HNRNPR介导的剪接变体UPF3B-S促进肝细胞癌浸润转移的机制研究

Tumor invasion and metastasis are the main causes of poor prognosis of hepatocellular carcinoma(HCC). Deregulation of alternative splicing has been implicated in the malignant progression of HCC. However, its underlying mechanism in HCC remains elusive. Our preliminary data showed that the expression of UPF3B-S (a truncated splice variant) was up-regulated in HCC, and associated with advanced tumor node metastasis (TNM) stage, vascular invasion and unfavorable prognosis. UPF3B-S silence inhibited the proliferation and migration of HCC cells and suppressed cyclinB1/Cdk1 signal pathway. Furthermore, UPF3B-S was regulated by HNRNPR. Based on our findings, we assume that splice variant UPF3B-S which was mediated by HNRNPR, promotes HCC invasion and metastasis via cyclinB1/Cdk1 signaling. In this study, we propose to determine the expression and clinicopathological significances of UPF3B-S and HNRNPR in HCC, to disclose the underlying mechanism of HNRNPR on regulating UPF3B-S expression, and to explore the role of HNRNPR/UPF3B-S/cyclinB1/Cdk1 in cell proliferation and migration of HCC. Our study probably provides not only the solid evidence of UPF3B-S involved in the malignant process of HCC, but also a prognostic biomarker and a promising therapeutic target in HCC.

侵袭转移是肝细胞癌（HCC）预后差的主要原因。研究表明基因异常剪接参与HCC恶性进展，但具体分子调控机制不明。我们前期研究发现剪接变体UPF3B-S在HCC中高表达，且与患者TNM分期晚、血管侵犯和预后差相关；沉默UPF3B-S抑制cyclinB1/Cdk1通路，抑制HCC细胞增殖与迁移；UPF3B-S表达受剪接因子HNRNPR调控。因此，我们提出HNRNPR介导的剪接变体UPF3B-S促进HCC浸润转移的假说。本项目拟采用一系列分子细胞生物学实验，利用多种细胞和动物模型，在细胞功能及基因表达调控上，明确UPF3B-S和HNRNPR在HCC中的表达及临床意义，揭示HNRNPR调控UPF3B-S的具体机制，探讨UPF3B-S对下游通路的调控方式，阐明HNRNPR/UPF3B-S/cyclinB1/Cdk1信号轴在HCC进展中的作用，为诠释HCC发生发展的分子机制和寻找治疗的新靶点提供依据。

有研究表明，可变剪接的异常调控参与肝细胞癌（HCC）细胞侵袭转移。对于侵袭性HCC细胞中异常可变剪接的深入研究有助于发现新的标志物和治疗靶点。HCC组织转录组分析显示，具侵袭特征的HCC组织中可变剪接通路相关基因高度富集。其中筛选出剪接变体UPF3B-S（UPF3B缺少8号外显子的短片段）为与侵袭性HCC相关的重要剪接变体。BaseScope检测表明，UPF3B-S在HCC中的表达显著高于癌旁肝组织，UPF3B-S在侵袭性HCC中的表达明显高于非侵袭性HCC组织。UPF3B-S高表达与HCC患者TNM分期晚、预后差显著相关。UPF3B-S对有侵袭表型HCC具有预测潜力。体内体外功能实验证实，UPF3B-S可促进HCC细胞侵袭、迁移能力。机制上，HnRNPR与UPF3B pre-mRNA结合并引起UPF3B基因8号外显子跳跃。其中，HnRNPR基因的RRM2结构域在上调UPF3B-S的表达中发挥了关键作用。UPF3B-S通过调节Hippo通路中LATS1-YAP1-BIRC5信号轴，促进了HCC细胞的侵袭和转移。进一步研究表明，UPF3B-S通过调节CDH1 mRNA 3'-UTR长度，降解CDH1 mRNA，导致LATS1和YAP1去磷酸化。综上，本研究表明，HnRNPR介导的UPF3B-S通过降解CDH1 mRNA抑制Hippo信号通路，促进HCC侵袭和转移，表明UPF3B-S可能作为侵袭性HCC的一个有前途的生物标志物和治疗HCC的一个潜在靶点。