Virus-Like Vesicles of Kaposi's Sarcoma-Associated Herpesvirus Activate Lytic Replication by Triggering Differentiation Signaling
Virus-Like Vesicles of Kaposi's Sarcoma-Associated Herpesvirus Activate Lytic Replication by Triggering Differentiation Signaling
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DOI：
10.1128/jvi.00362-17
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发表时间：
2017-08-01
影响因子：
5.4
通讯作者：
Sun, Ren
中科院分区：
文献类型：
--
作者：
Gong, Danyang;Dai, Xinghong;Sun, Ren
病毒样囊泡（VLV）是膜封闭的囊泡，在组织上类似于天然包膜病毒，但缺乏病毒衣壳和基因组。在肿瘤相关伽马疱疹病毒的生产性感染过程中，病毒粒子和 VLV 都会产生并释放到细胞外空间。然而，对伽玛疱疹病毒相关VLV的研究在很大程度上受到从成熟病毒颗粒中分离VLV的技术难度的限制。在这里，我们报告了一种利用卡波西肉瘤相关疱疹病毒（KSHV）突变体选择性分离 VLV 的策略，该突变体的小衣壳蛋白有缺陷，无法产生成熟的病毒体。通过质谱分析，我们发现 VLV 含有进入细胞所需的病毒糖蛋白，以及参与调节裂解复制的外皮蛋白，但缺乏衣壳蛋白。功能分析表明，VLV 诱导病毒裂解激活剂 RTA 的表达，从而启动 KSHV 裂解基因表达。此外，我们利用 RNA 测序对 VLV 触发的细胞反应进行了全基因组分析，发现细胞分化的主要调节因子 PRDM1 显着上调。在 KSHV 复制的背景下，我们证明 VLV 诱导的 PRDM1 上调对于通过激活其 RTA 启动子来重新激活 KSHV 是必要且充分的。总之，我们的研究系统地检查了 VLV 的组成，并证明了它们在操纵宿主细胞反应和促进 KSHV 裂解性复制方面的生物学作用。 重要性 被肿瘤相关疱疹病毒裂解感染的细胞会产生高比例的病毒样囊泡 (VLV)。 VLV 的组成和功能尚未明确，很大程度上是由于无法有效分离不含病毒颗粒的 VLV。使用能够建立潜伏 KSHV 感染和强大重新激活的细胞系统，我们成功地从小衣壳蛋白缺陷的 KSHV 突变体中分离出 VLV。我们定量分析了 VLV 中的蛋白质和 microRNA，并表征了 VLV 在操纵宿主细胞和促进病毒感染中的作用。更重要的是，我们证明，通过上调 PRDM1 表达，VLV 触发靶细胞中的分化信号，并通过激活 RTA 启动子促进病毒裂解感染。我们的研究不仅展示了分离 VLV 的新策略，而且还展示了 KSHV 相关 VLV 在细胞间通讯和病毒生命周期中的重要作用。
Virus-like vesicles (VLVs) are membrane-enclosed vesicles that resemble native enveloped viruses in organization but lack the viral capsid and genome. During the productive infection of tumor-associated gammaherpesviruses, both virions and VLVs are produced and are released into the extracellular space. However, studies of gammaherpesvirus-associated VLVs have been largely restricted by the technical difficulty of separating VLVs from mature virions. Here we report a strategy of selectively isolating VLVs by using a Kaposi's sarcoma-associated herpesvirus (KSHV) mutant that is defective in small capsid protein and is unable to produce mature virions. Using mass spectrometry analysis, we found that VLVs contained viral glycoproteins required for cellular entry, as well as tegument proteins involved in regulating lytic replication, but lacked capsid proteins. Functional analysis showed that VLVs induced the expression of the viral lytic activator RTA, initiating KSHV lytic gene expression. Furthermore, employing RNA sequencing, we performed a genome-wide analysis of cellular responses triggered by VLVs and found that PRDM1, a master regulator in cell differentiation, was significantly upregulated. In the context of KSHV replication, we demonstrated that VLV-induced upregulation of PRDM1 was necessary and sufficient to reactivate KSHV by activating its RTA promoter. In sum, our study systematically examined the composition of VLVs and demonstrated their biological roles in manipulating host cell responses and facilitating KSHV lytic replication.IMPORTANCE Cells lytically infected with tumor-associated herpesviruses produce a high proportion of virus-like vesicles (VLVs). The composition and function of VLVs have not been well defined, largely due to the inability to efficiently isolate VLVs that are free of virions. Using a cell system capable of establishing latent KSHV infection and robust reactivation, we successfully isolated VLVs from a KSHV mutant defective in the small capsid protein. We quantitatively analyzed proteins and microRNAs in VLVs and characterized the roles of VLVs in manipulating host cells and facilitating viral infection. More importantly, we demonstrated that by upregulating PRDM1 expression, VLVs triggered differentiation signaling in targeted cells and facilitated viral lytic infection via activation of the RTA promoter. Our study not only demonstrates a new strategy for isolating VLVs but also shows the important roles of KSHV-associated VLVs in intercellular communication and the viral life cycle.