Integrative imaging reveals SARS-CoV-2 induced reshaping of subcellular morphologies. Cortese et al.

In this study we combined light and electron microscopy imaging to determine the morphological organelle alterations induced by SARS-CoV-2 infection in lung epithelial cells. SARS-CoV-2, similar to other positive-strand RNA virus, replicates its genome in specialized virus-induced compartments called viral "replication organelles" (ROs). Additionally, ROs allow for enrichment of metabolites, viral enzymes and cofactors, and protect the viral RNA from degradation and sensing by pattern recognition receptors of the innate immune system. Double-membrane vesicles (DMVs) are the most abundant structure constituting coronaviral ROs. Whole cell 3D analysis of infected cells, generated through focus-ion beam scanning electron microscopy, revealed a network of DMVs embedded in an ER matrix (MOVIE S1). High-throughput electron microscopy allowed defining the 3D architecture of SARS-CoV-2 ROs (MOVIE S2, S3, S4) and their association with cellular organelles, such as mitochondria, peroxisomes, the Golgi apparatus and the cytoskeleton (MOVIE S5). In addition, super-resolution microscopy and live cell imaging shows how the cytoskeleton network is remodeled during the course of infection, forming a cage-like structure that surrounds the ROs (MOVIE S6 and S7). Pharmacological inhibition of cytoskeleton dynamics strongly impairs SARS-CoV-2 replication and virus particle secretion.本研究联用光学显微镜与电子显微镜成像技术，探究严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）感染肺上皮细胞所诱导的细胞器形态学改变。与其他正链RNA病毒类似，SARS-CoV-2会在病毒诱导形成的特化区室中完成基因组复制，这类区室被命名为病毒“复制细胞器（replication organelles，ROs）”。此外，复制细胞器可富集代谢物、病毒酶类及辅助因子，并能保护病毒RNA免遭先天免疫系统模式识别受体的降解与识别。双膜囊泡（double-membrane vesicles，DMVs）是构成冠状病毒复制细胞器的最主要结构组分。通过聚焦离子束扫描电子显微镜成像获取的感染细胞全细胞三维分析结果显示，双膜囊泡网络嵌入内质网（endoplasmic reticulum，ER）基质中（MOVIE S1）。高通量电子显微镜技术得以解析SARS-CoV-2复制细胞器的三维架构（MOVIE S2、S3、S4），以及其与线粒体、过氧化物酶体、高尔基体及细胞骨架等细胞细胞器的相互关联（MOVIE S5）。此外，超分辨率显微镜与活细胞成像技术揭示了感染进程中细胞骨架网络的重塑机制：其会形成包裹复制细胞器的笼状结构（MOVIE S6、S7）。对细胞骨架动力学进行药物抑制，可显著削弱SARS-CoV-2的复制效率与病毒粒子分泌能力。