Chemical Genetic and Biochemical Studies of Mitotic Proteolysis

有丝分裂蛋白水解的化学遗传学和生化研究

• 批准号: 7922343
• 负责人: RANDALL W KING
• 金额: $ 19.97万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7922343
• 关键词: Affect; Antimitotic Agents; Antineoplastic Agents; Binding; Biochemical; Biochemical Reaction; Biological; CDH1 gene; Cell Cycle; Cell Extracts; Cell division; Cell physiology; Cells; Chemicals; Chromatin; Complex; Conserved Sequence; Data; Development; Disease; Elements; Goals; Indium; Injury; Link; Lysine; Mediating; Mitosis; Mitotic; Nature; Paclitaxel; Pathway interactions; Permeability; Process; Protein Binding; Protein Kinase; Proteins; Proteolysis; Recruitment Activity; Regulation; Research; Research Personnel; Series; Signal Transduction; Structure; System; Tail; Testing; Tosylarginine Methyl Ester; Ubiquitin; Ubiquitin-Activating Enzymes; Ubiquitination; Xenopus; anaphase-promoting complex; axon growth; base; cancer therapy; chemical genetics; chemical synthesis; cyclin B1; drug candidate; high throughput screening; improved; inhibitor/antagonist; insight; multicatalytic endopeptidase complex; mutant; new therapeutic target; novel; preference; prevent; programs; protein degradation; receptor; reconstitution; research study; small molecule; tool; ubiquitin ligase

DESCRIPTION (provided by applicant): The ubiquitin-proteasome system regulates many essential cellular processes including cell division. The recent development of cancer drugs that inhibit the proteasome suggests that the ubiquitin-proteasome system may provide many new therapeutic targets. The goal of this project is to develop new chemical inhibitors of the ubiquitin-proteasome system, and to understand how this pathway regulates cell division. We first plan to characterize novel small molecule inhibitors of the Anaphase-Promoting Complex (APC), a ubiquitin ligase that is essential for cell division. APC inhibitors may be useful for the treatment of cancer, because they may inhibit cell division or sensitize cells to antimitotic drugs such as taxol. Additionally, APC inhibitors may be useful for the treatment of diseases that involve axonal injury, because the APC negatively regulates axon growth. Our studies of these small molecules will also provide important new insights into how the APC is regulated by activator proteins. Another goal of our studies is to characterize ubistatins, compounds that prevent ubiquitin- dependent proteolysis by binding to ubiquitin chains. However, the mechanism of ubiquitin chain binding remains unclear, and we plan to synthesize new ubistatin derivatives that will allow us to determine the structure of ubistatins bound to ubiquitin chains. The structural information will enable us to synthesize more potent and perhaps more cell-permeable versions of ubistatins, enhancing the utility of these compounds to investigators in the field, and accelerating the development of ubiquitin-binding compounds as drug candidates. A third goal of our research program is to understand how ubiquitinated substrates are recognized for degradation by the proteasome. It has been assumed that a specific type of ubiquitin chain (linked through lysine-48 of ubiquitin) is required for degradation by the proteasome. Our studies of ubiquitinated cyclin B1 suggest that other types of ubiquitin linkages may be sufficient to target the protein for degradation. We therefore plan to determine the types of ubiquitination that are required for degradation of cyclin B1 by purified proteasomes and in complex cell extracts. These studies are critical for understanding how small molecule inhibitors of ubiquitination and ubiquitin chain recognition may perturb degradation. Finally, we want to understand how the localization of substrates affects their degradation by the ubiquitin-proteasome system. We have identified a novel sequence element in cyclin B1 that recruits the protein to chromatin during mitosis. We plan to identify the protein that recruits cyclin B1 to chromatin, and to determine the functional consequences of cyclin B1 association with chromatin. Together these studies will provide new chemical tools for studying the ubiquitin-proteasome pathway, and will help us understand how a critical substrate of this pathway, cyclin B1, is regulated during the cell cycle.

描述（由申请人提供）：泛素-蛋白酶体系统调节许多重要的细胞过程，包括细胞分裂。最近开发的抑制蛋白酶体的癌症药物表明泛素-蛋白酶体系统可能提供许多新的治疗靶点。该项目的目标是开发泛素蛋白酶体系统的新化学抑制剂，并了解该途径如何调节细胞分裂。我们首先计划表征后期促进复合物（APC）的新型小分子抑制剂，APC是一种对细胞分裂至关重要的泛素连接酶。 APC 抑制剂可能有助于治疗癌症，因为它们可以抑制细胞分裂或使细胞对紫杉醇等抗有丝分裂药物敏感。此外，APC 抑制剂可用于治疗涉及轴突损伤的疾病，因为 APC 负向调节轴突生长。我们对这些小分子的研究也将为 APC 如何受激活蛋白的调节提供重要的新见解。我们研究的另一个目标是表征泛素，即通过与泛素链结合来防止泛素依赖性蛋白水解的化合物。然而，泛素链结合的机制仍不清楚，我们计划合成新的泛素衍生物，这将使我们能够确定泛素链结合的泛素结构。这些结构信息将使我们能够合成更有效且可能更具细胞渗透性的泛素，增强这些化合物对该领域研究人员的实用性，并加速泛素结合化合物作为候选药物的开发。我们研究计划的第三个目标是了解泛素化底物如何被蛋白酶体识别并降解。据推测，蛋白酶体降解需要特定类型的泛素链（通过泛素的赖氨酸 48 连接）。我们对泛素化细胞周期蛋白 B1 的研究表明，其他类型的泛素连接可能足以靶向该蛋白质进行降解。因此，我们计划确定纯化蛋白酶体和复杂细胞提取物降解细胞周期蛋白 B1 所需的泛素化类型。这些研究对于了解泛素化和泛素链识别的小分子抑制剂如何干扰降解至关重要。最后，我们想了解底物的定位如何影响泛素-蛋白酶体系统对其的降解。我们在细胞周期蛋白 B1 中发现了一个新的序列元件，它在有丝分裂过程中将该蛋白募集到染色质上。我们计划鉴定将细胞周期蛋白 B1 招募到染色质的蛋白质，并确定细胞周期蛋白 B1 与染色质关联的功能后果。这些研究将为研究泛素-蛋白酶体途径提供新的化学工具，并将帮助我们了解该途径的关键底物细胞周期蛋白 B1 在细胞周期中是如何受到调节的。