胃癌细胞中蛋白酶体介导的小G蛋白Rif降解及IncRNA表达谱

发布时间：2018-11-17 14:12

【摘要】：Rif是Rho小G蛋白家族成员,可以调控细胞骨架的结构,其活性通过结合GDP或GTP进行调控。近年来研究发现Rho小G蛋白的活性也可以通过泛素化介导的蛋白酶体降解来调控,然而,Rif的活性是否可以通过蛋白酶体降解进行调节至今还没有相关的研究。本研究发现Rif与泛素化连接酶SCF复合物的支架蛋白Cullin1存在相互作用关系,并可以改变SCF复合物调控蛋白NEDD8的细胞内定位,可能是一个SCF复合物的底物。同时,实验结果证实Rif是一个半衰期短的蛋白,其表达水平受到Cullin1表达的负调控。通过体内体外蛋白酶体降解实验,进一步证实Cullin1可以介导Rif在蛋白酶体中降解。在胃癌细胞中,抑制Rif蛋白表达会导致细胞周期抑制蛋白p27的表达上调,表明蛋白酶体介导的Rif降解可能有助于抑制胃癌细胞增殖。通过基因芯片检测手段,对胃癌细胞DNA复制期(S期)特异mRNA+lncRNA的表达谱进行分析,找出了胃癌细胞相对于正常胃上皮细胞在S期表达谱的差异。我们的研究结果首次揭示了蛋白酶体介导的Rif蛋白降解对胃癌的可能调控作用机制,筛选出了胃癌细胞DNA复制期特异的lncRNA及mRNA组合,为后续详细研究胃癌细胞的DNA复制调控奠定了基础。
[Abstract]:Rif is a member of small G protein family of Rho, which can regulate the structure of cytoskeleton and its activity is regulated by binding to GDP or GTP. In recent years, it has been found that the activity of Rho small G protein can also be regulated by ubiquitin mediated proteasome degradation. However, whether the activity of Rif can be regulated by proteasome degradation has not been studied. In this study, we found that Rif interacts with the scaffold protein Cullin1 of ubiquitin ligase SCF complex, and it can change the intracellular localization of SCF complex regulatory protein NEDD8, which may be a substrate of SCF complex. At the same time, the results showed that Rif is a short half-life protein, and its expression level is negatively regulated by Cullin1 expression. Through in vitro and in vivo proteasome degradation experiments, further confirmed that Cullin1 can mediate the degradation of Rif in proteasome. In gastric cancer cells, inhibition of Rif protein expression leads to up-regulation of cell cycle inhibitor p27, suggesting that proteasome mediated Rif degradation may contribute to the inhibition of gastric cancer cell proliferation. The expression profiles of DNA specific mRNA lncRNA in gastric cancer cells during DNA replication phase (S phase) were analyzed by gene chip analysis. The difference of expression profiles between gastric cancer cells and normal gastric epithelial cells in S phase was found out. Our results for the first time revealed the possible regulatory mechanism of proteasome mediated Rif degradation on gastric cancer and screened out specific lncRNA and mRNA combinations in the DNA replication phase of gastric cancer cells. It lays a foundation for further detailed study on the regulation of DNA replication in gastric cancer cells.
【学位授予单位】：内蒙古大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.2

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