The Human Respiratory Syncytial Virus Matrix Protein Is Required for Maturation of Viral Filaments
The Human Respiratory Syncytial Virus Matrix Protein Is Required for Maturation of Viral Filaments
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DOI：
10.1128/jvi.06744-11
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发表时间：
2012-04-01
影响因子：
5.4
通讯作者：
Oomens, Antonius G. P.
中科院分区：
文献类型：
--
作者：
Mitra, Ruchira;Baviskar, Pradyumna;Oomens, Antonius G. P.
开发了一个实验系统，用于从 cDNA 生成缺乏基质 (M) 蛋白表达的传染性人类呼吸道合胞病毒 (HRSV)（M 无效病毒）。然后通过用 M 无效病毒感染 HEp-2 和 Vero 细胞并评估对感染性病毒产生和病毒蛋白运输的影响来检查 M 蛋白在病毒组装中的作用。在缺乏 M 的情况下，感染性后代的产生会受到严重损害。使用针对核蛋白 (N)、附着蛋白 (G) 和融合蛋白 (F) 的抗体进行的免疫荧光 (IF) 显微镜分析未能检测到特征性病毒诱导的细胞表面细丝，而这些细丝被认为代表了感染性病毒体。此外，在称为包涵体（IB）的病毒复制工厂中检测到了很大一部分 N 蛋白。通过场发射扫描电子显微镜 (SEM) 对 M 无效病毒感染的细胞表面进行高分辨率分析，发现存在大面积的密集、均匀的短丝。尽管异常短，但这些细丝在其他方面与含有 M 的对照病毒诱导的细丝相似，包括病毒 G 和 F 蛋白的存在。在缺乏 M 的情况下，短而发育不良的丝的丰度表明，M 在丝形成的初始阶段不是必需的，但在这些结构的成熟或伸长中起着重要作用。此外，成熟病毒丝的缺失以及 IB 内 N 蛋白水平的同时增加表明，M 蛋白参与了病毒核糖核蛋白 (RNP) 复合物从细胞质 IB 到出芽位点的转运。
An experimental system was developed to generate infectious human respiratory syncytial virus (HRSV) lacking matrix (M) protein expression (M-null virus) from cDNA. The role of the M protein in virus assembly was then examined by infecting HEp-2 and Vero cells with the M-null virus and assessing the impact on infectious virus production and viral protein trafficking. In the absence of M, the production of infectious progeny was strongly impaired. Immunofluorescence (IF) microscopy analysis using antibodies against the nucleoprotein (N), attachment protein (G), and fusion protein (F) failed to detect the characteristic virus induced cell surface filaments, which are believed to represent infectious virions. In addition, a large proportion of the N protein was detected in viral replication factories termed inclusion bodies (IBs). High-resolution analysis of the surface of M-null virus-infected cells by field emission scanning electron microscopy (SEM) revealed the presence of large areas with densely packed, uniformly short filaments. Although unusually short, these filaments were otherwise similar to those induced by an M-containing control virus, including the presence of the viral G and F proteins. The abundance of the short, stunted filaments in the absence of M indicates that M is not required for the initial stages of filament formation but plays an important role in the maturation or elongation of these structures. In addition, the absence of mature viral filaments and the simultaneous increase in the level of the N protein within IBs suggest that the M protein is involved in the transport of viral ribonucleoprotein (RNP) complexes from cytoplasmic IBs to sites of budding.