Characterizing spatial and global host innate immune responses against SARS-CoV-1 and -2 infection in pathologically relevant human lung epithelial cells

Severe acute respiratory syndrome coronavirus-1 (SARS-CoV-1) and -2 (SARS-CoV-2) are beta-coronaviruses (ß-CoVs) that have caused significant morbidity and mortality worldwide. Therefore, a better understanding of host responses to ß-CoVs would provide insights into the pathogenesis of these viruses to identify potential targets for medical countermeasures. Our objective is to use a systems biology approach to explore the magnitude and scope of innate immune responses triggered by SARS-CoV-1 and -2 infection over time in pathologically relevant human lung epithelial cells (Calu-3/2B4 cells). Total RNA extracted at 12, 24, and 48 hours after ß-CoVs infection of Calu-3/2B4 cells were subjected to RNA sequencing, bioinformatics, and functional enrichment analysis to select genes whose expressions were significantly modulated upon infection over time. The results of our study demonstrate that SARS-CoV-1 and -2 stimulate similar yet distinct innate antiviral signaling pathways in pathologically relevant human lung epithelial cells. Upregulated genes found at multiple time points regulated the viral life cycle, interferons, and interferon-stimulated genes (ISGs). Based on their profound modulation upon infection, we selected four ISGs, i.e., bone marrow stromal cell antigen 2 (BST2), Z-DNA Binding Protein 1 (ZBP1), C-X-C Motif Chemokine Ligand 11 (CXCL11), and Interferon Induced Transmembrane Protein 1 (IFITM1) for characterization as potential drug targets against ß-CoVs. Overall design: Calu-3/2B4 cells were subjected to SARS-CoV-1 or SARS-CoV-2 infection at a multiplicity of infection (MOI) of 1.0. Our controls were uninfected (mock infected) Calu-3/2B4 cells. At three different time points (12-, 24-, and 48-hours post-infection), we collected total RNA. Afterwards, gene expression profiling analysis of RNA-seq data was performed comparing SARS-CoV-1- and SARS-CoV-2-infected to mock-infected cells and SARS-CoV-1- to SARS-CoV-2-infected cells.

严重急性呼吸综合征冠状病毒1型（SARS-CoV-1）与冠状病毒2型（SARS-CoV-2）均属于β冠状病毒（beta-coronaviruses），曾在全球范围内引发大规模发病与死亡。因此，深入解析宿主针对β冠状病毒的免疫应答，可为阐明此类病毒的致病机制、发掘医疗应对策略的潜在靶点提供理论支撑。本研究旨在借助系统生物学方法，探究病理相关人肺上皮细胞（Calu-3/2B4细胞）中，SARS-CoV-1与SARS-CoV-2感染随时间进程触发的固有免疫应答的强度与范围。研究人员在β冠状病毒感染Calu-3/2B4细胞后的12、24及48小时提取总RNA，随后开展RNA测序、生物信息学及功能富集分析，筛选出随感染进程表达发生显著变化的基因。本研究结果显示，SARS-CoV-1与SARS-CoV-2可在病理相关人肺上皮细胞中激活相似却存在差异的固有抗病毒信号通路。多个时间点上调的基因参与调控病毒生命周期、干扰素及干扰素刺激基因（interferon-stimulated genes, ISGs）。基于感染后表达的显著变化，我们筛选出4种干扰素刺激基因，即骨髓基质细胞抗原2（bone marrow stromal cell antigen 2, BST2）、Z-DNA结合蛋白1（Z-DNA Binding Protein 1, ZBP1）、C-X-C基序趋化因子配体11（C-X-C Motif Chemokine Ligand 11, CXCL11）以及干扰素诱导跨膜蛋白1（Interferon Induced Transmembrane Protein 1, IFITM1），将其作为对抗β冠状病毒的潜在药物靶点进行后续表征。整体实验设计：以感染复数（multiplicity of infection, MOI）为1.0的SARS-CoV-1或SARS-CoV-2感染Calu-3/2B4细胞，设置未感染（模拟感染）的Calu-3/2B4细胞作为对照组。分别在感染后12、24及48小时三个时间点收集总RNA。随后对RNA测序数据进行基因表达谱分析，比较SARS-CoV-1感染组、SARS-CoV-2感染组与模拟感染组的差异，同时对比SARS-CoV-1与SARS-CoV-2感染组之间的差异。