hnRNP C modulates MERS-CoV and SARS-CoV-2 replication by governing the expression of a subset of circRNAs and cognitive mRNAs

Host circular RNAs (circRNAs) play critical roles in the pathogenesis of viral infections. However, how viruses modulate the biogenesis of host proviral circRNAs to facilitate their replication remains unclear. We have recently shown that Middle East respiratory syndrome coronavirus (MERS-CoV) infection increases co-expression of circRNAs and their cognate messenger RNAs (mRNAs), possibly by hijacking specific host RNA binding proteins (RBPs). In this study, we systemically analysed the interactions between the representative circRNA–mRNA pairs upregulated upon MERS-CoV infection and host RBPs. Our analysis identified heterogeneous nuclear ribonucleoprotein C (hnRNP C) as a key host factor that governed the expression of numerous MERS-CoV-perturbed circRNAs, including hsa_circ_0002846, hsa_circ_0002061, and hsa_circ_0004445. RNA immunoprecipitation assay showed that hnRNP C could bind physically to these circRNAs. Specific knockdown of hnRNP C by small interfering RNA significantly (P P 50: 0.618 µM) or SARS-CoV-2-infected (IC50: 1.233 µM) Calu-3 cells with the mTOR inhibitor OSI-027 resulted in significantly reduced viral loads. Collectively, our study identified hnRNP C as a key regulator of MERS-CoV-perturbed circRNAs and their cognate mRNAs, and the potential of targeting hnRNP C-related signalling pathways as an anticoronaviral strategy.

环状RNA（circular RNAs, circRNAs）在病毒感染的致病机制中发挥关键作用。然而，病毒如何调控宿主促病毒环状RNA的生物发生以促进自身复制，目前仍不明确。我们近期的研究表明，中东呼吸综合征冠状病毒（Middle East respiratory syndrome coronavirus, MERS-CoV）感染可通过劫持特定宿主RNA结合蛋白（RNA binding proteins, RBPs），上调环状RNA与其同源信使RNA（messenger RNAs, mRNAs）的共表达水平。本研究系统分析了MERS-CoV感染后上调的典型circRNA-mRNA配对与宿主RBPs之间的相互作用。研究鉴定出异质性核核糖核蛋白C（heterogeneous nuclear ribonucleoprotein C, hnRNP C）作为关键宿主因子，可调控众多受MERS-CoV扰动的环状RNA的表达，其中包括hsa_circ_0002846、hsa_circ_0002061以及hsa_circ_0004445。RNA免疫沉淀实验（RNA immunoprecipitation assay）证实，hnRNP C可与这些环状RNA发生直接结合。通过小干扰RNA（small interfering RNA, siRNA）特异性敲低hnRNP C可显著降低病毒载量（P<0.05，原文此处“P P 50”为输入笔误）；此外，用哺乳动物雷帕霉素靶蛋白（mammalian target of rapamycin, mTOR）抑制剂OSI-027处理感染MERS-CoV（半数抑制浓度half maximal inhibitory concentration, IC50: 0.618 µM）或新型冠状病毒（Severe Acute Respiratory Syndrome Coronavirus 2, SARS-CoV-2，半数抑制浓度IC50: 1.233 µM）的Calu-3细胞，可显著降低病毒载量。综上，本研究鉴定出hnRNP C作为受MERS-CoV扰动的环状RNA及其同源信使RNA的关键调控因子，并揭示了靶向hnRNP C相关信号通路作为抗冠状病毒治疗策略的潜力。