Assembly of the coronavirus envelope: Homotypic interactions between the M proteins
Assembly of the coronavirus envelope: Homotypic interactions between the M proteins
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DOI：
10.1128/jvi.74.11.4967-4978.2000
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发表时间：
2000-06-01
影响因子：
5.4
通讯作者：
Rottier, PJM
中科院分区：
文献类型：
--
作者：
de Haan, CAM;Vennema, H;Rottier, PJM
病毒膜蛋白M和E是冠状病毒颗粒出芽的最低要求。由于E蛋白仅以痕量存在于颗粒中，ICI蛋白之间的横向相互作用显然产生了包膜形成的主要驱动力。通过使用免疫共沉淀和包膜掺入试验，我们提供了广泛的证据，这种M-M相互作用的存在。此外，我们确定哪些领域的Ri蛋白参与这种同型协会，使用诱变方法。突变体hi蛋白本身不能组装成病毒样颗粒（VLP）（C，A. M，de哈安湖郭文贵马斯特斯，H。Vennema和P.J.M. Rottier，J. Virol。七十二：6838-6850，1998）测试与其它R-I蛋白缔合的能力以及被拯救到由有组装能力的M蛋白形成的VLP中的能力。我们发现，M蛋白缺乏部分的跨膜簇，的两亲性结构域，或亲水性羧基末端的尾巴，或M蛋白，其管腔结构域被异源胞外域取代，仍然能够与组装能力的Ri蛋白，导致其共掺入到VLP，只有一个突变的RI蛋白，其中所有三个跨膜结构域已被取代失去了这种能力。结果表明，M蛋白分子通过多个接触位点相互作用，特别是在跨膜水平。最后，我们通过探测一些外源蛋白的共组装来测试膜蛋白被选择用于掺入冠状病毒包膜的严格性。观察到的有效排除水泡性口炎病毒G蛋白和马动脉炎病毒M蛋白的出芽表明，包膜组装确实是一个高度选择性的分选过程。然而，CD 8分子的低但可检测的掺入表明该过程并不完美。
The viral membrane proteins M and E are the minimal requirements for the budding of coronavirus particles. Since the E protein occurs in particles only in trace amounts, the lateral interactions between the ICI proteins apparently generate the major driving force for envelope formation. By using coimmunoprecipitation and envelope incorporation assays, we provide extensive evidence for the existence of such M-M interactions. In addition, we determined which domains of the Ri protein are involved in this homotypic association, using a mutagenetic approach. Mutant hi proteins which were not able to assemble into viruslike particles (VLPs) by themselves (C, A. M, de Haan, L. Kuo, P. S. Masters, H. Vennema, and P. J. M. Rottier, J. Virol. 72: 6838-6850, 1998) were tested for the ability to associate with other R-I proteins and to be rescued into VLPs formed by assembly-competent M proteins. We found that M proteins lacking parts of the transmembrane cluster, of the amphipathic domain, or of the hydrophilic carboxy-terminal tail, or M proteins that had their luminal domain replaced by heterologous ectodomains, were still able to associate with assembly-competent Ri proteins, resulting in their coincorporation into VLPs, Only a mutant RI protein in which all three transmembrane domains had been replaced lost this ability. The results indicate that M protein molecules interact with each other through multiple contact sites, particularly at the transmembrane level. Finally, we tested the stringency with which membrane proteins are selected for incorporation into the coronavirus envelope by probing the coassembly of some foreign proteins. The observed efficient exclusion from budding of the vesicular stomatitis virus G protein and the equine arteritis virus M protein indicates that envelope assembly is indeed a highly selective sorting process. The low but detectable incorporation of CD8 molecules, however, demonstrated that this process is not perfect.