CircRBM3细胞内转运HuR蛋白及其经外泌体细胞间传递调控膀胱癌进展的作用与机制研究

The important role of circular RNA (circRNA) in the regulation of gene expression has been gradually recognized in recent years, and its function and mechanism need to be further explored. Recent studies have shown that the exosomes of tumor cells have a high abundance of circRNA. Based on our previous study of bladder cancer circRNA expression, our preliminary data showed that the expression of circRBM3 in serum exosomes of patients with bladder cancer was significantly high, and the exosomes containing high level of circRBM3 could promote vascular formation of vascular endothelial cells. Moreover, circRBM3 also enhanced the invasion of bladder cancer cells. This study will further reveal the role and underlying mechanism of circRBM3/ miR-145 interaction in regulation of VEGF expression in vascular endothelial cells, and to elucidate the effect and mechanism of circRBM3 on binding and transporting HuR protein to regulate the expression of MMP-9 in bladder cancer cells. The successful implementation of this project will elucidate the mode and mechanism of exosome-mediated transmission of circRNA from a completely new perspective, as well as reveal the role of circRNA in intracellular transport of functional protein, and the research content has potential clinical application value in targeted therapy of bladder cancer.

环状RNA（circRNA）对基因表达调控的重要作用近年来逐渐被认知，其功能和作用机理有待进一步探究。新近研究表明，肿瘤细胞外泌体中含有丰富的circRNA。我们在前期测序分析得到膀胱癌组织circRNA表达谱基础上，初步研究显示：circRBM3在膀胱癌患者血清外泌体中显著高表达，富含circRBM3的膀胱癌细胞外泌体能够有效促进血管内皮细胞血管形成，circRBM3亦能增强膀胱癌细胞侵袭能力。本研究拟在此基础上，进一步解析circRBM3通过膀胱癌细胞外泌体传递至血管内皮细胞后吸附miR-145调控VEGF表达的作用及机制，并阐明circRBM3在膀胱癌细胞结合并转运HuR蛋白对mmp-9的表达调控作用及机制。本项目的成功实施将从全新的角度阐明外泌体传递circRNA的作用模式及机制，并有望首次揭示circRNA调控靶蛋白细胞定位作用，研究内容具有潜在膀胱癌靶向治疗临床应用价值。

环状RNA（circRNA）对基因表达调控的重要作用近年来逐渐被认知，其功能和作用机理有待进一步探究。新近研究表明，肿瘤细胞外泌体中含有丰富的circRNA。我们在前期测序分析得到膀胱癌组织circRNA表达谱基础上，初步研究显示：circRBM3在膀胱癌患者血清外泌体中显著高表达，circRBM3能够增强膀胱癌细胞侵袭能力。阐明了circRBM3在膀胱癌细胞结合并转运HuR蛋白对mmp-9的表达调控作用及其促进膀胱癌肺转移的作用。此外，circRBM3在膀胱癌组织及细胞中显著高表达并与膀胱癌病人的不良预后密切相关，circRBM3与WTAP在膀胱癌组织中的表达呈显著正相关；RIP及RNA pull-down实验进一步证明circRBM3与WTAP可以在膀胱癌细胞核中直接结合。关于circRBM3的作用机制，我们证明circRBM3可以与WTAP结合促进m6A甲基转移酶复合物组装。细胞功能学实验及裸鼠成瘤实验证实，敲降circRBM3可以在体内体外水平显著促进细胞凋亡，而过表达WTAP能够逆转这种效果。此外，我们筛选前期测序数据并验证发现TNFAIP3在膀胱癌组织及细胞中显著高表达且其表达水平与膀胱癌分级、侵袭以及淋巴结转移等临床指标正相关。细胞与动物实验证实干扰TNFAIP3的表达能够促进细胞凋亡。circRBM3/WTAP复合物以依赖m6A的方式增加其mRNA稳定性来促进TNFAIP3表达。最后我们的研究证明了circRBM3/WTAP/TNFAIP3通路沉默提高膀胱癌CDDP化疗敏感性的作用，为膀胱癌CDDP耐药的突破提供了新思路。