小分子量G蛋白Rap2及其效应因子调控细胞运动与肿瘤细胞侵袭和转移的研究

Ras superfamily proteins play important roles in cells through mediating a variety of extracellular signals including growth factors and cytokines, and regulate essential life activities of the cells. Rap2 is a member of Rap subfamily in this superfamily, but the roles for Rap2 and its specific effectors and signaling pathway is still remain to be clearified. Our previous results have shown that Rap2 activity was largely increased in renal cancer tissues as compared to the normal ajancent tissues, and it was closely related to increased cell migration and invasion. To elucidate the correlations of Rap2 with cell migration and cancer metastasis, and Rap2 specific effecors and signaling pathway involved in cell migration and invasion, we are planing to use cellular and nude mice model to clearify the Rap2 specific sinaling pathway involved in cell migration and invasion by GST-pull down, Co-IP, cell time lapse microscopy and GTPase activaty assay. Then we will analyze the cross talk between Rap2 and Rho pathways through expressing or knocking down of Rap2 downstream effectors. To identify the key kinases activation downsteam of Rap2 signaling pathway involved in cell migration and invasion,the activated kinase network will be compared between normal and cancer cells, cancer tissues and normal tissues using kinase substrate chip assay.Dicephering the corelation between Rap2 and cell migration and cancer invasion and the mechanism of signaling pathway will deepen our knowledge about function of the small G protein,therefore, provide useful cues for elucidating the mechanism of cancer cell migration and metastasis.

Ras超家族分子是一类重要的功能蛋白，介导生长因子、细胞因子等多种细胞外因子参与的信号通路，调节着细胞的基本生命活动。Rap2是该超家族中Rap家族成员，但其特异性效应因子及所调控的分子通路尚不清楚。本课题前期证实肿瘤组织Rap2活性明显升高，且与肿瘤细胞迁移及转移关系密切。为探讨Rap2与细胞运动及肿瘤转移的关系，解析所调控的特异性效应因子及信号通路，本研究拟从Rap2特异效应因子入手，采用GST-pull down、Co-IP、活细胞运动观察与Rap2活性分析等方法，并通过沉默其下游因子等方式，分析Rap2效应因子的作用及其与Rho信号通路关系。进而利用激酶底物芯片比较Rap2底物在正常细胞、肿瘤组织及癌旁组织中的活化水平，获取其诱导激活的关键激酶，剖析Rap2与细胞运动及肿瘤转移关系及其信号通路的活化机制，从而丰富对小分子G蛋白的功能认识，并揭示对肿瘤细胞迁移与转移的影响。