AFAP1-AS1结合SNIP1上调c-Myc转录活性促进肺癌侵袭转移的机制研究

In our previous study, we first found that long non-coding RNA AFAP1-AS1 was abnormally highly expressed in lung cancer. AFAP1-AS1 promote the invasion and metastasis，but its molecular mechanism is still unclear. In this project, we found that AFAP1-AS1 can bind to SNIP1 through the RNA pull down experiment and the high-resolution mass spectrometry analysis. AFAP1-AS1 is involved in the regulation process of SNIP1 on c-Myc protein level. AFAP1-AS1 mediates the combination of SNIP1 and c-Myc. Combined with the existing work foundation and literature analysis, the scientific hypothesis of this project is: AFAP1-AS1 binds to SNIP1 and up-regulates the transcriptional activity of c-Myc. On the one hand, AFAP1-AS1 synergistically with SNIP1 inhibits the ubiquitination of c-Myc and up-regulates the protein level of c-Myc.On the other hand, AFAP1-AS1 mediates the formation of SNIP1/ c-Myc /p300 protein complex, and directs the complex to the promoter region of c-Myc target gene to initiate target gene transcription. The completion of this project will further clarify the molecular mechanism of lung cancer invasion and metastasis, provide new targets for the diagnosis and treatment of lung cancer, and provide new molecular markers for prognosis judgment.

本课题组前期首次发现长链非编码RNA AFAP1-AS1在肺癌中异常高表达，促进肺癌细胞的侵袭转移，但其分子机制尚不明确。在本项目中，我们通过RNA pull down结合高分辨质谱分析，发现AFAP1-AS1可与SNIP1结合；AFAP1-AS1参与SNIP1对c-Myc蛋白水平的调控过程；AFAP1-AS1介导SNIP1与c-Myc结合。结合已有工作基础及文献分析，本项目的科学假说为：AFAP1-AS1与SNIP1结合，上调c-Myc的转录活性，一方面，AFAP1-AS1协同SNIP1，抑制c-Myc泛素化修饰，上调c-Myc的表达；另一方面，AFAP1-AS1介导SNIP1/c-Myc/p300蛋白复合物的形成，并指引该复合物至c-Myc靶基因启动子区，启动靶基因转录。本项目的完成，将进一步明确肺癌侵袭转移的分子机制，为肺癌患者的诊断及治疗提供新的靶点，为预后判断提供新的分子标记。

肌动蛋白丝相关蛋白1反义RNA 1 (AFAP1-AS1)是一个在多种肿瘤中异常高表达的长链非编码RNA。本研究旨在探讨AFAP1-AS1在肺癌发生发展中的作用及机制。我们首先采用原位杂交技术检测了187例石蜡包埋肺癌和36例正常肺上皮组织中AFAP1-AS1的表达，研究了AFAP1-AS1在肺癌细胞中的迁移和侵袭能力。为了揭示AFAP1-AS1促进肺癌发生发展的分子机制，我们通过RNA pull down和质谱分析筛选出AFAP1-AS1的潜在相互作用蛋白。我们的研究结果发现，AFAP1-AS1在肺癌临床组织中高表达，其表达与肺癌患者的不良预后呈正相关。体内实验证实AFAP1-AS1可促进肺癌转移。AFAP1-AS1通过与Smad核相互作用蛋白1(SNIP1)相互作用促进肺癌细胞的迁移和侵袭，抑制c-Myc蛋白的泛素化，进而影响其降解。c-Myc分子的上调又促进了ZEB1、ZEB2和SNAIL基因的表达，最终促进了上皮向间充质转化(epithelial - to mesenchymal transition, EMT)和肺癌转移。明确AFAP1-AS1促进肺癌迁移和侵袭的分子机制，可能为肺癌患者的早期诊断和治疗提供新的治疗靶点。