Laboratory of Immunobiochemistry, Division of Bacterial, Parasitic and Allergenic Products, Office of Vaccines Research and Review, Center for Biologics Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, Maryland, USA.
Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
J Virol. 2018 Aug 16;92(17). doi: 10.1128/JVI.00518-18. Print 2018 Sep 1.
Host-influenza virus interplay at the transcript level has been extensively characterized in epithelial cells. Yet, there are no studies that simultaneously characterize human host and influenza A virus (IAV) genomes. We infected human bronchial epithelial BEAS-2B cells with two seasonal IAV/H3N2 strains, Brisbane/10/07 and Perth/16/09 (reference strains for past vaccine seasons) and the well-characterized laboratory strain Udorn/307/72. Strand-specific RNA sequencing (RNA-seq) of the infected BEAS-2B cells allowed for simultaneous analysis of host and viral transcriptomes, in addition to pathogen genomes, to reveal changes in mRNA expression and alternative splicing (AS). In general, patterns of global and immune gene expression induced by the three IAVs were mostly shared. However, AS of host transcripts and small nuclear RNAs differed between the seasonal and laboratory strains. Analysis of viral transcriptomes showed deletions of the polymerase components (defective interfering-like RNAs) within the genome. Surprisingly, we found that the neuraminidase gene undergoes AS and that the splicing event differs between seasonal and laboratory strains. Our findings reveal novel elements of the host-virus interaction and highlight the importance of RNA-seq in identifying molecular changes at the genome level that may contribute to shaping RNA-based innate immunity. The use of massively parallel RNA sequencing (RNA-seq) has revealed insights into human and pathogen genomes and their evolution. Dual RNA-seq allows simultaneous dissection of host and pathogen genomes and strand-specific RNA-seq provides information about the polarity of the RNA. This is important in the case of negative-strand RNA viruses like influenza virus, which generate positive (complementary and mRNA) and negative-strand RNAs (genome) that differ in their potential to trigger innate immunity. Here, we characterize interactions between human bronchial epithelial cells and three influenza A/H3N2 strains using strand-specific dual RNA-seq. We focused on this subtype because of its epidemiological importance in causing significant morbidity and mortality during influenza epidemics. We report novel elements that differ between seasonal and laboratory strains highlighting the complexity of the host-virus interplay at the RNA level.
宿主-流感病毒在转录水平上的相互作用在上皮细胞中得到了广泛的研究。然而，目前还没有研究同时对人类宿主和甲型流感病毒（IAV）基因组进行特征分析。我们用两种季节性的 IAV/H3N2 株，布里斯班/10/07 和珀斯/16/09（过去疫苗季节的参考株）和经过充分研究的实验室株乌汶/307/72 感染人支气管上皮 BEAS-2B 细胞。感染 BEAS-2B 细胞的链特异性 RNA 测序（RNA-seq）允许同时分析宿主和病毒的转录组，以及病原体基因组，以揭示 mRNA 表达和选择性剪接（AS）的变化。总的来说，三种 IAV 诱导的全局和免疫基因表达模式大多是共享的。然而，季节性和实验室株之间宿主转录物和小核 RNA 的 AS 存在差异。病毒转录组的分析显示聚合酶成分（缺陷干扰样 RNA）在基因组内缺失。令人惊讶的是，我们发现神经氨酸酶基因发生了 AS，而且这种剪接事件在季节性和实验室株之间存在差异。我们的研究结果揭示了宿主-病毒相互作用的新元素，并强调了 RNA-seq 在识别基因组水平上可能导致 RNA 固有免疫形成的分子变化方面的重要性。大规模平行 RNA 测序（RNA-seq）的使用揭示了人类和病原体基因组及其进化的见解。双 RNA-seq 允许同时对宿主和病原体基因组进行剖析，而链特异性 RNA-seq 提供了关于 RNA 极性的信息。这在流感病毒等负链 RNA 病毒的情况下很重要，因为它们产生的正（互补和 mRNA）和负链 RNA（基因组）在触发固有免疫的潜力上有所不同。在这里，我们使用链特异性双 RNA-seq 来描述人支气管上皮细胞与三种甲型流感病毒/H3N2 株之间的相互作用。我们之所以关注这种亚型，是因为它在流感流行期间引起严重发病率和死亡率方面具有流行病学上的重要性。我们报告了季节性和实验室株之间存在差异的新元素，突出了宿主-病毒在 RNA 水平上相互作用的复杂性。
