Receptor-independent infection of murine coronavirus: Analysis by spinoculation
Receptor-independent infection of murine coronavirus: Analysis by spinoculation
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DOI：
10.1128/jvi.80.10.4901-4908.2006
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发表时间：
2006-05-01
影响因子：
5.4
通讯作者：
Taguchi, Fumihiro
中科院分区：
文献类型：
--
作者：
Watanabe, Rie;Matsuyama, Shutoku;Taguchi, Fumihiro
已知高神经毒性鼠冠状病毒 JHMV（野生型 [wt] JHMV）从通过鼠冠状病毒小鼠肝炎病毒受体（MHVR）感染的细胞传播到没有 MHVR（MHVR 独立感染）的细胞，而它的突变病毒分离前 wt JHMV，srr7，仅以 MVHR 依赖性方式传播。这些观察结果是通过将 JHMV 感染的细胞覆盖到对 wt JHMV 感染具有抗性的受体阴性细胞上而获得的。假设认为不依赖于 MHVR 的感染归因于 wt JHMV S 蛋白的自然发生的融合激活，而在 srr7 的情况下并未发生这种情况。在S蛋白激活过程中，没有MHVR的S蛋白油细胞的附着似乎是一个关键条件。因此，在本研究中，我们试图了解 wt JHMV 病毒粒子是否附着在 MHVR 上。阴性细胞能够感染这些细胞。为了使病毒颗粒在没有MHVR的情况下附着在细胞表面，我们使用了旋接种，即将细胞与接种的病毒一起以3,000rpm离心2小时。正如通过实时 PCR 定量估计所附着的病毒体所揭示的那样，该过程迫使病毒附着到细胞表面，并且还促进了 wt JHMV 感染 MHVR 阴性细胞，但对于 srr7 却未能做到这一点。在可溶形式的 MHVR 存在下，通过旋转接种附着在 MHVR 阴性细胞上的 wt 和 srr7 病毒颗粒有利于感染，诱导 wt 和 srr7 的构象变化。进一步揭示，当S蛋白在细胞上表达时，wt JHMV S1而不是srr7被释放到细胞表面。这些观察结果支持这样的假设：病毒体附着到 MHVR 阴性细胞是关键步骤，并且 wt JHMV S1 在自然发生的事件中在 S2 之前释放的独特特征涉及 MHVR 独立感染。
A highly neurovirulent murine coronavirus JHMV (wild-type [wt] JHMV) is known to spread from cells infected via the murine coronavirus mouse hepatitis virus receptor (MHVR) to cells without MHVR (MHVR-independent infection), whereas it mutant virus isolated front wt JHMV, srr7, spread only in in MVHR-dependent fashion. These observations were obtained by the overlay of JHMV-infected cells onto receptor-negative cells that are otherwise resistant to wt JHMV infection. MHVR-independent infection is hypothetically thought to be attributed to a naturally occurring fusion activation of the wt JHMV S protein, which did not occur in the case of srr7. Attachment of S protein oil cells without MHVR during the S-protein activation process seems to be a key condition. Thus, in the present study, we tried to see whether wt JHMV virions that are attached on MHVR. negative cells are able to infect those cells. In order to make virions attach to the cell surface without MHVR, we have used spinoculation, namely, the centrifugation of cells together with inoculated virus at 3,000 rpm for 2 h. This procedure forces viruses to attach to the cell surface, as revealed by quantitative estimation of attached virions by real-time PCR and also facilitated wt JHMV infection to MHVR-negative cells, but failed to do so for srr7. Virions of both wt and srr7 attached on MHVR-negative cells by, spinoculation were facilitated for infection in the presence of a soluble form of MHVR that induces conformational changes of both wt and srr7. It was further revealed that wt JHMV S1, but not srr7, was released front the cell surface when S protein was expressed on cells. These observations support the hypothesis that attachment of the virion to MHVR-negative cells is it critical step and that a unique feature of wt JHMV S1 to be released front S2 in a naturally, occurring event is involved in an MHVR-independent infection.