hNUDC与Mpl和LIS-1复合体的形成及其双重功能的研究

项目名称： hNUDC与Mpl和LIS-1复合体的形成及其双重功能的研究

项目编号： No.31271230

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 生物科学

项目作者： 徐培林

作者单位： 中山大学

项目金额： 79万元

中文摘要： 早期的研究认为NUDC与LIS-1组成复合物,调控细胞有丝分裂和胞质分裂。然而本实验室近期研究结果表明人体NUDC与血小板生成素受体（Mpl）也有结合，且在体内、体外均有促进巨核细胞成熟的功能。进一步研究证明Mpl与hNUDC在细胞的内质网、高尔基、膜均共定位，并且Mpl可以介导hNUDC向胞外释放。 因此我们推测hNUDC/Mpl和hNUDC/LIS-1之间存在不同的亚细胞共定位以及不同的生物学功能。本项将运用双分子荧光互补法（BiFC）和荧光共振能量转移技术(FRET)在活体贴壁细胞和巨核细胞中同时直观hNUDC/Mpl和hNUDC/ LIS-1复合体的共定位。另外通过噬菌体展示、基因干扰等手段验证Mpl和LIS-1之间与hNUDC竞争结合的机制。还将通过基因沉默技术分析这三种蛋白在巨核细胞的成熟分化过程中的联系。本项目首次提出了hNUDC具有内源与外源双重功能。

中文关键词： 迁移蛋白；受体；蛋白相互作用；miR-19；癌症

英文摘要： The earlier studies have revealed that hNUDC forms the complex with LIS-1, a nuclea migration protein, involved in cell mitosis and cytokineses. However, other mechanistic studies by which hNUDC and Mpl interaction has been provided by our laboratory studies recently. hNUDC has in addition been shown to act as a second naturally ligand for thrombopoietin receptor (Mpl) and it plays a biological role in maturation of megakaryocyte cells in both in vitro and in vivo. Furthermore, recent studies suggest that the co-expression of hNUDC and Mpl effectively elevates hNUDC secretion, leading us to conclude a mechanism of Mpl-dependent secretory of hNUDC. We suggest that hNUDC/Mpl and hNUDC/LIS-1 may have different funtions and locat at different sucellular compartments. For further comfirm this idea, the present investigation was undertaken to elucidate the complex formation of hNUDC/Mpl or hNUDC/LIS-1 in living cells by direct visualization of the molecular interactions between proteins using BiFC and FRET. Moreoever, the applicability of BiFC system in conjunction with co-expression of the reference subcellular location markers allow detection of BiFC fusions in various subcellular compartments. Using phage-disply ELISA and small RNA interferon techeniques, the presented study will further detecte the competition b

英文关键词： hNUDC；Mpl；protein interaction；miR-194；cancer