The Tribbles-COP1 Complex in Leukemia

描述（由申请方提供）：拟定研究的总体目标是表征AML形成中Tribbles蛋白（TRIB）和E3连接酶COP 1之间的分子相互作用，利用结构研究的见解作为未来发现该复合物靶向抑制剂的基础。我们还打算确定TRIB-COP 1介导的泛素化和降解与人类疾病相关的其他目标。TRIB过表达在100%转化率的小鼠中驱动白血病形成。人类AML样本的表达阵列研究支持TRIB在人类白血病中的作用。在小鼠模型中，TRIB抑制骨髓分化的能力取决于其结合COP 1的能力，COP 1是一种复合物，可导致靶向骨髓分化因子C/EBP？降解。在这里，我们提出了一个模型，其中TRIB通过直接结合COP 1的一个结构域来指导COP 1对C/EBP的特异性。如果正确的话，该模型将邀请针对TRIB-COP 1界面的抑制剂的设计。本文所建议的工作将在Stephen Blacklow博士的指导下进行。候选人是一名医生，科学家，具有临床病理学委员会认证和输血医学亚专业培训。这里概述的研究将构成候选人独立研究的基础。目标1.确定功能性TRIB-COP 1复合物组装的结构基础。子1.1确定Tribbles和COP 1之间的结合模式。我们的第一个结构目标将是COP 1的TRIB结合区与TRIB 1的COP 1结合基序之间的复合物，这将为TRIB结构-功能关系提供新的见解，并为设计直接破坏TRIB-COP 1相互作用的选择性抑制剂提供必要的基础。子1.2确定Tribbles和COP 1识别降解底物的机制。我们将联合收割机诱变与生物物理和结构方法相结合，以确定底物识别的基础，长期目标是解决含有TRIB蛋白，COP 1和底物的三元复合物的结构。目标2.发现TRIB-COP 1复合物在哺乳动物细胞中的新的、功能重要的底物。我们现在正在使用底物捕获策略发现TRIB-COP 1介导的降解的新靶标，其中COP 1连接酶活性已被灭活。这一目标的完成将加深我们对TRIB在哺乳动物中功能的理解，并为TRIB-COP 1复合物在哺乳动物细胞中降解的底物范围提供更多的见解。总之，这些目标的完成将揭示TRIB-COP 1复合物促进AML的分子机制，并作为未来设计这些复合物的靶向抑制剂的模板。

DESCRIPTION (provided by applicant): The overarching goal of the proposed studies is to characterize the molecular interactions between Tribbles proteins (TRIBs) and the E3 ligase COP1 in the formation of AML, harnessing insights from the structural studies as the foundation for future discovery of targeted inhibitors of this complex. We also intend to identify additional targets of TRIB-COP1-mediated ubiquitination and degradation relevant to human disease. TRIB overexpression drives leukemia formation in mice with 100% penetrance. Expression array studies of human AML samples support a role for TRIB in human leukemia. In the mouse model, TRIB's ability to inhibit myeloid differentiation hinges on its ability to bind COP1, formin a complex that causes targets the myeloid differentiation factor C/EBP� for degradation. Here, we propose a model in which TRIB directs the specificity of COP1 for C/EBP� by binding directly to one domain of COP1. If correct, this model would invite the design of inhibitors that target the TRIB-COP1 interface. The work proposed here will be conducted under the mentorship of Dr. Stephen Blacklow. The candidate is a physician-scientist with board certification in Clinical Pathology and subspecialty training in Transfusion Medicine. The studies outlined here will form the foundation for the candidate's independent research. Aim 1. To determine the structural basis for assembly of functional TRIB-COP1 complexes. Subaim1.1 To determine the mode of binding between Tribbles and COP1. Our first structural target will be a complex between the TRIB-binding region of COP1 bound to the COP1 binding motif of TRIB1, which will give new insights into TRIB structure-function relationships and provide the foundation necessary to design selective inhibitors that directly disrupt the TRIB-COP1 interaction. Subaim1.2 To identify the mechanism by which Tribbles and COP1 recognize substrates destined for degradation. We will combine mutagenesis with biophysical and structural approaches to determine the basis for substrate recognition, with the long-term goal of solving the structure of a ternary complex containing a TRIB protein, COP1, and substrate. Aim 2. To discover new, functionally important substrates of TRIB-COP1 complexes in mammalian cells. We are now discovering new targets of TRIB-COP1 mediated degradation using a substrate-trapping strategy in which the COP1 ligase activity has been inactivated. Completion of this aim will deepen our understanding of TRIB function in mammals, and provide additional insight into the range of substrates targeted for degradation by TRIB-COP1 complexes in mammalian cells. Together, completion of these aims will reveal the molecular mechanism by which TRIB-COP1 complexes promote AML, and serve as the template for future design of targeted inhibitors of these complexes.