Studies on ROC1-dependent E3 ubiquitin ligases

描述（由申请人提供）：在细胞中，大多数不需要的和有害的蛋白质通过称为泛素-蛋白酶体途径的过程被消除。在这个过程中，靶蛋白被标记有泛素分子链，然后所产生的标记蛋白被蛋白酶体识别，从而驱动蛋白质降解。泛素标记和蛋白酶体活性的正常运作对于细胞和生物体的功能至关重要。泛素-蛋白酶体系统的调节缺陷在包括癌症在内的人类疾病中是明显的。我们的长期目标是了解SCF E3泛素连接酶复合物的精确功能，SCF E3泛素连接酶复合物是细胞中负责用泛素链标记大量蛋白质进行降解的关键机制。我们项目的短期目标是确定泛素链是如何建立在蛋白质上的，这是降解的信号。蛋白质目标有一个内置的“降解代码”，决定它的破坏。在泛素分子中，存在“缀合密码”以指导形成泛素链或将泛素连接至蛋白质靶标的键。泛素标记反应可以被视为一个过程，开始于蛋白质靶上的“降解密码”和泛素“缀合密码”的“阅读”，通过蛋白质促进剂，包括SCF E3和酶，如UbcH 5和Cdc 34。随后，SCF，UbcH 5和Cdc 34作用于“构建”一个大的蛋白质网络，将蛋白质靶标与泛素链连接起来。然而，关于泛素“共轭密码”的性质，这些密码如何被“读取”，以及泛素标记所必需的网络“构建”过程，仍然知之甚少。本研究利用生物化学、分子生物学和细胞生物学的工具，主要目的是：1）鉴定能够形成蛋白质靶点和SCF E3复合物的新分子组分，SCF E3复合物能够触发泛素标记过程; 2）了解SCF、UbcH 5和Cdc 34如何构建蛋白质网络，从而用泛素标记靶蛋白; 3）确定泛素的“共轭密码”，并揭开Cdc 34如何阅读这些“密码”的奥秘。" 公共卫生相关性：该项目旨在揭开用泛蛋白分子链标记蛋白质靶点从而导致蛋白质降解的奥秘。这项研究中获得的知识将促进对这种泛素标记过程的理解，这对细胞和生物体的功能至关重要。重要的是，该项目揭示的分子见解将改善有效治疗包括癌症在内的人类疾病的药物设计。

DESCRIPTION (provided by applicant): In cells, a majority of unwanted and harmful proteins are eliminated through a process called the ubiquitin-proteasome pathway. In this process, a target protein is tagged with a chain of ubiquitin molecules and the resulting marked protein is then recognized by the proteasome, which drives protein degradation. Proper functioning of the ubiquitin tagging and proteasomal activities is of key importance for the function of a cell and an organism. Defective regulation of the ubiquitin-proteasome system is manifest in human diseases including cancers. Our long-range goal is to understand the precise functioning of the SCF E3 ubiquitin ligase complex, which is the key machinery in cells responsible for tagging a large set of proteins with a chain of ubiquitin for degradation. The short-term objective of our project is to determine how a chain of ubiquitin is built on a protein, which signals degradation. A protein target has a built-in "degradation code" that dictates its destruction. In an ubiquitin molecule, "conjugation codes" are present to direct the formation of an ubiquitin chain, or a linkage that connects an ubiquitin to the protein target. The ubiquitin tagging reaction can be viewed as a process, beginning with the "reading" of the "degradation code" on the protein target, and ubiquitin "conjugation codes," by protein facilitators that include SCF E3 and enzymes such as UbcH5 and Cdc34. Subsequently, SCF, UbcH5, and Cdc34 act to "build" a large protein network that links the protein target with a chain of ubiquitin. However, it remains poorly understood regarding the nature of the ubiquitin "conjugation codes," how these codes are "read," as well as the network "building" process necessary for ubiquitin tagging. Using the tools of biochemistry, molecular biology and cell biology, our project is to: 1) identify new molecular components that help form the complex of a protein target and an SCF E3, which triggers the ubiquitin tagging process; 2) understand how SCF, UbcH5, and Cdc34 build protein networks for tagging a target protein with ubiquitin; 3) identify the ubiquitin "conjugation codes" and uncover the mystery of how Cdc34 reads these "codes." PUBLIC HEALTH RELEVANCE: This project intends to uncover the mystery of tagging a protein target with a chain of ubiquitin molecules, which leads to protein degradation. Knowledge gained in this study will advance understanding of this ubiquitin tagging process that is fundamental to the function of a cell and an organism. Importantly, the molecular insights revealed in this project will improve the designing of drugs effective for treating human diseases that include cancer.