F-box proteins: deregulated cell cycle control and proteolysis in cancer

描述(由申请人提供)：泛素依赖的蛋白质分解确保特定的蛋白质功能在正确的时间、正确的位置和单向的方式被关闭。许多细胞周期调节蛋白的降解是由两类泛素连接酶控制的：SCF(Skp1-cull-F-box Protein)复合体和后期促进复合体/环体(APC/C)。在人类中有68个SCF连接酶，每个连接酶都有一个不同的F-box蛋白亚基，通过直接将底物招募到连接酶的其余部分，并最终招募到泛素结合酶来提供特异性。尽管有大量的F-box蛋白，但只有三种人类干细胞因子泛素连接酶(SCFskp2、SCF？trcp和SCFFbw7)具有成熟的功能和底物，其中许多参与细胞周期调控(如CDc25A、Cyclin E、Emi1、p21、p27和Wee1)。鉴于细胞周期机制的关键作用，细胞周期调节器的蛋白分解改变显然是癌细胞典型的不受限制的增殖的决定因素。值得注意的是，在三个特征的F-box蛋白中，Skp2是原癌基因的产物，Fbw7是肿瘤抑制因子，而？Trcp的过表达有助于转化，至少在某些上皮组织中是这样。在最初的九年中，CA76584支持阐明三种泛素连接酶复合体(SCFSkp2、SCF？Trcp和APC/CCdh1)通过降解调节CDKs活性的与癌症相关的底物来控制细胞周期进展的分子和细胞机制。此外，还揭示了发生在癌症中的这些通路的腐败。新的初步研究表明，高水平的F-box蛋白Emi1和Emi2与人类癌症中Skp2的稳定性增加有关，并表明Emi1和Emi2作为癌蛋白发挥作用。基于这些结果，CA76584第三周期下提出的新目标集中在泛素连接酶/细胞周期网络的新一层解除调节及其与癌症的关系：确定EmIL和Emi2在肿瘤中的表达是否解除调节，并研究癌细胞中Skp2稳定性的解除调节机制(目标1)；利用组织培养系统和体内实验研究Emi2在癌症发展中的作用(目标2)；研究Emi2在癌细胞中的细胞周期功能并确定其具有生物意义的底物(目标3)。随着泛素介导的细胞周期调节蛋白分解机制的揭开，该团队致力于将其基础研究成果与对恶性转化的理解相结合。

DESCRIPTION (provided by applicant): Ubiquitin-dependent proteolysis ensures that specific protein functions are turned off at the right time, in the right place and in a unidirectional fashion. The degradation of many cell cycle regulatory proteins is controlled by two classes of ubiquitin ligases: the SCF (Skp1-Cull-F-box protein) complexes and the Anaphase Promoting Complex/Cyclosome (APC/C). In humans there are sixty-eight SCF ligases, each characterized by a different F-box protein subunit that provides specificity by directly recruiting the substrate to the rest of the ligase and, ultimately, to the ubiquitin conjugating enzyme. Despite the large number of F-box proteins, only three human SCF ubiquitin ligases (SCFskp2, SCF?Trcp and SCFFbw7) have well- established functions and substrates, many of which are involved in cell cycle control (e.g., Cdc25A, cyclin E, Emi1, p21, p27 and Wee1). Given the crucial function of the cell cycle machinery, altered proteolysis of cell cycle regulators is clearly a contributing determinant of the unrestrained proliferation typical of cancer cells. Significantly, of the three characterized F-box proteins, Skp2 is the product of a proto-oncogene, Fbw7 is a tumor suppressor, and overexpression of ?Trcp contributes to transformation, at least in certain epithelial tissues. During the first nine years, CA76584 supported the elucidation of the molecular and cellular mechanisms by which three ubiquitin ligase complexes (SCFSkp2, SCF?Trcp and APC/CCdh1) control cell cycle progression through the degradation of cancer-relevant substrates that regulate the activity of CDKs. Furthermore, the corruption of these pathways occurring in cancer was revealed. Novel preliminary studies show that high levels of the F-box proteins Emi1 and Emi2 correlate with an increased stability of Skp2 in human cancers, and suggest that Emi1 and Emi2 function as oncoproteins. Based on these results, the new aims proposed under the third cycle of CA76584 are focused on a new tier of deregulation of the ubiquitin ligase/cell cycle network and its involvement in cancer: To determine whether the expression of Emil and Emi2 is deregulated in tumors and to investigate the mechanisms deregulating Skp2 stability in cancer cells (Aim 1); To study the contribution of Emi2 to cancer development using tissue culture systems and in vivo experiments (Aim 2); To study the cell cycle functions of Emi2 in cancer cells and to identify its biologically significant substrates (Aim 3). As the mechanisms of the ubiquitin-mediated proteolysis of cell cycle regulators are unraveled, this team is committed to the integration of its basic research results with an understanding of malignant transformation.