PTEN与TGFBR1互为ceRNA调控口腔鳞癌自噬的分子机制

项目名称： PTEN与TGFBR1互为ceRNA调控口腔鳞癌自噬的分子机制

项目编号： No.81472528

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 孙志军

作者单位： 武汉大学

项目金额： 85万元

中文摘要： CeRNA(competing endogenous RNA) 是一种通过竞争共享miRNA而调控靶基因的内源性RNA。通过前期研究,本项目提出假说：口腔黏膜上皮及口腔癌中抑癌基因PTEN及TGFBR1互为ceRNA,通过竞争共享miRNA调控对方mRNA的表达，进而调控自噬信号通路及自噬相关基因的表达，影响口腔癌的自噬及癌发生。拟从以下方面开展工作：①利用口腔癌细胞系、基因敲除动物模型和人类组织标本分析和验证PTEN及TGFBR1共享miRNA，并通过转染彼此 3'UTR及敲减DICER的策略验证PTEN及TGFBR1的ceRNA关系。②明确共享miRNA调控自噬相关基因对自噬及上皮恶性转化的分子机制。③设计外源竞争性miRNA海绵，探索体外及转基因动物口腔癌的实验性治疗。研究结果将加深对口腔癌mRNA-miRNA-mRNA分子网络调控机制的理解，为口腔癌的治疗提供新的靶点及策略。

中文关键词： C01_口腔肿瘤；微小RNA；自噬；转基因动物；竞争内源性RNA

英文摘要： CeRNA(competing endogenous RNA) is a kind of endogenous RNAs that share multiple MREs(miRNA response elements) and regulate their targets in a microRNA(miRNA) competing based mechanism. In this project,we hypothesize that PTEN and TGFBR1, two key tumor suppressor gene in oral squamous cell carcinoma(OSCC) may have the potential interaction as ceRNA, which regulate autophagy and carcinogenesis of OSCC by regulate a panel of shared miRNAs. This project include three parts:Firstly,we testify the hypothesis by bioinformatic analysis and data mining.The expression pattern of PTEN and TGFBR1 shared miRNA in human OSCC cell lines, transgenic mice OSCC and human OSCC samples were detected by in situ hybridization and qPCR.Biological validation of PTEN and TGFBR1 regulated miRNA were performed using knock down and RNA-IP method.Furthermore,functional PTEN and TGFBR1 3'UTR transfection and DICER knock down assay were performed to confirm the ceRNA interaction of PTEN and TGFBR1. Secondly,we explore the molecular mechanism of PTEN and TGFBR1 shared miRNA's influence on autophagy and OSCC initiation.We will focus potent shared oncomiR such as miR-130a and miR-181a on their target autophagy related gene(ATGs) using qPCR, confocal microscope and living cell imaging in OSCC cell lines as well as PTEN/TGFBR1 knock out mice. In vitro foci formation assay and in vivo carcinogenesis were performed using shared miRNA inhibitor or mimics. Thirdly, we will further design combined exogenous competing miRNA sponges to explore experimental therapeutics of OSCC by inhibiting PTEN and TGFBR1 shared oncomiR in vitro and in vivo. Achievments of this project will better understand the multiple layer regulation of key tumor suppressor gene in autophagy and carcinogenesis of OSCC and develop new strategies for OSCC treatment.

英文关键词： oral cancer;microRNA;autophagy;transgenic mice;ceRNA