Specific domains of nucleolin interact with Hdm2 and antagonize Hdm2-mediated p53 ubiquitination.
Specific domains of nucleolin interact with Hdm2 and antagonize Hdm2-mediated p53 ubiquitination.
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DOI：
10.1111/j.1742-4658.2011.08430.x
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发表时间：
2012-02
期刊：
影响因子：
--
通讯作者：
Saxena A
中科院分区：
文献类型：
--
作者：
Bhatt P;d'Avout C;Kane NS;Borowiec JA;Saxena A
核仁素是一种丰富的多功能核仁蛋白，在核糖体RNA加工、RNA聚合酶I催化的转录和细胞凋亡调节中具有明确的作用。早些时候，我们报告说，人类核仁素结合p53-拮抗剂HDM 2确定互惠共免疫沉淀试验使用细胞裂解物。我们还表明，核仁素拮抗HDM 2介导的p53降解。在这里，我们确定了特定的结构域的核仁素和HDM 2蛋白，支持相互作用，并研究了p53-泛素化和蛋白质水平上的复合物形成的影响。我们的数据表明，核仁素N-末端以及中央RNA结合结构域（RBD）主要参与Hdm 2的结合。核仁素RBD在体外与Hdm 2的NLS/内斯（nuclear localization and export signals）结构域结合，而核仁素N端优先与细胞中表达的Hdm 2 RING结构域结合。我们进一步证明核仁素的C-末端GAR（富含甘氨酸-精氨酸）结构域是与p53直接相互作用的主要结合结构域。虽然核仁素或其各种结构域的过表达对Hdm 2自身泛素化没有显著影响，但核仁素RBD拮抗Hdm 2 E3连接酶对p53的活性，导致p53稳定。相反，核仁素的相邻GAR结构域与p53相互作用，对p53泛素化产生适度的刺激作用。这些数据表明，核仁素构象的变化可以改变这些结构域在体内的可用性，以调节核仁素对Hdm 2活性的整体影响，从而对p53稳定性。
Nucleolin is an abundant multifunctional nucleolar protein with defined roles in ribosomal RNA processing, RNA polymerase I-catalyzed transcription, and the regulation of apoptosis. Earlier we reported that human nucleolin binds to the p53-antagonist Hdm2 as determined by reciprocal co-immunoprecipitation assays using cell lysates. We also demonstrated that nucleolin antagonizes Hdm2-mediated degradation of p53. Here, we identify specific domains of nucleolin and Hdm2 proteins that support mutual interaction and investigated the implications of complex formation on p53-ubiquitination and protein levels. Our data indicate that the nucleolin N-terminus as well as the central RNA-binding domain (RBD) are predominantly involved in binding to Hdm2. The nucleolin RBD robustly bound to the NLS/NES (nuclear localization and export signals) domain of Hdm2 in vitro, while N-terminus of nucleolin preferentially associated with the Hdm2 RING domain expressed in cells. We further demonstrate that the C-terminal GAR (Glycine-Arginine Rich) domain of nucleolin serves as the predominant binding domain for direct interaction with p53. While over-expression of nucleolin or its various domains had no significant effect on Hdm2 auto-ubiquitination, the nucleolin RBD antagonized the Hdm2 E3 ligase activity against p53, leading to p53 stabilization. Conversely, the adjacent GAR domain of nucleolin interacted with p53 causing a modest stimulatory effect on p53 ubiquitination. These data suggest that changes in nucleolin conformation can alter the availabilities of such domains in vivo to modulate the overall impact of nucleolin on Hdm2 activity and hence on p53 stability.
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影响因子：
5.3
作者：
Daniely, Y;Dimitrova, DD;Borowiec, JA
通讯作者：
Borowiec, JA
影响因子：
21.3
作者：
Colombo, E;Marine, JC;Pelicci, PG
通讯作者：
Pelicci, PG
影响因子：
10.5
作者：
Jayaraman, L;Moorthy, NC;Prives, C
通讯作者：
Prives, C
影响因子：
3.7
作者：
FANG, SH;YEH, NH
通讯作者：
YEH, NH
影响因子：
14.9
作者：
Greasley, PJ;Bonnard, C;Amati, B
通讯作者：
Amati, B