Function of HNRNPC in breast cancer cells by controlling the dsRNA-induced interferon response

Elevated expression of RNA binding protein HNRNPC has been reported in cancer cells, while the essentialness and functions of HNRNPC in tumors were not clear. We showed that repression of HNRNPC in the breast cancer cells MCF7 and T47D inhibited cell proliferation and tumor growth. Our computational inference of the key pathways and extensive experimental investigations revealed that the cascade of interferon responses mediated by RIG-I was responsible for such tumor-inhibitory effect. Interestingly, repression of HNRNPC resulted in accumulation of endogenous double-stranded RNA (dsRNA), the binding ligand of RIG-I. These up-regulated dsRNA species were highly enriched by Alu sequences and mostly originated from pre-mRNA introns that harbor the known HNRNPC binding sites. Such source of dsRNA is different than the recently well-characterized endogenous retroviruses that encode dsRNA. In summary, essentialness of HNRNPC in the breast cancer cells was attributed to its function in controlling the endogenous dsRNA and the down-stream interferon response. This is a novel extension from the previous understandings about HNRNPC in binding with introns and regulating RNA splicing. Overall design: Cytoplasmic total RNAs were profiled upon HNRNPC knock-down in MCF7 (2 replicates) and T47D cells, NC and LAMN knock-down cells served as controls.已有研究报道，RNA结合蛋白异质性核核糖核蛋白C（HNRNPC）在癌细胞中呈高表达，但该蛋白在肿瘤中的必要性与功能尚未明确。本研究证实，在乳腺癌细胞MCF7与T47D中敲低HNRNPC可显著抑制细胞增殖与肿瘤生长。通过对关键通路的计算推演与大规模实验验证，本研究揭示：视黄酸诱导基因蛋白I（RIG-I）介导的干扰素应答级联反应是该抗肿瘤效应的核心机制。值得注意的是，敲低HNRNPC会导致内源性双链RNA（double-stranded RNA, dsRNA）的积累，而这类分子正是RIG-I的结合配体。上述上调的dsRNA分子高度富集Alu序列，且大多来源于携带已知HNRNPC结合位点的前mRNA（pre-mRNA）内含子区域。该类dsRNA的来源与近期已被充分表征的编码dsRNA的内源性逆转录病毒截然不同。综上，乳腺癌细胞中HNRNPC的必要性可归因于其调控内源性dsRNA及其下游干扰素应答的功能。这一发现是对既往认知的全新拓展——既往学界认为HNRNPC仅通过结合内含子参与调控RNA剪接。实验整体设计：在MCF7细胞（设2个生物学重复）与T47D细胞中敲低HNRNPC后，对其胞质总RNA进行表达谱分析；以阴性对照（negative control, NC）敲低细胞与LAMN敲低细胞作为对照。