HIF-1α/ESR1介导RhoE表达调控机制及其在胃癌上皮间质转化和侵袭中的机制研究

Invasion and metastasis are the most devastaing features of gastric cancer，while the underlying molecular mechnisms are still elusive.Our previos study proved that transactivation of RhoE by HIF-1α during hypoxia could enhance epithilial-mesenchymal transiton (EMT) and invasiveness of gastric cacner. But the regulating mechanisms of RhoE at transcriptional level are still unclear. Other studies have indicated that ESR1 can bind to the NDA of RhoE, while P53 mutants and HIF-1αcan inhibit the transcription of ESR1; Beside，P53 and HIF-1αcould interact with each other. We propose that the three transcription factors abovementioned may compose a complex molecular network to modulate RhoE expression and the EMT and invasion of gastric cancer. Methods including gene recombination,mutation,virus infection,gene reportor will be adopted in this study to investigate the regulatory roles of P53 mutants and HIF-1αfor ESR1 tanscription,RhoE expression，EMT and invasion of gastric cancer; furthermore, Co-IP、Pull-down，ChIP assays will be applied to the examination of interaction between P53，HIF-1α，ESR1 and RhoE at different levels.This study improve our understanding of the complex relationships between hypoxia, P53 mutation, ESR1 silencing and RhoE upregulation in the EMT and invasion process of gastric cancer, thus providing a novel theoretic basis for gastric cancer treatment.

侵袭和转移是胃癌最具破坏性的特征，但其分子机制仍不明了。我们前期研究发现HIF-1α在缺氧时激活RhoE表达，进而促进胃癌细胞的EMT和转移。然而RhoE DNA启动子区并不存在HIF-1α的结合位点，提示HIF-1α可能通过其他分子的中介间接调控RhoE转录。生物信息学分析提示RhoE的启动子区域存在ESR1结合位点，而且在肿瘤中ESR1与HIF-1α和RhoE的表达密切相关。同时基于在胃癌中普遍存在的突变型P53可与HIF-1α相互作用促进肿瘤恶性表型的进展，我们提出突变型P53与HIF-1α通过ESR1共同调控RhoE的表达，促进胃癌细胞EMT和转移。本研究拟采用基因克隆、报告基因、Co-IP、EMSA等技术，从细胞、动物和临床组织标本水平阐明HIF-1α、P53通过ESR1调控RhoE表达的分子调控网络并验证其在胃癌细胞发生EMT和侵袭转移中的作用机制，为胃癌侵袭治疗提供理论基础。

侵袭和转移是胃癌最具破坏性的特征，但其分子机制仍不明了。我们前期研究发现HIF-1α在缺氧时激活RhoE表达，进而促进胃癌细胞的EMT和转移。然而RhoE DNA启动子区并不存在HIF-1α的结合位点，提示HIF-1α可能通过其他分子的中介间接调控RhoE转录。生物信息学分析提示RhoE的启动子区域存在ESR1结合位点，而且在肿瘤中ESR1与HIF-1α和RhoE的表达密切相关。同时基于在胃癌中普遍存在的突变型P53可与HIF-1α相互作用促进肿瘤恶性表型的进展，我们提出突变型P53与HIF-1α通过ESR1共同调控RhoE的表达，促进胃癌细胞EMT和转移。本研究拟采用基因克隆、报告基因、Co-IP、EMSA等技术，从细胞、动物和临床组织标本水平阐明HIF-1α、P53通过ESR1调控RhoE表达的分子调控网络并验证其在胃癌细胞发生EMT和侵袭转移中的作用机制，为胃癌侵袭治疗提供理论基础。.我们通过质谱分析发现RhoE可以和LAMP2A 和HSC70 结合，提示它是伴侣分子介导的自噬（CMA）的底物。实验验证发现通过下调LMAP2A抑制CMA活性后RhoE水平上调，阐明了CMA从蛋白水平调控RhoE表达的新的分子机制。同时，我们发现上调RhoE促进胃癌细胞的EMT和迁移能力，突变型p53在缺氧条件下与HIF1α共同负性调控ESR1，而ESR1则负性调控RhoE的表达，从而参与调控胃癌细胞的侵袭转移。