Rap2信号通路调控肾癌细胞迁移及侵袭的机制

发布时间：2018-04-25 05:36

  本文选题：Rap2 + 细胞迁移　； 参考：《哈尔滨工业大学》2014年硕士论文

【摘要】：Rap2属于Ras家族Rap亚族，具有Rap2A、Rap2B和Rap2C三种异构体，调节多种细胞生命活动。由于Rap2与Ras家族其他成员具有相似的结构，因此Rap2能够结合多种Ras家族共有的相互作用蛋白，另外Rap2还可能通过与特异性相互作用蛋白的结合的方式完成信号转导。目前已知的人源Rap2相互作用蛋白有PARG、TNIK、MINK和MAP4K4四种，，但是对其在肿瘤发生及发展中的机制尚无阐述。 本课题组前期研究发现活性化Rap2在肾癌组织中的水平远高于癌旁正常组织，且Rap2的高表达能够促进细胞的运动和迁移，然而其分子机制都尚待阐明。本文首先通过GST-Pull down检测了肾癌细胞系中Rap2的活性化水平，并通过半定量PCR检测了对Rap2活性具有调控作用的Rap1GAP1和RasGEF1B在不同肾癌细胞系的表达水平，结果发现肾癌细胞系中Rap2的活性化水平与上述调控因子的差异表达相关。为了研究Rap2调控细胞侵袭和迁移的作用通路，通过划痕实验和Transwell实验对具有高活性化水平Rap2的肾癌细胞系进行了针对TNIK和MAP4K4的RNA干扰和迁移及侵袭分析，结果显示：Rap2通过TNIK和MAP4K4促进肾癌细胞的迁移及侵袭能力。已有研究报道FAK正调控细胞的铺展和运动，本文通过免疫荧光和western blot的实验证明Rap2及其激活突变体的过表达可上调FAK的表达，而DN突变体Rap2-17N则对FAK表达无显著调控作用。本研究增进了对于Rap2在肾癌细胞系中高活性化的机制，以及Rap2调控细胞运动和侵袭的分子机制的了解。
[Abstract]:Rap2 belongs to the Rap subfamily of Ras family. It has three isomers Rap2An Rap2B and Rap2C and regulates many cell life activities. Because Rap2 has similar structure with other members of Ras family, Rap2 can bind to many kinds of interaction proteins shared by Ras family, and Rap2 may also complete signal transduction by binding with specific interacting proteins. At present, there are four known human Rap2 interaction proteins: Rap2 TNIKMIK and MAP4K4, but the mechanism of them in tumorigenesis and development has not been elucidated. Our previous study found that the level of activated Rap2 in renal cell carcinoma tissues is much higher than that in adjacent normal tissues, and the high expression of Rap2 can promote cell movement and migration, but its molecular mechanism remains to be clarified. In this study, the activation level of Rap2 was detected by GST-Pull down, and the expression of Rap1GAP1 and RasGEF1B, which regulate the activity of Rap2, was detected by semi-quantitative PCR in different RCC cell lines. The results showed that the activation level of Rap2 was related to the differential expression of these regulatory factors in renal cell carcinoma cell line. In order to study the pathway of Rap2 regulating cell invasion and migration, the RNA interference, migration and invasion of TNIK and MAP4K4 in renal cell lines with high Rap2 activity were analyzed by scratch test and Transwell assay. The results showed that: Rap2 promoted the migration and invasion of renal cancer cells through TNIK and MAP4K4. It has been reported that FAK is regulating the proliferation and movement of cells. The results of immunofluorescence and western blot have shown that the overexpression of Rap2 and its activated mutants can up-regulate the expression of FAK, while the Rap2-17N of DN has no significant effect on FAK expression. This study enhanced the understanding of the mechanism of high activation of Rap2 in renal cell line and the molecular mechanism of Rap2 regulating cell movement and invasion.
【学位授予单位】：哈尔滨工业大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R737.11

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1 蒋新农,周柔丽;肿瘤细胞粘附、迁移与转移的相关性[J];生物化学与生物物理进展;1998年05期

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