一个新的环状RNA分子circRNF13在舌鳞癌细胞耐药中的作用机制研究

Our previous data demonstrated that the circular RNA circRNF13 was down-regulated in tongue squamous cell carcinoma (TSCC), and low circRNF13 expression was associated with poor prognosis and drug resistance in TSCC; overexpression of circRNF13 in TSCC cells arrested cell cycle progression, induced apoptosis, and reversed drug resistance phenotype. We also confirmed that the circRNF13 and RNF13 mRNA were processed by the same precursor, and their expression was negatively correlated. Using the latest proteomics platform, we obtained differential expression protein profile from TSCC cells before and after overexpression and knockdown of circRNF13. Through further bioinformatics analyses, we proposed the following hypothesis: circRNF13 can competitive inhibit the mRNA splicing and protein expression of the E3 ubiquitin linked enzyme, RNF13, and can function as a miR-3147 sponge to relieve the inhibition of miR-3147 on the SUMOylation enzyme SUMO2, thus circRNF13 catalyzes post-translational modification (ubiquitination and/or SUMOylation) of important proteins through RNF13 and SUMO2 in cells, and finally regulates cell cycle progression, apoptosis and other related signal pathways, and affects the drug resistance of TSCC cells. In the proposed project, we will reveal the biological function of circRNF13 through a series of experiments in vivo and in vitro and provide new molecular markers and targets for the diagnosis and treatment of TSCC.

我们在舌鳞癌中发现circRNF13表达下调，且与患者不良预后及癌细胞耐药相关；过表达circRNF13可阻滞细胞周期、促进凋亡和逆转耐药表型。我们还证实circRNF13与RNF13基因的mRNA由相同前体加工而成，且它们的表达呈负相关。利用最新蛋白组学平台获得过表达和敲低circRNF13前后舌鳞癌细胞中差异表达蛋白谱后，结合生物信息学分析，我们提出了如下科学假说：circRNF13可竞争性抑制RNF13这一E3泛素链接酶的mRNA剪接和蛋白表达，也可作为miR-3147的“海绵”解除miR-3147对SUMO2这一SUMO化调节酶的抑制作用，从而通过RNF13和SUMO2催化细胞内重要蛋白的翻译后修饰(泛素化和/或SUMO化)，最终调控细胞周期、凋亡等相关信号通路，影响舌鳞癌的耐药。本项目将通过一系列体内外实验揭示circRNF13的生物学功能并为舌鳞癌的诊疗提供新的分子标记和靶点。

越来越多的研究表明环状RNAs (circular RNAs, circRNAs) 在肿瘤的发生发展过程中发挥着重要的功能，且在肿瘤标志物的开发应用方面circRNA也具有明显优势。本项目研究发现顺铂可诱导舌鳞癌细胞中circRNA circRNF13表达上调，circRNF13通过阻滞细胞周期进程和诱导细胞凋亡抑制舌鳞癌细胞的增殖，过表达circRNF13可逆转舌鳞癌细胞的耐药；circRNF13在舌鳞癌组织中表达下调且表达水平与患者预后相关；circRNF13通过竞争性抑制RNF13 mRNA的剪接加工和RNF13蛋白的表达，阻滞舌鳞癌细胞周期进程并促进舌鳞癌细胞凋亡；circRNF13通过circRNF13-miR-3147-SUMO2轴以及直接结合SUMO2影响SUMO2的稳定性，调控舌鳞癌细胞周期进程和细胞凋亡；RNF13和SUMO2通过调控FOXO1影响舌鳞癌细胞周期和凋亡，从而调节舌鳞癌细胞的耐药。此外，我们还在鼻咽癌中研究了circRNA circRNF13的功能和作用机制：发现circRNA circRNF13在鼻咽癌中同样表达下调，且circRNF13能抑制鼻咽癌细胞的增殖、迁移和侵袭；circRNA circRNF13通过促进GLUT1泛素化抑制糖酵解，从而抑制肿瘤细胞的迁移和侵袭；circRNA circRNF13通过直接结合SUMO2 mRNA的3′‑UTR来促进SUMO2的稳定性，进而通过SUMO2促进GLUT1的泛素化；circRNA circRNF13通过SUMO2抑制肿瘤细胞的增殖、迁移和侵袭。上述研究结果表明，circRNA circRNF13在肿瘤细胞中发挥抑瘤基因的作用，相关结果已获授权发明专利和发表在SCI期刊上。本课题的相关研究结果为进一步阐明头颈肿瘤的发病机理提供了重要的理论基础和并提供了新的治疗靶点。