Structural basis for CBP/p300 transcriptional regulation

描述（由申请人提供）：CREB结合蛋白（CBP）和p300是模块化转录共激活因子，在真核细胞中整合了许多信号转导途径，对生长、分化、凋亡和DNA修复至关重要。CBP/p300在胚胎发育中起关键作用，是许多致癌事件的靶点。它们也作为肿瘤抑制因子发挥作用，并且是许多细胞和肿瘤病毒癌蛋白的转录活性所必需的。CBP和p300在介导肿瘤抑制因子p53的稳定性和转录活性中起重要作用。这些蛋白质之间的相互作用是高度动态的，并在多个位点的p53反式激活结构域的磷酸化，以响应遗传毒性应激调制。腺病毒癌蛋白E1 A与细胞因子竞争螯合CBP/p300，破坏正常细胞周期控制以诱导细胞永生化。本研究将利用广泛的蛋白质构建体库来研究p53和E1 A与CBP的TAZ 1、TAZ 2、KIX和核辅激活因子结合结构域的相互作用。将确定由p53的二分N-末端反式激活结构域与CBP结构域形成的复合物的结构，以获得对结构域之间的相互作用的动态相互作用的新见解，并进入CBP/p300激活p53调节的转录的分子基础。将研究在单个和多个位点磷酸化p53反式激活结构域对与这些结构域相互作用的影响。将确定E1 A（非致癌AdV 5和强致癌AdV 12血清型）与CBP/p300结构域形成的复合物的结构，以深入了解E1 A与其他转录因子竞争螯合CBP/p300并破坏细胞转录机制的分子基础。最后，两个三元复合物的特点，HDM 2，p53，和TAZ 1结构域的CBP之间，和之间的口袋结构域的视网膜母细胞瘤蛋白Rb，E1 A和TAZ 2结构域。由于其强调弱的竞争性相互作用以及多个结合位点和多个合作伙伴之间的协同相互作用，这项研究将为CBP/p300的分子识别机制提供新的和基本的见解，以及它们介导与p53和病毒癌蛋白在肿瘤抑制，生长和进展中的关键相互作用的分子基础。

DESCRIPTION (provided by applicant): CREB binding protein (CBP) and p300 are modular transcriptional coactivators that integrate numerous signal transduction pathways in eukaryotic cells, and are essential for growth, differentiation, apoptosis, and DNA repair. CBP/p300 play critical roles in embryonic development and are the targets of many oncogenic events. They also function as tumor suppressors and are required for the transcriptional activities of many cellular and tumor virus oncoproteins. CBP and p300 play an essential role in mediating the stability and transcriptional activity of the tumor suppressor p53. The interactions between these proteins are highly dynamic and are modulated by phosphorylation of the p53 transactivation domain at multiple sites in response to genotoxic stress. The adenoviral oncoprotein E1A competes with cellular factors to sequester CBP/p300, disrupting normal cell cycle control to induce cell immortalization. This research will utilize an extensive library of protein constructs to investigate the interactions of p53 and E1A with the TAZ1, TAZ2, KIX, and nuclear coactivator binding domains of CBP. The structures of the complexes formed by the bipartite N-terminal transactivation domain of p53 with the CBP domains will be determined, to obtain new insights into the dynamic interplay of interactions between the domains and into the molecular basis for CBP/p300 activation of p53-regulated transcription. The effects of phosphorylation of the p53 transactivation domain, at single and multiple sites, on interactions with these domains will be investigated. Structures will be determined for the complexes formed by E1A (both non-oncogenic AdV5 and strongly oncogenic AdV12 serotypes), with domains of CBP/p300, in order to obtain insights into the molecular basis by which E1A competes with other transcription factors to sequester CBP/p300 and subvert the cellular transcription machinery. Finally, two ternary complexes will be characterized, between HDM2, p53, and the TAZ1 domain of CBP, and between the pocket domain of the retinoblastoma protein Rb, E1A and the TAZ2 domain. With its emphasis on weak, competing interactions and the synergistic interplay between multiple binding sites and multiple partners, this research will provide new and fundamental insights into the mechanisms of molecular recognition by CBP/p300, and the molecular basis by which they mediate critical interactions with p53 and viral oncoproteins in tumor suppression, growth and progression.