Data_Sheet_4_Long non-coding RNA SNHG9 regulates viral replication in rhabdomyosarcoma cells infected with enterovirus D68 via miR-150-5p/c-Fos axis.PDF

BackgroundThe Enterovirus D68 (EV-D68) epidemic has increased knowledge of the virus as a pathogen capable of causing serious respiratory and neurological illnesses. It has been shown that long noncoding RNAs (lncRNAs) regulate viral replication and infection via multiple mechanisms or signaling pathways. However, the precise function of lncRNAs in EV-D68 infection remains unknown.
MethodsThe differential expression profiles of lncRNA in EV-D68-infected and uninfected rhabdomyosarcoma (RD) cells were studied using high-throughput sequencing technology. The knockdown through small interfering RNA (siRNA) and overexpression of lncRNA SNHG9 (small ribonucleic acid host gene 9) were applied to investigate how lncRNA SNHG9 regulates EV-D68 propagation. The targeted interactions of lncRNA SNHG9 with miR-150-5p and miR-150-5p with c-Fos were validated using dual luciferase reporter system. LncRNA SNHG9 knockdown and miR-150-5p inhibitor were co-transfected with RD cells. QRT-PCR and western blot were used to detect RNA and protein levels, of c-Fos and VP1, respectively. Median tissue culture infectious dose (TCID50) was applied to detect viral titers.
ResultsThe results demonstrated that a total of 375 lncRNAs were highly dysregulated in the EV-D68 infection model. In the EV-D68 infection model, lncRNA SNHG9 and c-Fos were increased in EV-D68-infected RD cells. However, the expression level of miR-150-5p was downregulated. In addition, overexpression of SNHG9 in RD cells resulted in decreased viral replication levels and viral titers following infection with EV-D68, and further experiments revealed that overexpression of SNHG9 inhibited the viral replication by targeting increased miR-150-5p binding and significantly increased c-Fos expression in RD cells.
ConclusionOur findings indicate that the SNHG9/miR-150-5p/c-Fos axis influences EV-D68 replication in host cells and that SNHG9 may be a possible target for anti-EV-D68 infection therapies.

【背景】肠道病毒D68（Enterovirus D68, EV-D68）的流行加深了学界对该病毒的认知：其作为病原体可引发严重的呼吸道与神经系统疾病。已有研究证实，长链非编码RNA（long noncoding RNAs, lncRNAs）可通过多种机制或信号通路调控病毒复制与感染过程，但lncRNAs在EV-D68感染过程中的具体功能仍未明确。
【方法】本研究采用高通量测序技术，分析EV-D68感染与未感染的横纹肌肉瘤（rhabdomyosarcoma, RD）细胞中lncRNAs的差异表达谱。通过小干扰RNA（small interfering RNA, siRNA）介导的基因敲低以及lncRNA SNHG9（小核糖核酸宿主基因9, small ribonucleic acid host gene 9）过表达，探究SNHG9对EV-D68增殖的调控作用。利用双荧光素酶报告系统验证SNHG9与miR-150-5p、miR-150-5p与c-Fos的靶向结合关系。将SNHG9敲低体系与miR-150-5p抑制剂共转染至RD细胞中，分别采用实时荧光定量PCR（QRT-PCR）与蛋白质印迹（western blot）检测c-Fos与VP1的RNA及蛋白表达水平，并通过半数组织培养感染剂量（median tissue culture infectious dose, TCID₅₀）测定病毒滴度。
【结果】研究结果显示，EV-D68感染模型中共有375个lncRNAs呈现显著表达失调。在该感染模型中，EV-D68感染的RD细胞内SNHG9与c-Fos的表达水平均升高，而miR-150-5p的表达水平则呈下调趋势。此外，在RD细胞中过表达SNHG9可降低EV-D68感染后的病毒复制水平与病毒滴度；进一步实验证实，SNHG9过表达可通过增强miR-150-5p的结合能力，显著上调RD细胞内c-Fos的表达，从而抑制病毒复制。
【结论】本研究结果表明，SNHG9/miR-150-5p/c-Fos调控轴可影响宿主细胞内EV-D68的复制过程，SNHG9或可成为抗EV-D68感染治疗的潜在靶点。