【作者】 吴晓玲；

【导师】 李勤喜；

【作者基本信息】 厦门大学 ， 生物化学与分子生物学， 2007， 硕士

【摘要】 体轴发育抑制因子Axin是一种多功能构架蛋白,在多条信号传导途径中发挥重要作用:在Wnt信号通路中,Axin作为负调控因子,下调β-catenin的水平;在JNK信号通路中,Axin与MEKK1或MEKK4相互结合,激活JNK,诱导细胞凋亡;在TGF-β信号通路中,Axin通过磷酸化Smad3增强转录活性;在p53信号通路中,Axin与p53,HIPK2形成三聚体复合物,调节p53的转录活性。这些信号途径对细胞的各种生理过程如细胞的生长,增殖,分化,癌变,凋亡及细胞周期起着重要的调控作用。PML是一个很重要的抑癌因子,最初是从急性早幼粒白血病(acute promyelocytic leukemia,APL)病人的细胞中,鉴定并克隆出来。细胞中,PML主要分布在nuclear body上。PML-NB的成分错综复杂,包含SUMO-1, Sp100, Sp140, CBP, BLM, Daxx, pRB, p53等多种重要的信号分子,可见PML-NB参与多条信号转导途径。在APL中,PML与RARα基因融合,融合蛋白与野生型的PML发生异源二聚化,引起PML的异常分布,破坏NB,导致肿瘤抑制,细胞增殖、分化、存活的紊乱。众所周知,Axin和PML都是抑癌因子,都参与p53诱导的凋亡,但两者之间的直接联系还不清楚。因此,本论文采用生物化学,分子生物学和细胞生物学的方法对Axin和PML之间的关系进行研究和阐述,初步揭示了PML在Axin诱导细胞死亡中的作用。本论文发现:( 1 ) UV刺激可促使Axin从细胞浆转移到细胞核,并定位于PML-NB。( 2 ) Axin可与PML相互作用。( 3 ) Axin诱导的p53的活化需要PML的参与,PML的显性负作用突变体及其siRNA都可强烈抑制Axin激活p53。( 4 ) PML和Axin均参与UV诱导的细胞死亡。( 5 ) Axin诱导的细胞死亡需要PML的参与,PML的显性负作用突变体及其siRNA可以抑制Axin引起的细胞死亡。综上所述,本论文发现,PML在UV引起的Axin介导的细胞死亡中发挥重要作用,对我们今后进一步认识Axin作为多功能构架蛋白,在多信号通路中的作用及生物学效应具有重要意义。更多还原

【Abstract】 Axin (Axis inhibitor) is a scaffold protein which plays important roles in multiple signal-transduction pathways. In the Wnt signaling pathway, Axin stimulates degradation ofβ-catenin by forming a large multimeric protein complex with APC, GSK-3βandβ-catenin. In JNK signaling, Axin activates JNK by directly binding to MEKK1 or MEKK4. In TGF-βsignaling, Axin enhances the transcriptional activity of Smad3. Besides, Axin regulates the transcriptional activity of p53 by forming a protein complex with p53 and HIPK2. These different signaling pathways control cellular processes, including cell proliferation, differentiation, transformation, apoptosis and cell cycle arrest.PML, an important tumor suppressor, was originally cloned and identified from APL patient. It locates on the PML-NB, which is a multimetric complex containing SUMO-1, Sp100, Sp140, CBP, BLM, Daxx, pRB, p53 and so on. In APL patients, PML fuses with RARα, and the resulting fusion protein dimerizes with PML and interfere with the formation of nuclear body, leading to the disorder of survival, proliferation, differentiation of normal cell and tumor suppression.It has been shown that both Axin and PML are tumor suppressors and are involved in p53-induced apoptosis, but the relationship between them is not clear. In this study, taking advantage of the methods of biochemistry, molecular biology and cell biology, I focused on the relationship between Axin and PML, and discovered that PML plays an important role in Axin induced cell death. (1) UV irradiation stimulates redistribution of Axin from cytoplasm to PML-NB in the nucleus; (2) Axin interacts with PML; (3) PML is required for Axin-mediated p53 activation, and this activation can be blocked by its dominate forms and siRNA; (4) Both PML and Axin are involved in UV-induced cell death; (5) PML is required for Axin-induced cell death, both dominant negative forms and siRNA of PML can block the process. In conclusion, all data give evidence of the effect of PML on UV-induced, Axin-mediated cell death, as sheds a new light on our understanding of Axin.更多还原