蛋白酶体激活因子REGγ对糖原合成酶激酶GSK3β蛋白活性及降解机制的研究

发布时间：2018-09-07 10:59

【摘要】：糖原合成酶激酶(Glycose synthase kinase 3, GSK3),是最初在糖原合成中发现的一个重要的丝氨酸／苏氨酸激酶,在细胞各种信号通路包括WNT通路,胰岛素敏感性调节,造血干细胞,细胞周期和凋亡,肿瘤细胞的存活中都扮演着重要的角色。GSK3具有两个由不同基因编码翻译的剪切体：GSK3α(51KD)和GSK3β(47kD)。llS调节蛋白复合物γ亚单位REGy(11S regulator complex gamma sub unit),是一类蛋白酶体激活因子,它能与20s蛋白酶体结合而参与泛素与ATP非依赖的蛋白降解。本文研究了蛋白酶体激活因子REGγ与糖原合成酶激酶GSK3p之间的相互作用。首先,我们发现蛋白酶体激活因子REGγ能结合并降解糖原合成酶激酶GSK3p。过量表达REGγ会增加GSK3β降解,敲低REGγ会增加内源GSK3p蛋白水平。REGγ基因敲除小鼠MEF细胞内GSK3β蛋白水平也会增加。另外,REGγ能影响GSK3β蛋白在细胞内的定位。同时我们探究了REGγ降解GSK3β的机制。然后,我们探究了REGγ与GSK3β之间的作用,所引起的细胞内生物学功能的变化。在REGγ敲低的细胞内,我们发现GSK3β蛋白水平上升,肿瘤抑制因子P53的Ser315位磷酸化修饰升高。已有报道表明GSK3β能磷酸化修饰p53的Ser315,而p53的Ser315磷酸化修饰能促进p53的转录活性,增加p53的泛素化降解。因此,REGγ可能通过GSK3β,间接参与P53的磷酸化修饰、转录活性及降解调控。同时我们发现GSK3β下游蛋白β-catenin蛋白水平降低。REGγ也有可能通过GSK3β间接调节β-catenin,从而调控β-catenin参与的肿瘤相关基因的转录。流式细胞仪FACS技术检测细胞周期和凋亡实验时,发现过量表达REGγ会抑制GSK3β诱导的细胞凋亡。综上所述我们主要研究了REGγ能结合并降解GSK3β蛋白,从而影响GSK3β下游蛋白的活性,生物功能和与之相关的肿瘤发生,并为以后研究GSK3β及其所参与的细胞信号通路提供一些线索。
[Abstract]:Glycose synthase kinase 3 (GSK3) is an important serine/threonine kinase first discovered in glycogen synthesis. GSK3 plays an important role in various cell signaling pathways including WNT pathway, insulin sensitivity regulation, hematopoietic stem cells, cell cycle and apoptosis, and tumor cell survival. There are two splicers encoded and translated by different genes: GSK3alpha (51KD) and GSK3beta (47kD). llS regulator complex gamma sub unit (REGy), a class of proteasome-activating factors that bind to 20S proteasome and participate in ubiquitin-and ATP-independent protein degradation. The interaction between live factor REG gamma and glycogen synthase kinase GSK3p.
First, we found that proteasome-activated factor REGgamma binds to and degrades glycogen synthase kinase GSK3p. Overexpression of REGgamma increases the degradation of GSK3p, and knockdown of REGgamma increases the level of endogenous GSK3p protein. At the same time, we explored the mechanism of REG GSK3 degradation.
We then explored the role of REG-gamma and GSK3-beta in the changes in cellular biological functions. In REG-gamma knocked-down cells, we found that GSK3-beta protein levels rose and the phosphorylation of tumor suppressor P53 Ser315 increased. It has been reported that GSK3-beta phosphorylated p53 Ser315, while p53 Ser315 phosphorylated p53. Therefore, REG-gamma may indirectly participate in the phosphorylation modification, transcriptional activity and degradation regulation of P53 through GSK3 beta. At the same time, we found that the level of downstream protein beta-catenin of GSK3 beta decreased. REG-gamma may also indirectly regulate beta-catenin through GSK3 beta, thereby regulating the participation of beta-catenin. Flow cytometry (FACS) was used to detect cell cycle and apoptosis. Overexpression of REGgamma inhibited GSK3 beta-induced apoptosis.
In summary, we mainly studied the binding and degradation of GSK3 beta protein by REGgamma, thereby affecting the activity, biological function and tumorigenesis of GSK3 beta downstream protein, and providing some clues for the future study of GSK3 beta and its involved cell signaling pathways.
【学位授予单位】：华东师范大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R341

【相似文献】