eIF5A2调控食管癌细胞代谢重编程机制研究

In the previous study, we found that eIF5A2 could promote tumor metastasis and angiogenesis. The overall survival of esophageal squamous cell carcinoma (ESCC) patients with eIF5A2 overexpression was poorer than patients with eIF5A2 normal expression. EIF5A2 could be induced by gene amplification and hypoxia; Bidirectional regulation between eIF5A2 and HIF1a was also observed. cDNA microarray analysis demonstrated that some genes related to glucose and lipid metabolism changed in eIF5A2 overexpressed cells compared with vector control cells. HIF1a plays an important role in metabolism net,and the preliminary data indicated that eIF5A2 was involved in cell metabolism reprograming. In the present study, 1) Chromosome immuno precipitation (CHIP) assay and luciferase assay are performed to investigate the molecular mechanism between eIF5A2-HIF1a bidirectional regulation; 2) Mass spec analyses are performed to investigate the metabolism reprograming regulated by eIF5A2; 3)to study whether eIF5A2 regulating cell metabolism reprograming is dependent on HIF1a. Our research will focus on these three points and investigate the metabolism reprograming in ESCC.

我们前期研究发现eIF5A2具有促进肿瘤转移和血管生成的作用，与食管鳞癌患者不良预后密切相关。深入研究发现缺氧和基因扩增都可引起eIF5A2表达增高，并且eIF5A2与HIF1a互相调控。表达谱芯片分析显示eIF5A2升高导致一些与糖代谢和脂类代谢相关的基因表达发生变化。已知HIF1a是代谢网络中重要转录因子，预实验结果又提示eIF5A2参与调控细胞代谢重编程。因此，本研究中我们将：1）采用染色质免疫沉淀（CHIP）和萤光素酶实验研究eIF5A2与HIF1a互相调控的分子机制；2）采用超高压液相色谱联合质谱及磁共振技术进行细胞代谢流分析，研究eIF5A2调控细胞代谢重编程的分子机制；3）研究eIF5A2调控细胞代谢重编程是否HIF1a依赖？本研究将结合分子生物学和代谢组学技术研究eIF5A2代谢调控，深入探索食管癌发生、发展中代谢重编程发生的分子机制，为临床研究提供可能的治疗靶点。

我们的研究表明缺氧环境下可诱导食管鳞癌细胞中EIF5A2的表达，同时细胞中EIF5A2的表达可以上调HIF1α的表达；我们发现缺氧、HIF1α和EIF5A2三者之间存在相互调控的关系。EMSA结果和染色质免疫共沉淀实验结果发现EIF5A2通过结合HIF1α启动子区域调控HIF1α的表达， 我们并将该区域定位；临床分析发现EIF5A2与食管癌的转移和不良预后显著相关。体外、体内功能实验发现EIF5A2促进食管鳞状细胞癌的侵袭转移，并参与血管生成；EIF5A2引起了细胞糖代谢和脂质代谢的变化：敲低EIF5A2时葡萄糖摄入减少，乳酸生成减少，糖酵解通路上的几个关键酶也发生了变化。采用同位素标记的培养液进行细胞培养，然后做LC-MS/MS质谱分析,发现代谢产物如Glucose-6-phosphate（G6P）, citrate和malate发生了变化。采用质谱进行非靶向脂质分析，发现EIF5A2引起脂质代谢产物改变。