Genetic analysis of determinants for spike glycoprotein assembly into murine coronavirus virions: Distinct roles for charge-rich and cysteine-rich regions of the endodomain
Genetic analysis of determinants for spike glycoprotein assembly into murine coronavirus virions: Distinct roles for charge-rich and cysteine-rich regions of the endodomain
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DOI：
10.1128/jvi.78.18.9904-9917.2004
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发表时间：
2004-09-01
影响因子：
5.4
通讯作者：
Masters, PS
中科院分区：
文献类型：
--
作者：
Ye, R;Montalto-Morrison, C;Masters, PS
冠状病毒刺突蛋白（S）形成了独特的病毒体表面结构，这是该病毒家族的特征，在负染电子显微镜下显示为被球形球覆盖的茎。这些结构对于通过病毒附着于细胞受体随后融合于宿主细胞膜而引发感染是必不可少的。S蛋白还可以介导感染细胞中合胞体的形成。S蛋白是一种I型跨膜蛋白，与其他病毒融合蛋白相比非常大，并且除了S分子的短羧基末端区段之外的所有区段都构成胞外域。对于原型冠状病毒小鼠肝炎病毒（MHV），先前已经确定S蛋白组装成病毒体是由羧基末端区段指定的，所述羧基末端区段包括跨膜结构域和胞内结构域。我们已经从遗传学上解剖了MHV S蛋白中的这些结构域，以定位S掺入病毒体的决定因素。我们的研究结果表明，装配能力映射到S的胞内结构域，这被证明是足够的目标异源整合膜蛋白纳入MHV病毒粒子。特别是，突变分析表明，一个主要的作用，电荷丰富的羧基末端区域的胞内域。此外，我们发现胞内结构域的邻近富含半胱氨酸的区域对于感染细胞的融合是至关重要的，证实了先前用S蛋白表达系统获得的结果。
The coronavirus spike protein (S) forms the distinctive virion surface structures that are characteristic of this viral family, appearing in negatively stained electron microscopy as stems capped with spherical bulbs. These structures are essential for the initiation of infection through attachment of the virus to cellular receptors followed by fusion to host cell membranes. The S protein can also mediate the formation of syncytia in infected cells. The S protein is a type I transmembrane protein that is very large compared to other viral fusion proteins, and all except a short carboxy-terminal segment of the S molecule constitutes the ectodomain. For the prototype coronavirus mouse hepatitis virus (MHV), it has previously been established that S protein assembly into virions is specified by the carboxy-terminal segment, which comprises the transmembrane domain and the endodomain. We have genetically dissected these domains in the MHV S protein to localize the determinants of S incorporation into virions. Our results establish that assembly competence maps to the endodomain of S, which was shown to be sufficient to target a heterologous integral membrane protein for incorporation into MHV virions. In particular, mutational analysis indicated a major role for the charge-rich carboxy-terminal region of the endodomain. Additionally, we found that the adjacent cysteine-rich region of the endodomain is critical for fusion of infected cells, confirming results previously obtained with S protein expression systems.