研究白藜芦醇(resveratrol,Res)对内质网应激途径细胞凋亡和糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)/Tau蛋白磷酸化作用的影响。体外原代培养神经元细胞,采用衣霉素(tunicamycin,TM)建立内质网应激模型,Western blo...研究白藜芦醇(resveratrol,Res)对内质网应激途径细胞凋亡和糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)/Tau蛋白磷酸化作用的影响。体外原代培养神经元细胞,采用衣霉素(tunicamycin,TM)建立内质网应激模型,Western blot法检测内质网分子伴侣蛋白葡萄糖调节蛋白78(glucose-regulated protein 78,GRP78)、未折叠蛋白反应相关的肌醇需要酶1α(inositol-requiring enzyme 1α,IRE1α)的Ser724磷酸化、剪接的X盒结合蛋白1(spliced form of X-box binding protein 1s,XBP1s)表达、GSK-3β的Ser9和Tau蛋白的Ser396磷酸化水平。生物化学方法分析细胞质中半胱天冬酶-12(Caspase-12)和半胱天冬酶-3(Caspase-3)活性、细胞凋亡水平。结果显示,TM能够诱导内质网应激作用,导致神经元细胞GSK-3β的活化和Tau蛋白磷酸化水平升高(P<0.01)、神经元细胞经内质网途径凋亡(P<0.05)。与内质网应激抑制剂4-苯基丁酸结果相似,Res组显著降低了GRP78的表达(P<0.01)、降低了IRE1α-XBP1通路的活性(P<0.01)。Res可以减缓TM诱导的内质网途径细胞凋亡级联反应中Caspase-12和Caspase-3的活性(P<0.01)。Res抑制了TM诱导的GSK-3β的Ser9位点磷酸化水平和Tau蛋白Ser396位点的磷酸化水平(P<0.01)。结果表明,Res能够降低TM诱导的IRE1α-XBP1途径内质网应激作用、GSK-3β的活性及Tau蛋白发生磷酸化水平,减弱神经元细胞经内质网途径凋亡的级联反应作用。展开更多
目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GS...目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GSK-3β在102例宫颈癌、52例高级别鳞状上皮内病变(high-grade squamous intraepithelial lesion,HSIL)、46例低级别鳞状上皮内病变(low-grade squamous intraepithelial lesion,LSIL)及40例慢性宫颈炎组织中的表达情况,分析三者的表达与宫颈癌患者临床病理特征的关系,分析三者之间的相关性,COX比例风险模型分析影响宫颈癌患者预后的影响因素。结果随着宫颈病变进展,KIF11、β-catenin阳性率逐渐升高,GSK-3β阳性率逐渐减低(P<0.05)。KIF11、β-catenin、GSK-3β在宫颈癌组织中的阳性表达在国际妇产科联盟(International Federation of Gynecology and Obstetrics,FIGO)分期、分化程度、淋巴结转移方面比较,差异有统计学意义(P<0.05),但在不同年龄及病理类型间比较,差异无统计学意义(P>0.05)。宫颈癌患者组织中KIF11与β-catenin的表达呈正相关(r=0.461,P<0.05),β-catenin与GSK-3β的表达呈负相关(r=-0.692,P<0.05),KIF11与GSK-3β的表达呈负相关(r=-0.336,P<0.05)。KIF11、β-catenin阳性表达患者的平均生存时间短于阴性表达患者,GSK-3β阳性表达患者的平均生存时间长于阴性表达患者。COX回归分析显示,FIGO分期、淋巴结转移、KIF11、β-catenin为宫颈癌患者预后的独立危险因素,GSK-3β为独立保护因素。结论KIF11、β-catenin、GSK-3β在宫颈癌患者组织中异常表达,KIF11可能参与宫颈癌发生、发展中Wnt/β-catenin通路的调节,三者联合检测可为宫颈癌的诊断及预后提供新的参考。展开更多
NORE1A (RASSF5) is a tumor suppressor of the RASSF family that is often down-regulated in human tumors. NORE1A has multiple roles in controlling cellular homeostasis, one of them being regulating levels of β-catenin ...NORE1A (RASSF5) is a tumor suppressor of the RASSF family that is often down-regulated in human tumors. NORE1A has multiple roles in controlling cellular homeostasis, one of them being regulating levels of β-catenin by binding and modulating the ubiquitin ligase substrate recognition factor β-TrCP. β-catenin is a major executor of the Wnt pathway. The ubiquitin SCF-β-TrCP ligase complex acts on a phospho-degron site in β-catenin that can be phosphorylated by GSK-3β. We now show that in addition to binding β-TrCP, NORE1A also promotes the phosphorylation of the β-catenin phospho-degron by complexing with the kinase GSK-3β. Indeed, NORE1A enhances the formation of a GSK-3β/β-TrCP complex. A structural mutant of NORE1A that retains β-TrCP binding but will no longer interact with GSK-3β inhibits the β-catenin degrading action of NORE1A. The GSK-3β interaction with NORE1A plays an important role in the biology of NORE1A as a GSK-3β inhibitor blocks NORE1A induced senescence. Thus, we identify a new role for the tumor suppressor NORE1A: The regulation of GSK-3β. GSK-3β has many other substrates including multiple transcription factors and co-activators such as p53 and the Hippo component TAZ. The work implies that NORE1A may be able to influence all of them via this new kinase scaffolding interaction.展开更多
