用该系统，我们对 VSV G 介导的细胞融合进行了实时记录和定量分析。我们发现，VSV G 介导的细胞-细胞融合所需的时间尺度约为 1-2 分钟。接下来，我们通过用其他融合蛋白的跨膜（TM）区域替换 TM 区域，特异性地研究了 VSV G 蛋白 TM 区域在膜融合中的功能。TM 区域替换显著损害了 VSV G 蛋白在细胞-细胞融合测定中的功能，并降低了 VSV G 介导的慢病毒和重组 VSV 的感染效率。进一步的实验表明，TM 区域在从半融合到完全融合的转变中起作用。TM 区域内的几个残基被确定对膜融合很重要。总的来说，我们的发现揭示了 TM 区域在 VSV G 介导的病毒融合中的重要功能。
