p33ING1b核定位信号肽入核引导作用及其融合蛋白表达纯化的研究
发布时间：2018-08-22 16:09
【摘要】：[目的]生长抑制因子基因1(Inhibitor of growth gene 1,ING1)定位于人类染色体13q33-34,是1996年发现的一个生长抑制基因。其主要生理功能为调节细胞周期,抑制细胞增殖,参与细胞衰老和凋亡调控,并在维持基因组稳定方面发挥重要作用。p33~(ING1b)是其最主要的编码蛋白。以往的研究证实多数恶性肿瘤细胞中NG1基因突变率很低;其mRNA转录丰度及p33~(ING1b)蛋白表达水平目前各家研究结果不一,尚无定论,但较一致的发现是胶质瘤细胞中有p33~(ING1b)蛋白的亚细胞定位异常,即从正常情况下定位于胞核变为定位于胞浆,这可能是导致胶质瘤发生、发展的重要因素。一般情况下细胞浆向细胞核内的蛋白转运通过Importin α/β蛋白转运系统实现,Importin α作为适配器既与被运载蛋白的核定位信号肽(Nuclear localization signal;NLS)结合,又与Importin β结合,形成三联转运复合物,并通过importin β与核孔复合体(Nuclear-pore complex;NPC)作用,启动一系列入核转运机制。然而,生理情况下p33~(ING1b蛋白的入核转运机制怎样?是什么原因导致了胶质瘤细胞中p33~(ING1b)蛋白的亚细胞定位异常?均尚不清楚,有必要作深入系统的探讨。 为此,本研究以p33~(ING1b)蛋白核定位信号肽(p33~(ING1b)NLS)的功能研究入手,采用一系列分子生物学实验方法,初步探讨了生理情况下p33~(NG1b)NLS在p33~(ING1b)入核转运过程中的作用。旨在为进一步深入探讨p33~(ING1b)蛋白正常入核机制及其在胶质瘤细胞中入核失败的原因提供重要的线索及有用的研究工具。 [方法]在本研究第一部分,我们首先应用逆转录PCR(Reverse transcript
[Abstract]:[objective] the growth suppressor gene 1 (Inhibitor of growth gene 1 (ING1) was located on human chromosome 13q33-34. It was identified as a growth suppressor gene in 1996. Its main physiological functions are to regulate cell cycle, inhibit cell proliferation, participate in the regulation of cell senescence and apoptosis, and play an important role in maintaining genomic stability. P33 ~ (ING1b) is the most important coding protein. Previous studies have confirmed that the mutation rate of NG1 gene is very low in most malignant tumor cells, and its mRNA transcription abundance and the expression level of p33 ~ (ING1b) protein are different. However, it was found that the subcellular localization of p33 ~ (ING1b) protein in glioma cells was abnormal, that is, the localization from nucleus to cytoplasm of glioma cells was normal, which may be an important factor leading to the occurrence and development of glioma. In general, the translocation of proteins from cytoplasm to nucleus is realized by Importin 伪 / 尾 protein transport system. Importin 伪, as an adapter, binds to the nuclear localization signal peptide (Nuclear localization signalling peptide (Nuclear localization signaln NLS) of the carrying protein and binds to Importin 尾 to form a triplex transport complex. A series of nuclear transport mechanisms were initiated by the interaction of importin 尾 with Nuclear-pore complex. However, what is the nuclear transport mechanism of p33 ~ (ING1b) protein under physiological conditions? What causes subcellular localization of p33 ~ (ING1b) protein in glioma cells? All are not clear, it is necessary to make a thorough and systematic discussion. Therefore, the function of p33 ~ (ING1b) nuclear localization signal peptide (p33 ~ (ING1b) NLS) was studied, and a series of molecular biological methods were used to explore the role of p33 ~ (NG1b) NLS in the nuclear transport of p33 ~ (ING1b) under physiological conditions. The aim of this study is to provide important clues and useful tools for further study on the mechanism of p33- (ING1b) normal nucleation and the failure of p33- (ING1b) protein entry in glioma cells. [methods] in the first part of this study, we first applied reverse transcription PCR (Reverse transcript
【学位授予单位】：天津医科大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R346
本文编号：2197623
[Abstract]:[objective] the growth suppressor gene 1 (Inhibitor of growth gene 1 (ING1) was located on human chromosome 13q33-34. It was identified as a growth suppressor gene in 1996. Its main physiological functions are to regulate cell cycle, inhibit cell proliferation, participate in the regulation of cell senescence and apoptosis, and play an important role in maintaining genomic stability. P33 ~ (ING1b) is the most important coding protein. Previous studies have confirmed that the mutation rate of NG1 gene is very low in most malignant tumor cells, and its mRNA transcription abundance and the expression level of p33 ~ (ING1b) protein are different. However, it was found that the subcellular localization of p33 ~ (ING1b) protein in glioma cells was abnormal, that is, the localization from nucleus to cytoplasm of glioma cells was normal, which may be an important factor leading to the occurrence and development of glioma. In general, the translocation of proteins from cytoplasm to nucleus is realized by Importin 伪 / 尾 protein transport system. Importin 伪, as an adapter, binds to the nuclear localization signal peptide (Nuclear localization signalling peptide (Nuclear localization signaln NLS) of the carrying protein and binds to Importin 尾 to form a triplex transport complex. A series of nuclear transport mechanisms were initiated by the interaction of importin 尾 with Nuclear-pore complex. However, what is the nuclear transport mechanism of p33 ~ (ING1b) protein under physiological conditions? What causes subcellular localization of p33 ~ (ING1b) protein in glioma cells? All are not clear, it is necessary to make a thorough and systematic discussion. Therefore, the function of p33 ~ (ING1b) nuclear localization signal peptide (p33 ~ (ING1b) NLS) was studied, and a series of molecular biological methods were used to explore the role of p33 ~ (NG1b) NLS in the nuclear transport of p33 ~ (ING1b) under physiological conditions. The aim of this study is to provide important clues and useful tools for further study on the mechanism of p33- (ING1b) normal nucleation and the failure of p33- (ING1b) protein entry in glioma cells. [methods] in the first part of this study, we first applied reverse transcription PCR (Reverse transcript
【学位授予单位】：天津医科大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R346
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，本文编号：2197623
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