【作者】 周萍；

【导师】 李晓涛；

【作者基本信息】 华东师范大学 ， 生物化学与分子生物学， 2011， 硕士

【摘要】 REGγ,又称11S蛋白酶体激活因子,它不仅可以降解细胞周期蛋白激酶抑制因子p21,在细胞周期调控中发挥重要作用,而且还能够降解抑癌基因表达产物p53,参与肿瘤的发生发展过程。TGFβ,即转化生长因子β,能够上调细胞内的p21和p15,抑制细胞生长,也参与细胞周期的调控。P53作为肿瘤发生的第一大因素,和它下游基因一起参与肿瘤的发生发展过程。由此可见,它们均是参与细胞周期调控的重要因子。然而,到目前为止,关于REGγ和TGFβ, REGγ和p53下游基因关系的研究还未见报道。在关于REGγ和TGFβ的研究中,我们发现TGFβ能够抑制REGγ的表达,REGγ和TGFβ下游的细胞周期蛋白激酶抑制因子p21和p15之间存在相互作用,这些结果表明TGFβ诱导的细胞生长抑制受REGγ的调控,TGFβ诱导其下游基因p21和p15的活化与REGγ功能抑制有关。该研究次证实了REGγ和TGFβ之间确实存在相互作用。P53作为细胞内的重要抑癌基因之一,其下游基因也与肿瘤的发生发展密切相关。通过已有的基因芯片数据库的元分析,我们在结肠、甲状腺、肝癌和肺癌四种肿瘤组织里筛选出了一些和REGγ表达具有相关性的基因,主要包括：p53下游基因,与myc相关的基因,和一些其他癌基因。采用分子生物学实验,我们再次证明了这些基因中70%和REGγ呈现明显相关性。我们重点集中在p53下游基因,研究发现：29个p53下游基因与REGγ相关,相关性基因高达80%。其中,备受大家关注的重要基因BAX、PTEN、GADD45, CDK2,均被发现和REGγ的表达呈现显著相关性。而且,部分基因与REGγ的相关性存在肿瘤特异性,部分基因REGγ的相关性受p53活性的影响。该部分研究不仅证实了REGγ和多种肿瘤组织中p53下游基因的表达具有相关性,而且暗示了REGγ与p53下游基因可能存在相互作用,这为进一步探索REGγ的靶蛋白提供了新思路。REGγ能够增加MDM2介导p53的降解作用,但是REGγ和p53是否存在直接的相互作用尚不清楚。我们通过实验证明了REGγ和p53可以直接相互作用,相互作用位点在100-290AA这个片段。该结果表明除了通过MDM2, REGγ或许能够直接介导p53降解,因此细胞内可能存在p53降解的新机制。但是,REGγy和p53相互作用的更精确位点需要进一步研究。本课题初步探讨了REGγ和TGFβ,p53及其下游基因的关系,证实了：TGFβ诱导的细胞生长抑制受REGγ的调控,80%的p53下游基因与REGγ存在相互作用,REGγ能够直接与p53相互结合,这些研究不仅有助于探索REGγ新的生物学功能,还为进一步揭示TGFβ信号通路的新作用机制和肿瘤发生机理奠定了理论基础。更多还原

【Abstract】 REGy, also known as 11S proteasome activator, plays an important role in cell cycle regulation through the degradation of p21(CDKI) mediated by REGy, and that it is concerned with the tumorigenesis through the degradation of p53. TGF(3, that is, transforming growth factor beta, can inhibit the cell growth through the up regulation of p15 and p21.It is also involved in cell cycle regulation. p53, as a most important factor of the tumorigenesis, exerts its function together with its downstream genes. Obviously, they are all important regulators of the cell cycle. Up to now, there is no report about the relation between REGγand TGFβ, REGy and P53 downstream genes.In the research of the REGγand TGFβ,we found that the expression of REGy can be inhibited by TGF(3, and that REGy has relation with the p21 and p15,which are downstream genes of TGFβ. All these data reveal that the cell growth induced by TGF(3 is regulated by REGy. The high expression of p21 and p15 are involved in the REGy inhibition. This study demonstrates that REGγhas some relation with TGFβ.As one of the most important tumor suppressor genes, p53 is combined with its downstream genes to play an important role in the tumorigenesis. Through the meta-analysis of the publicly microarray database, we find some genes whose expression has some relation with REGy in four tumor tissues, including colon, thyroid, liver and lung. There are three kinds of genes:p53 downstream genes, myc genes and some others.We focused our attention on the p53 downstream genes and find that 29 of the p53 downstream gene are related with REGy. That is,80% of the p53 downstream genes are consistent with the computer analysis. Some of the attention-attracting genes, such as BAX, PTEN, GADD45, CDK2, are related with REGy expression. Some of these genes have the tumor specificity, and some are affected by p53 activation. This study demonstrates that REGγhave some relation with the p53 downstream genes. Maybe there are some interaction between them, which provide a new idea for the exploration of the new REGγtarget genes.REGγcan degrade p53 by enhancing the MDM2 mediated ubiquitination. But we have no idea about the direct interation between REGγand p53. We have proved that they can interact with each other, and the binding site is the 100-290AA subunit of p53.This new result has provided us that REGγmay degrade the p53 directly, except the MDM2 mediated method. But the specific binding site need our further study.In this project, we discuss the relation of REGγand TGFβ, p53 and its dowmstream genes. We get the results that TGFβinduced cell growth is regulated by REGγ,80% of the computer analysis p53 genes are related with REGy expression, and that REGy can interact with p53 directly. These new findings help us to explore the unknown biological function of REGy and pave the way for revealing new mechanism of TGFβsignaling path way and tumorigenesis.更多还原