COPII Vesicle Transport Is Required for Rotavirus NSP4 Interaction with the Autophagy Protein LC3 II and Trafficking to Viroplasms
COPII Vesicle Transport Is Required for Rotavirus NSP4 Interaction with the Autophagy Protein LC3 II and Trafficking to Viroplasms
复制标题
DOI：
10.1128/jvi.01341-19
复制
发表时间：
2020-01-01
影响因子：
5.4
通讯作者：
Estes, Mary K.
中科院分区：
文献类型：
--
作者：
Crawford, Sue E.;Criglar, Jeanette M.;Estes, Mary K.
许多在细胞质中复制的病毒通过知之甚少的机制显著地重塑和刺激宿主细胞膜的积累以进行有效复制。对于轮状病毒，病毒体组装的关键步骤需要在病毒复制中心附近积累膜，称为病毒膜。早期的电子显微镜研究将病毒质相关的膜描述为“肿胀的”内质网（ER）。我们以前证明，轮状病毒感染启动细胞自噬和自噬标志物蛋白LC 3和轮状病毒ER合成的跨膜糖蛋白NSP 4的交通病毒，表明NSP 4必须退出ER的膜。本研究的目的是解决NSP 4退出ER的机制，并确定是否病毒相关的膜ER衍生。我们报告说：（i）NSP 4退出COPII囊泡中的ER，导致COPII囊泡转运和ER退出位点中断;（ii）COPII囊泡被LC 3 II劫持，LC 3 II与NSP 4相互作用;和（iii）含NSP 4/LC 3 II的膜积聚在病毒质附近。此外，ER跨膜蛋白SERCA和calnexin在病毒质相关膜中未检测到，提供了轮状病毒成熟过程“出芽”通过自噬劫持COPII囊泡膜发生的证据。这些研究结果揭示了一个新的机制轮状病毒成熟依赖于细胞内宿主蛋白质运输和自噬的积累所需的病毒replication.IMPORTANCE在一个形态发生的步骤，这是非常罕见的无包膜病毒，不成熟的轮状病毒颗粒组装在复制中心称为病毒囊，并通过细胞质细胞膜芽获得感染性颗粒组装的外壳蛋白。历史上，用于颗粒出芽的细胞内膜被认为是内质网（ER），因为与未成熟颗粒相互作用以触发出芽的轮状病毒非结构蛋白NSP 4被合成为ER跨膜蛋白。目前的研究表明，NSP 4退出ER的COPII囊泡和NSP 4的COPII囊泡被劫持的细胞自噬机制，介导的NSP 4的运输病毒。改变轮状病毒成熟的范例，我们提出，不成熟轮状病毒颗粒出芽所需的细胞膜不是ER的延伸，而是COPII衍生的自噬隔离膜。
Many viruses that replicate in the cytoplasm dramatically remodel and stimulate the accumulation of host cell membranes for efficient replication by poorly understood mechanisms. For rotavirus, a critical step in virion assembly requires the accumulation of membranes adjacent to virus replication centers called viroplasms. Early electron microscopy studies describe viroplasm-associated membranes as "swollen" endoplasmic reticulum (ER). We previously demonstrated that rotavirus infection initiates cellular autophagy and that membranes containing the autophagy marker protein LC3 and the rotavirus ER-synthesized transmembrane glycoprotein NSP4 traffic to viroplasms, suggesting that NSP4 must exit the ER. This study aimed to address the mechanism of NSP4 exit from the ER and determine whether the viroplasm-associated membranes are ER derived. We report that (i) NSP4 exits the ER in COPII vesicles, resulting in disrupted COPII vesicle transport and ER exit sites; (ii) COPII vesicles are hijacked by LC3 II, which interacts with NSP4; and (iii) NSP4/LC3 II-containing membranes accumulate adjacent to viroplasms. In addition, the ER transmembrane proteins SERCA and calnexin were not detected in viroplasm-associated membranes, providing evidence that the rotavirus maturation process of "budding" occurs through autophagy-hijacked COPII vesicle membranes. These findings reveal a new mechanism for rotavirus maturation dependent on intracellular host protein transport and autophagy for the accumulation of membranes required for virus replication.IMPORTANCE In a morphogenic step that is exceedingly rare for nonenveloped viruses, immature rotavirus particles assemble in replication centers called viroplasms, and bud through cytoplasmic cellular membranes to acquire the outer capsid proteins for infectious particle assembly. Historically, the intracellular membranes used for particle budding were thought to be endoplasmic reticulum (ER) because the rotavirus nonstructural protein NSP4, which interacts with the immature particles to trigger budding, is synthesized as an ER transmembrane protein. This present study shows that NSP4 exits the ER in COPII vesicles and that the NSP4-containing COPII vesicles are hijacked by the cellular autophagy machinery, which mediates the trafficking of NSP4 to viroplasms. Changing the paradigm for rotavirus maturation, we propose that the cellular membranes required for immature rotavirus particle budding are not an extension of the ER but are COPII-derived autophagy isolation membranes.