Contribution of host an viral small non-coding RNAs to SARS-CoV lung pathology

Severe acute respiratory syndrome coronavirus (SARS-CoV) causes lethal disease in humans, with viral E protein promoting the exacerbated inflammatory response. By deep sequencing RNAs from the lungs of infected mice, we have addressed the relevance of small, non-coding RNAs in SARS-CoV pathology. Host microRNAs (miRNAs) expressed during infection by a virulent virus encoding the E protein were significantly enriched for cytokine-mediated inflammatory pathways when compared with attenuated SARS-CoV-∆E, suggesting contribution of miRNAs to E protein-induced inflammation. The discovery of three 18-22 nt small viral RNAs (svRNAs) derived from the nsp3 and N genomic regions of SARS-CoV in mouse lung and cell cultures is also described. Depletion of these svRNAs significantly reduced viral titers and genomic RNA levels, indicating their positive contribution to virus growth. Remarkably, svRNA-N antagomirs significantly reduced in vivo lung pathology and pro-inflammatory cytokine expression, indicating that svRNAs contribute to SARS-CoV pathogenesis and highlighting the potential of these antagomirs as antivirals. Strand-specific, single-end, reads were generated for detecting smallRNAs (18 nts or more) in lungs (mouse). Three or four replicates were prepared per sample type.

严重急性呼吸综合征冠状病毒（SARS-CoV）可引发人类致死性疾病，其病毒E蛋白（E protein）会介导过度炎症反应。本研究通过对感染小鼠肺部的RNA进行深度测序，探究了小型非编码RNA（small non-coding RNAs）在SARS-CoV致病过程中的作用。相较于缺失E蛋白的减毒SARS-CoV（SARS-CoV-∆E），编码E蛋白的强毒株感染过程中诱导产生的宿主微小RNA（microRNAs，以下简称miRNAs），在细胞因子介导的炎症通路中显著富集，提示miRNAs参与了E蛋白诱导的炎症反应。本研究还报道了在小鼠肺部及细胞培养物中发现的3种长度为18~22 nt的病毒小型RNA（small viral RNAs，以下简称svRNAs），其序列源自SARS-CoV的非结构蛋白3（nsp3）及N基因组区域。敲低这些svRNAs可显著降低病毒滴度及基因组RNA水平，表明其对病毒增殖具有正向促进作用。值得注意的是，svRNA-N反义拮抗剂（antagomirs）可显著减轻体内肺部病理损伤并下调促炎细胞因子表达，证实svRNAs参与了SARS-CoV的致病过程，同时凸显了这类反义拮抗剂作为抗病毒药物的应用潜力。本研究采用链特异性单端测序技术，对小鼠肺部的小型RNA（长度≥18 nt）进行测序，每种样本类型设置3~4次生物学重复。