BARD1与HUWE1在BRCA1依赖的乳腺癌发生发展中的作用及其机理研究

BRCA1 is an important tumor suppressor associated with breast cancer. As a key molecular, BRCA1 protein level is tightly regulated and turnover of BRCA1 protein directly affects its various functions and antitumor activities. It is known that BRCA1 can form a heterodimer with BARD1, which greatly increases the stability of each other. The underlying mechanism is largely unknown. Our previous results demonstrate that E3 ligase HUWE1 enhances BRCA1 ubiquitination and degradation and regulates BRCA1 activity ,acting as a negative regulator for BRCA1,plays critical role in regulation of BRCA1 in cell.Immunoprecipitation demonstrated that BARD1 not only readily interacted with HUWE1, but als overexpression of BARD1 resulted in the down regulation of HUWE1 protein.Moreover, overexpression of BARD1 abrogated the ubiquitination of HUWE1, suggesting that BARD1 may play an important role in regulating the stability as well as the E3 ligase activity of HUWE1 and thus affect HUWE1-dependent ubiquitination and degradation of BRCA1.Our results indicated that HUWE1-bound BARD1 is preferentially modified, most likely through Neddylation. In addition, BARD1 is shown to be modified by NEDD8 in vivo. This finding has important implication in our understanding of the molecular mechanism underlying the tumor suppression function of HUWE1 by BARD1. Our research revealed a new network between BRCA1, BARD1 and HUWE1 that is important in our understanding of the tumor suppression function of BRCA1/BARD1. Moreover, our study will also provide new model to elucidate the mechanisms on how ubiquitin E3 ligases are regulated by each other.But we should further investigate the role and the possible relationship in between BARD1 and HUWE1, what specific function of BARD1 neddylation, what specific network between BRCA1, BARD1 and HUWE1,that should be further stood.

BRCA1是一个重要的抑癌分子，其蛋白分子的稳定性对于其抑癌功能的发挥起到重要作用。BARD1能起到稳定BRCA1的作用，但其具体机制不详。在前期工作中我们发现了BRCA1的一个新的负调控因子-E3泛素化连接酶HUWE1，进一步证实BARD1不仅能与HUWE1相互作用，而且能够调节HUWE1的蛋白和泛素化水平，从而抑制HUWE1对BRCA1的降解。我们还发现与HUWE1结合的BARD1被NEDD8所修饰，此修饰对于BARD1发挥对HUWE1的调控可能起到重要作用。基于此，我们提出以下假设：BRCA1、HUWE1、BARD1 三者之间可形成复杂的网络调控模式，即BARD1被NEDD8修饰后，通过调节HUWE1而稳定BRCA1，此模式将为解析BRCA1/BARD1相互稳定的机制和阐明其抑癌功能提供了新的思路。本课题将以BARD1与HUWE1为切点，研究其在BRCA1依赖的乳腺癌发生发展中的机理

在前期工作中我们发现了BRCA1的一个新的负调控因子-E3泛素化连接酶HUWE1，进一步证实BARD1能与HUWE1相互作用，而且能够调节HUWE1的蛋白和泛素化水平，从而抑制HUWE1对BRCA1的降解。本研究我们证实，与HUWE1结合的BARD1被NEDD8所修饰，此修饰的BARD1可以调控HUWE1的蛋白稳定性及泛素化水平。BARD1被NEDD8修饰后，通过调节HUWE1而稳定BRCA1，但BARD1突变体Q564H、C557S缺乏neddylation修饰，缺乏对HUWE1的抑制作用。