文摘研究白藜芦醇(resveratrol,Res)对内质网应激途径细胞凋亡和糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)/Tau蛋白磷酸化作用的影响。体外原代培养神经元细胞,采用衣霉素(tunicamycin,TM)建立内质网应激模型,Western blot法检测内质网分子伴侣蛋白葡萄糖调节蛋白78(glucose-regulated protein 78,GRP78)、未折叠蛋白反应相关的肌醇需要酶1α(inositol-requiring enzyme 1α,IRE1α)的Ser724磷酸化、剪接的X盒结合蛋白1(spliced form of X-box binding protein 1s,XBP1s)表达、GSK-3β的Ser9和Tau蛋白的Ser396磷酸化水平。生物化学方法分析细胞质中半胱天冬酶-12(Caspase-12)和半胱天冬酶-3(Caspase-3)活性、细胞凋亡水平。结果显示,TM能够诱导内质网应激作用,导致神经元细胞GSK-3β的活化和Tau蛋白磷酸化水平升高(P<0.01)、神经元细胞经内质网途径凋亡(P<0.05)。与内质网应激抑制剂4-苯基丁酸结果相似,Res组显著降低了GRP78的表达(P<0.01)、降低了IRE1α-XBP1通路的活性(P<0.01)。Res可以减缓TM诱导的内质网途径细胞凋亡级联反应中Caspase-12和Caspase-3的活性(P<0.01)。Res抑制了TM诱导的GSK-3β的Ser9位点磷酸化水平和Tau蛋白Ser396位点的磷酸化水平(P<0.01)。结果表明,Res能够降低TM诱导的IRE1α-XBP1途径内质网应激作用、GSK-3β的活性及Tau蛋白发生磷酸化水平,减弱神经元细胞经内质网途径凋亡的级联反应作用。
文摘目的探讨驱动蛋白家族成员11(kinesin family member 11,KIF11)、β-连环蛋白(β-catenin)、糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)在宫颈癌中的表达情况及临床意义。方法采用免疫组化法检测KIF11、β-catenin、GSK-3β在102例宫颈癌、52例高级别鳞状上皮内病变(high-grade squamous intraepithelial lesion,HSIL)、46例低级别鳞状上皮内病变(low-grade squamous intraepithelial lesion,LSIL)及40例慢性宫颈炎组织中的表达情况,分析三者的表达与宫颈癌患者临床病理特征的关系,分析三者之间的相关性,COX比例风险模型分析影响宫颈癌患者预后的影响因素。结果随着宫颈病变进展,KIF11、β-catenin阳性率逐渐升高,GSK-3β阳性率逐渐减低(P<0.05)。KIF11、β-catenin、GSK-3β在宫颈癌组织中的阳性表达在国际妇产科联盟(International Federation of Gynecology and Obstetrics,FIGO)分期、分化程度、淋巴结转移方面比较,差异有统计学意义(P<0.05),但在不同年龄及病理类型间比较,差异无统计学意义(P>0.05)。宫颈癌患者组织中KIF11与β-catenin的表达呈正相关(r=0.461,P<0.05),β-catenin与GSK-3β的表达呈负相关(r=-0.692,P<0.05),KIF11与GSK-3β的表达呈负相关(r=-0.336,P<0.05)。KIF11、β-catenin阳性表达患者的平均生存时间短于阴性表达患者,GSK-3β阳性表达患者的平均生存时间长于阴性表达患者。COX回归分析显示,FIGO分期、淋巴结转移、KIF11、β-catenin为宫颈癌患者预后的独立危险因素,GSK-3β为独立保护因素。结论KIF11、β-catenin、GSK-3β在宫颈癌患者组织中异常表达,KIF11可能参与宫颈癌发生、发展中Wnt/β-catenin通路的调节,三者联合检测可为宫颈癌的诊断及预后提供新的参考。
文摘NORE1A (RASSF5) is a tumor suppressor of the RASSF family that is often down-regulated in human tumors. NORE1A has multiple roles in controlling cellular homeostasis, one of them being regulating levels of β-catenin by binding and modulating the ubiquitin ligase substrate recognition factor β-TrCP. β-catenin is a major executor of the Wnt pathway. The ubiquitin SCF-β-TrCP ligase complex acts on a phospho-degron site in β-catenin that can be phosphorylated by GSK-3β. We now show that in addition to binding β-TrCP, NORE1A also promotes the phosphorylation of the β-catenin phospho-degron by complexing with the kinase GSK-3β. Indeed, NORE1A enhances the formation of a GSK-3β/β-TrCP complex. A structural mutant of NORE1A that retains β-TrCP binding but will no longer interact with GSK-3β inhibits the β-catenin degrading action of NORE1A. The GSK-3β interaction with NORE1A plays an important role in the biology of NORE1A as a GSK-3β inhibitor blocks NORE1A induced senescence. Thus, we identify a new role for the tumor suppressor NORE1A: The regulation of GSK-3β. GSK-3β has many other substrates including multiple transcription factors and co-activators such as p53 and the Hippo component TAZ. The work implies that NORE1A may be able to influence all of them via this new kinase scaffolding interaction.
