Ferreon Josephine C, Lee Chul Won, Arai Munehito, Martinez-Yamout Maria A, Dyson H Jane, Wright Peter E
Department of Molecular Biology, The Scripps Research Institute, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA.
Proc Natl Acad Sci U S A. 2009 Apr 21;106(16):6591-6. doi: 10.1073/pnas.0811023106. Epub 2009 Apr 8.
The tumor suppressor activity of p53 is regulated by interactions with the ubiquitin ligase HDM2 and the general transcriptional coactivators CBP and p300. Using NMR spectroscopy and isothermal titration calorimetry, we have dissected the binding interactions between the N-terminal transactivation domain (TAD) of p53, the TAZ1, TAZ2, KIX, and nuclear receptor coactivator binding domains of CBP, and the p53-binding domain of HDM2. The p53 TAD contains amphipathic binding motifs within the AD1 and AD2 regions that mediate interactions with CBP and HDM2. Binding of the p53 TAD to CBP domains is dominated by interactions with AD2, although the affinity is enhanced by additional interactions with AD1. In contrast, binding of p53 TAD to HDM2 is mediated primarily by AD1. The p53 TAD can bind simultaneously to HDM2 (through AD1) and to any one of the CBP domains (through AD2) to form a ternary complex. Phosphorylation of p53 at T18 impairs binding to HDM2 and enhances affinity for the CBP KIX domain. Multisite phosphorylation of the p53 TAD at S15, T18, and S20 leads to increased affinity for the TAZ1 and KIX domains of CBP. These observations suggest a mechanism whereby HDM2 and CBP/p300 function synergistically to regulate the p53 response. In unstressed cells, CBP/p300, HDM2 and p53 form a ternary complex that promotes polyubiquitination and degradation of p53. After cellular stress and DNA damage, p53 becomes phosphorylated at T18 and other residues in the AD1 region, releases HDM2 and binds preferentially to CBP/p300, leading to stabilization and activation of p53.
p53的肿瘤抑制活性通过与泛素连接酶HDM2以及一般转录共激活因子CBP和p300的相互作用来调节。利用核磁共振光谱和等温滴定量热法，我们剖析了p53的N端反式激活结构域（TAD）、CBP的TAZ1、TAZ2、KIX和核受体共激活因子结合结构域以及HDM2的p53结合结构域之间的结合相互作用。p53 TAD在AD1和AD2区域包含两亲性结合基序，介导与CBP和HDM2的相互作用。p53 TAD与CBP结构域的结合主要由与AD2的相互作用主导，尽管与AD1的额外相互作用会增强亲和力。相比之下，p53 TAD与HDM2的结合主要由AD1介导。p53 TAD可以同时与HDM2（通过AD1）和任何一个CBP结构域（通过AD2）结合形成三元复合物。p53在T18位点的磷酸化会损害其与HDM2的结合，并增强对CBP KIX结构域的亲和力。p53 TAD在S15、T18和S20位点的多位点磷酸化导致对CBP的TAZ1和KIX结构域的亲和力增加。这些观察结果提示了一种机制，即HDM2和CBP/p300协同发挥作用来调节p53反应。在未受应激的细胞中，CBP/p300、HDM2和p53形成三元复合物，促进p53的多聚泛素化和降解。细胞应激和DNA损伤后，p53在T18和AD1区域的其他残基处发生磷酸化，释放HDM2并优先与CBP/p300结合，导致p53的稳定和激活。
