剪接变体USO1S促进肝癌浸润转移的机制研究

Hepatocellular carcinoma (HCC) is of high morbidity and mortality. It has been demonstrated that alternative splicing plays an important role in cancer progression. However, its underlying mechanism in HCC remains elusive. Recently, we found that USO1 gene was aberrantly spliced in 62.5% (5/8) of HCC samples. The splicing variant USO1S was highly expressed in HCC. Further studies showed that knockdown of USO1S inhibited cell proliferation and migration. In addition, bioinformatics analyses suggested USO1S was a potential downstream target of the splicing factor SRSF3. Based on our findings, we assume that splicing variant USO1S, regulated by SRSF3, promotes HCC cell proliferation and migration. In this study, we propose to determine the expression and clinical significance of USO1S and SRSF3 in HCC, to disclose the mechanism through which SRSF3 regulates USO1S, and to identify the role of USO1S in HCC progression and its targeting signaling pathway. Our study is likely to provide solid evidence that USO1S serves as a promising therapeutic target for HCC treatment.

肝细胞癌恶性程度高，预后差。已有研究表明基因可变剪接在肿瘤的发生发展中发挥重要作用，但其在肝细胞癌中的机制尚未明了。我们通过高通量转录组测序发现USO1基因在62.5%（5/8）的肝细胞癌样本中发生可变剪接，其剪接变体USO1S在肝癌细胞中高表达；另外，体外实验发现沉默USO1S抑制肝癌细胞增殖与迁移；同时，生物信息学及共表达分析发现，USO1S受剪接因子SRSF3调控。因此，我们提出SRSF3介导的剪接变体USO1S促进肝癌浸润转移的假说。本项目拟采用一系列分子生物学方法，利用人体标本、多种细胞株和动物模型，在细胞功能和基因表达调控上，明确SRSF3和USO1S在肝细胞癌中的表达及其临床意义，揭示SRSF3调控USO1S的分子机制，寻找USO1S促进肝癌细胞增殖迁移的下游靶点，并解析其作用的信号传导通路，为发展USO1S作为肝细胞癌治疗新靶点提供依据。

研究背景：可变剪接可以使一个基因转录出多样化的mRNA并翻译出不同功能的蛋白产物。研究表明，可变剪接的失调控常与肿瘤的恶性进展相关。在本研究中，我们着力于筛选影响肝细胞癌恶性进展的剪接变体事件，并探索其在肝细胞癌发生发展中的生物学功能及临床应用价值。.主要研究内容：我们从TCGA网站下载肝细胞癌的剪接变体事件及预后数据并作相应分析。应用BaseScope技术在肝癌新鲜组织及组织芯片中分析USO1S在肝癌细胞中的表达。运用CCK8和克隆形成等体外实验研究USO1S在促进肝癌细胞增殖中的作用。通过裸鼠皮下成瘤模型评估USO1S在体内对肝癌细胞增殖的作用。通过流式细胞学技术研究USO1S对肝癌细胞凋亡的作用。通过高通量测序探索USO1S下游靶点/相关信号通路。.重要结果及关键数据：我们构建了外显子跳跃事件预测模型来预测肝细胞癌患者的预后。我们发现USO1S在肝细胞癌组织中显著高表达，且伴有USO1SmRNA阳性表达的肝癌患者疾病生存时间短。沉默USO1S表达导致肝癌细胞生长受抑制且细胞凋亡增加。RNA测序结果表明，USO1S可能通过p53信号通路促进肝细胞癌恶性进展。.科学意义：我们的研究结果证实，USO1S通过促进肝癌细胞增殖、抑制肝癌细胞凋亡参与肝细胞癌恶性进展，为诠释肝细胞癌发生发展的分子机制提供科学依据。USO1S在肝细胞癌中高表达且预示预后不良，提示USO1S的mRNA水平有望成为肝细胞癌精准诊断及预后预测的分子标记物。