β-TrCP1和β-TrCP2降解的分子机制及其在细胞凋亡和自噬中生物学功能的差异研究

项目名称： β-TrCP1和β-TrCP2降解的分子机制及其在细胞凋亡和自噬中生物学功能的差异研究

项目编号： No.31470753

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 生物科学

项目作者： 赵永超

作者单位： 浙江大学

项目金额： 85万元

中文摘要： CRL泛素连接酶β-TrCP1/2通过降解一系列重要底物而在许多生物学进程中发挥着关键的调节作用。β-TrCP1/2与肿瘤发生密切相关，且具有明显的原癌基因特性，因此，可以作为潜在的肿瘤治疗靶点。但是，其自身的降解及两者的功能差异却非常不清楚。我们的预实验结果显示这两个蛋白代谢很快，提示两者的稳定性可能对其功能的发挥起着重要的调节作用。我们初步确定了这两个蛋白能互相降解对方。在两者高表达的乳腺癌细胞中沉默β-TrCP1诱导自噬而沉默β-TrCP2诱导凋亡，提示它们不是功能冗余的。本项目拟通过突变分析、生物信息分析等手段，系统阐明这两个蛋白的降解机制及其在细胞凋亡和自噬中的功能差异，并在裸鼠模型上研究两者的稳定性对肿瘤的生长和化疗药物敏感性的影响。另外，本项目也拟深入分析沉默β-TrCP1或β-TrCP2对相关信号通路的影响，并证明这两个酶对底物具有选择性，为两者成为肿瘤治疗的靶点打下基础。

中文关键词： β-TrCP1/2；蛋白质翻译后修饰；泛素化；蛋白质降解；肿瘤治疗靶点

英文摘要： CRLβ-TrCP1/2 (Cullin-RING Ligase) play a critical role in the regulation of a vast array of biological processes via the degradation of many key regulators. Accumulating data indicate that β-TrCP1/2 have important roles in tumorigenesis and possess mainly oncogenic characteristics. Thus, β-TrCP1/2 may serve as potential target(s) for anticancer therapy. However, how these two family members are degraded and whether they possess different biological functions are totally unknown. Our preliminary data showed that β-TrCP1/2 are fast turnover proteins, indicating that their stability is likely to play a critical role in their biological functions. We also found that β-TrCP1/2 target each other for degradation. More interestingly, β-TrCP1 knock-down induces autophagy, whereas β-TrCP2 knock-down induces apoptosis in β-TrCP1/2 activated breast cancer cells, suggesting that these two paralogues may be not functional redundant. In this proposed study, we plan to elucidate the molecular mechanism of β-TrCP1/2 degradation via approaches including gene mutation, bioinformatics, cellular and molecular biology. Moreover, we will determine how β-TrCP1/2 stability regulates tumor cell growth and anticancer drug sensitivity in both cell-based assays and xenograft mouse models. Finally, we will investigate the affected signaling pathways after knock-down of β-TrCP1 or β-TrCP2 and confirm that β-TrCP1 or β-TrCP2 indeed have preference for different substrates associated with autophagy or apoptosis. Taken together, our study will fully elucidate the molecular mechanism by which β-TrCP1/2 degrades each other and how they function differently in regulation of apoptosis and autophagy. Our study will provide additional piece of proof-of-concept evidence validating β-TrCP1/2 as promising anti-cancer targets.

英文关键词： β-TrCP1/2;Post-translational modification;Ubiquitination;Protein degradation;Anticancer target