Deubiquitinases in Cell Cycle Control

细胞周期控制中的去泛素酶

• 批准号: 10725061
• 负责人: Michael James Emanuele
• 金额: $ 3.93万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10725061
• 关键词: Address; Aging; Binding; Biochemical; Biochemistry; Cardiac health; Cell Cycle; Cell Cycle Progression; Cell Cycle Regulation; Cell Proliferation; Cell division; Cell physiology; Cells; Cellular biology; Chromosomal Stability; Chromosome Segregation; Data; Data Set; Disease; Drug Targeting; Enzymatic Biochemistry; Enzymes; Family; Future; G1/S Transition; Genome; Genome Stability; Health; Human; Infection; Kinetics; Knowledge; Ligase; Maintenance; Malignant Neoplasms; Mitotic; Mitotic Checkpoint; Molecular Biology; Normal Cell; Outcome; Pathologic; Pathology; Pathway interactions; Patients; Peptide Hydrolases; Phenotype; Play; Polyubiquitin; Positioning Attribute; Proliferating; Proteins; Proteome; Proteomics; Regulation; Role; S phase; Signal Transduction; Specific qualifier value; Substrate Interaction; System; Techniques; Therapeutic; Ubiquitin; Ubiquitination; Work; cancer cell; genome integrity; in silico; inhibitor; loss of function; multicatalytic endopeptidase complex; nervous system disorder; ovarian neoplasm; protein degradation; restraint; therapeutic target; ubiquitin isopeptidase; ubiquitin-protein ligase; virtual

DEUBIQUITINASES IN CELL CYCLE CONTROL Project Summary Ubiquitin signaling contributes to virtually all aspects of cell physiology and is implicated in aging and disease. The covalent conjugation of polyubiquitin chains onto substrates triggers their degradation by the proteasome, as well as various other cellular outcomes. Ubiquitination is carried out by an enzymatic cascade of ubiquitin activators (E1), conjugators (E2) and ligases (E3). During normal cell cycles the ubiquitin system plays an essential and conserved role in remodeling the protein landscape. Ubiquitin substrates are determined by E3 ligases and much of our understanding of ubiquitin signaling has focused on the identity, substrates and mechanisms of E3s. However, ubiquitination is reversible, and ubiquitin is removed from substrates by catalytic proteases termed DUBs (deubiquitinases). Despite their critical role in sculpting the proteome, much less is known about the identity, substrates and mechanisms of DUBs in cell cycle progression, when compared to their E3 counterparts. Nevertheless, dysregulation of both E3s and DUBs alters cell cycle progression and has deleterious effects on genome integrity. Moreover, both E3s and DUBs can be perturbed in pathologies such as cancer, contributing to the biochemical and phenotypic features of disease. Thus, defining the identity, substrates and mechanisms of DUBs in cell cycle is essential to understanding how normal cell proliferation and genome stability are maintained and coordinated. We hypothesize that DUBs are essential for cell cycle progression and chromosome stability and are equally important as their E3 counterparts. We address this hypothesis in three specific aims, that combine complementary techniques, and which focus on the role of DUBs in major cell cycle transitions. In Aims 1 and 2 we investigate Cezanne/OTUD7B, an ovarian tumor family deubiquitinase, that we recently demonstrated is cell cycle regulated and which controls the M to G1 transition. In Aim 1, we will determine substrates for Cezanne using proteomics approaches, define mechanisms of DUB-substrate interactions, and the role of Cezanne in the degradation of substrates at M/G1. In Aim 2, we will expand this analysis to determine how Cezanne is itself regulated, both at the level of its abundance and activity, and then determine how these regulatory systems influence its role in cell division. Finally, in Aim 3, we determine the role of DUBs in a second major cell cycle transition, G1/S. The G1/S boundary is a major barrier to proliferation in normal and cancer cell cycles and relies heavily on ubiuqitin signaling. However, little is known about DUBs involved in G1/S. We will use computational approaches and loss-of-function screens, to identify and then investigate DUBs that control G1/S. Collectively, this proposal will fill significant knowledge gaps in the cell cycle, ubiquitin and DUB fields, related to roles and mechanisms of DUBs in proliferation and genome maintenance. My lab is uniquely positioned to address these questions, illustrated by our prior work, that includes global analysis of ubiquitination networks, detailed analysis of specific ubiquitin pathways and their role in cell cycle, and determination of the function and enzymology of cell cycle DUBs.

细胞周期控制中的去泛素酶 项目概要 泛素信号传导几乎对细胞生理学的所有方面都有贡献，并且与衰老和疾病有关。 多聚泛素链与底物的共价缀合触发了蛋白酶体的降解， 以及各种其他细胞结果。泛素化是通过泛素的酶级联进行的 激活剂 (E1)、接合剂 (E2) 和连接酶 (E3)。在正常细胞周期中，泛素系统发挥着 在重塑蛋白质景观中发挥重要且保守的作用。泛素底物由 E3 测定 连接酶和我们对泛素信号传导的大部分理解都集中在其身份、底物和 E3 的机制。然而，泛素化是可逆的，泛素通过催化作用从底物上去除。 称为 DUB（去泛素酶）的蛋白酶。尽管它们在塑造蛋白质组方面发挥着关键作用，但更重要的是 与它们相比，了解 DUB 在细胞周期进展中的身份、底物和机制 E3对应。然而，E3 和 DUB 的失调会改变细胞周期进程，并导致 对基因组完整性的有害影响。此外，E3 和 DUB 都可能受到以下病症的干扰： 癌症，导致疾病的生化和表型特征。因此，定义身份、底物 DUBs 在细胞周期中的作用和机制对于理解正常细胞增殖和基因组如何进行至关重要 保持稳定、协调一致。我们假设 DUB 对细胞周期进展至关重要，并且 染色体稳定性与 E3 染色体同等重要。我们分三步来阐述这个假设 具体目标，结合互补技术，重点关注 DUB 在主要细胞周期中的作用 过渡。在目标 1 和 2 中，我们研究了 Cezanne/OTUD7B，这是一种卵巢肿瘤家族去泛素化酶，我们 最近证明它可以调节细胞周期并控制 M 到 G1 的转变。在目标 1 中，我们将 使用蛋白质组学方法确定 Cezanne 的底物，定义 DUB 底物的机制 相互作用，以及塞尚在 M/G1 底物降解中的作用。在目标 2 中，我们将扩展此 分析以确定塞尚本身如何在其丰度和活性水平上受到调节，然后 确定这些调节系统如何影响其在细胞分裂中的作用。最后，在目标 3 中，我们确定 DUB 在第二个主要细胞周期转变（G1/S）中的作用。 G1/S边界是扩散的主要障碍 在正常细胞和癌细胞周期中，严重依赖泛素信号传导。然而，人们对 DUB 知之甚少 参与G1/S。我们将使用计算方法和功能丧失屏幕来识别然后 研究控制 G1/S 的 DUB。总的来说，该提案将填补细胞周期方面的重大知识空白， 泛素和 DUB 领域，与 DUB 在增殖和基因组维护中的作用和机制有关。 我的实验室具有独特的优势来解决这些问题，我们之前的工作就说明了这一点，其中包括全球 泛素化网络分析，具体泛素通路及其在细胞周期中的作用的详细分析， 以及细胞周期 DUB 的功能和酶学测定。