Mechanisms of Chaperone-Mediated Control in the Assembly of the Proteasome Holoenzyme

蛋白酶体全酶组装中分子伴侣介导的控制机制

• 批准号: 10224660
• 负责人: Soyeon Park
• 金额: $ 31.77万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10224660
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Affect; Aging; Binding; Binding Sites; Biochemical; Biological; Biological Assay; Cell physiology; Cells; Complex; Critical Pathways; Data; Dependence; Development; Disease; Escherichia coli; Event; Goals; Growth; Health; Holoenzymes; Human; In Vitro; Link; Malignant Neoplasms; Mediating; Mediator of activation protein; Medical; Modeling; Molecular Chaperones; Molecular Machines; Nerve Degeneration; Neurodegenerative Disorders; Nucleosome Core Particle; Nucleotides; Outcome; Pathologic; Pathway interactions; Phenotype; Polyubiquitination; Process; Proteins; Proteolysis; Public Health; Publishing; Quality Control; Regulation; Research; Role; Route; Saccharomycetales; Structure; Syndrome; System; Testing; Time; Ubiquitin; Ubiquitination; Variant; Work; Yeast Model System; Yeasts; base; cell growth; combat; human disease; in vivo; innovation; multicatalytic endopeptidase complex; mutant; new therapeutic target; prevent; protein degradation; reconstitution; therapeutic target; ubiquitin-protein ligase; yeast genetics

PROJECT SUMMARY The proteasome is a complex molecular machine and an essential mediator of ubiquitin-dependent protein degradation. Polyubiquitination of target proteins has been long considered the major regulator of proper protein degradation. However, little is known about a new and potent regulator of cellular protein degradation — mechanisms that control the rate and quality of proteasome assembly. It is well established that cells require multiple evolutionarily conserved chaperones to regulate proteasome assembly. A catalyzed assembly mechanism serves as an active regulator of proper proteasome assembly, and therefore of proteolysis itself. Under some pathological situations, cells are known to exploit these chaperones to alter the rate of proteasome- mediated proteolysis. Moreover, chaperone-mediated assembly of proteasomes has also been linked to major pathways for growth control. Therefore, understanding how chaperones control proteasome assembly is of wide- ranging biological and biomedical significance. The objective of this proposal is to elucidate how these chaperones function to regulate the rate and the quality of proteasome formation. Our central hypothesis is that the chaperones determine correct versus incorrect assembly of the proteasome by regulating both the ATP hydrolysis and the ubiquitination of the ATPase subunits en route to proteasome formation. Aim 1 will determine how chaperone-mediated inhibition of ATP hydrolysis by the proteasome subunits controls proteasome assembly events. Both in vitro and in vivo proteasome assembly assays will be used while the ATPase rates are directly modulated by specific yeast mutants. Aim 2 will determine how ubiquitinations of the proteasome subunits provide quality control during chaperone-mediated proteasome assembly. Biochemical approaches similar to Aim 1 will be used in combination with yeast genetics. The goal is to identify how the chaperones regulate ubiquitination of the proteasome subunits, and how ubiquitination affects assembly intermediates and the assembled proteasome holoenzyme. The expected outcome is the identification of the mechanisms by which the chaperones determine incorrect versus correct assembly events, and provide quality control, thereby enabling only the correct assembly events to proceed in forming the proteasome holoenzyme. The proposed research is innovative because it aims to identify mechanisms that regulate the number of functional proteasomes in the cell, which have been missing from the current paradigm of proteasome-mediated proteolysis. This contribution is significant for human health to help understand how this critical pathway is exploited to alter protein degradation in pathological conditions, including cancer, neurodegenerative disease and aging. These conditions are known to be impacted by the ubiquitin-proteasome system, a proven therapeutic target for altering protein degradation in disease. We anticipate that understanding chaperone-mediated regulation of the proteasome subunits will contribute to the development of new targeted drugs.

项目总结 蛋白酶体是一个复杂的分子机器，是泛素依赖的重要介体 蛋白质降解。长期以来，靶蛋白的多泛素化一直被认为是适当的主要调节因素。 蛋白质降解。然而，人们对一种新的、有效的细胞蛋白质降解调节剂知之甚少-- 控制蛋白酶体组装速度和质量的机制。众所周知，细胞需要 多个进化上保守的伴侣调节蛋白酶体组装。一种催化组装 机制是适当的蛋白酶体组装的活性调节器，因此也是蛋白降解本身的调节器。 在某些病理情况下，已知细胞利用这些伴侣来改变蛋白酶体的速率- 介导的蛋白质分解。此外，伴侣介导的蛋白酶体组装也与主要的 控制生长的途径。因此，了解伴侣如何控制蛋白酶体组装具有广泛的意义。 具有广泛的生物学和生物医学意义。这项提议的目的是阐明这些 伴侣的作用是调节蛋白酶体形成的速度和质量。我们的中心假设是 伴侣通过调节两个ATP来决定蛋白酶体的正确和不正确的组装 蛋白酶体形成过程中ATPase亚基的水解和泛素化。目标一号将决定 伴侣蛋白酶体亚基抑制ATP水解酶如何控制蛋白酶体 装配事件。将使用体外和体内蛋白酶体组装分析，而ATPase速率为 由特定的酵母突变体直接调节。目标2将确定蛋白酶体的泛素化 在伴侣介导的蛋白酶体组装过程中，亚基提供质量控制。生化方法 与AIM 1类似，将与酵母遗传学结合使用。我们的目标是确定监护人是如何 调节蛋白酶体亚基的泛素化，以及泛素化如何影响组装中间体和 组装好的蛋白酶体全酶。预期的结果是确定通过哪些机制 监护人确定不正确和正确的装配事件，并由此提供质量控制 使正确的组装事件能够在形成蛋白酶体全酶的过程中进行。建议数 这项研究具有创新性，因为它的目的是确定调节功能细胞数量的机制 细胞中的蛋白酶体，这些蛋白酶体在目前的蛋白酶体介导的蛋白分解范例中已经缺失。 这一贡献对人类健康有助于理解这一关键途径是如何被利用来改变的 蛋白质在病理条件下的降解，包括癌症、神经退行性疾病和衰老。这些 已知条件受到泛素-蛋白酶体系统的影响，泛素-蛋白酶体系统是一种已被证实的改变治疗靶点。 疾病中的蛋白质降解。我们预计，对伴侣调节的调节的理解 蛋白酶体亚基将有助于新靶向药物的开发。