乏氧微环境下蛋白激酶PAK4在胃癌侵袭转移中的作用及机制研究

The p21-actived kinase(PAK4) is a serine/threonine protein kinase involved in a variety of cellular processes including regulation of cytoskeletal reorganization and oncogenic transformation.The PAK4 plays a crucial role in cellular signal transduction. Using yeast two-hybrid screen, we identified a novel PAK4 binding protein,HIF-1α, which is playing an important role under hypoxia enviroment and is closely associated with epithelial mesenchymal transition(EMT) and vascular mimicry(VM).In our previous study we found that the expression of PAK4 was incresed and PAK4 was translocated into nucleus in gastric cancer cells under hypoxia enviroment，and we confirmed PAK4 binding to the HIF-1αby GST pull-down and Co-IP assays. Furthermore, silence of PAK4 upregulated E-cadherin and downregulated Twist1 protein expression. In addition, the expression of PAK4 was associated with VM in gastric cancer samples. So we presumed that the interaction of PAK4 and HIF-1αmay play an important role in gastric cancer inviasion and migration. The aim of this project is to demonstrate the mechanisms of PAK4 modulation under hypoxia and the interaction of PAK4 an HIF-1α,and the effects of PAK4 and HIF-1αinteraction on EMT, VM and metastasis in gastric cancer. The project will provide novel research ideas and targets for gastric cancer therapy.

PAK4是丝/苏氨酸激酶，参与调节细胞骨架重组和肿瘤转化等多种细胞活动，是癌细胞信号转导网络的枢纽。HIF-1α是我们用酵母双杂交技术筛选到的PAK4新的互作蛋白，在肿瘤乏氧微环境中起着枢纽作用且与EMT及血管生成拟态密切相关。我们发现乏氧可诱导胃癌细胞PAK4表达上调及入核增多，GST pull-down等实验显示PAK4与HIF-1α相互作用，且在乏氧条件下干扰PAK4可影响EMT始动标志E-cadherin 及关键转录因子Twist1的表达，同时在胃腺癌中PAK4的表达与血管生成拟态有关，提示在乏氧微环境下PAK4可能通过与 HIF-1α相互作用影响胃癌EMT和血管生成拟态。本课题旨在阐明胃癌乏氧微环境中PAK4的表达调控机制及与HIF-1α相互作用模式，并深入研究这种相互作用对胃癌EMT、血管生成拟态和侵袭转移的影响，为临床胃癌的综合治疗提供新的研究思路和治疗靶点。

PAK4是丝/苏氨酸激酶，参与调节细胞骨架重组和肿瘤转化等多种细胞活动，是癌细胞信号转导网络的枢纽。HIF-1α是我们用酵母双杂交技术筛选到的PAK4新的互作蛋白，在肿瘤乏氧微环境中起着枢纽作用且与EMT及血管生成拟态密切相关。我们发现在乏氧条件下PAK4-HIF-1a-MTA1三者形成复合体，进一步研究表明在胃癌细胞中PAK4上调MTA1和HIF-1a的表达，及PAK4与MTA1共同调控HIF-1a的信号通路。在三维培养条件下检测不同稳定转染细胞系对胃癌细胞形成VM管状能力的影响。并在基质胶中探讨PAK4对血管生成拟态的影响，以及对VM相关因子在蛋白水平和RNA 水平的影响。最终我们利用胃癌组织标本，进一步研究PAK4与VM相关因子在临床标本中的关系及与TNM分期的相关性。本课题旨在阐明胃癌乏氧微环境中PAK4的表达调控机制及与HIF-1α相互作用模式，并深入研究这种相互作用对胃癌EMT、血管生成拟态和侵袭转移的影响，为临床胃癌的综合治疗提供新的研究思路和治疗靶点。