Deubiquitination regulation of c-Myc

c-Myc 的去泛素化调控

基本信息

批准号：
9245658
负责人：
Mu-Shui Dai
金额：
$ 37.65万
依托单位：
OREGON HEALTH & SCIENCE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2020-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9245658
关键词：
Affect Binding Biochemical Biogenesis Biology Burkitt Lymphoma Cell Culture Techniques Cell Nucleolus Cell Proliferation Cells Complex Cultured Cells DNA Polymerase II DNA Polymerase III Deubiquitinating Enzyme Deubiquitination Event Family member Gene Targeting Genes Genetic Transcription Goals Growth Homeostasis Hot Spot Human In Vitro Investigation Lead Link MCF10A cells MYC Box Malignant Neoplasms Mammary Tumorigenesis Mediating Messenger RNA Molecular Mus Mutation Normal Cell Nucleoplasm Oncogenic Oncoproteins Phosphorylation Play Post-Translational Protein Processing Proteins Proteolysis Proto-Oncogene Proteins c-myc Recombinant DNA Recruitment Activity Regulation Reporting Research Ribosomes Role Serine Serum Signal Transduction Stress System Testing Threonine Translations Ubiquitin Ubiquitination base c-myc Genes cancer therapy cell growth feeding high throughput screening in vivo insight killings knock-down malignant breast neoplasm mammary epithelium mouse model multicatalytic endopeptidase complex mutant novel novel therapeutics overexpression promoter protein degradation public health relevance response small molecule inhibitor therapeutic development therapeutic target tumor xenograft tumorigenesis ubiquitin ligase ubiquitin-protein ligase ubiquitin-specific protease

项目摘要

DESCRIPTION (provided by applicant): The c-Myc oncoprotein is essential for normal cell growth and proliferation. However, overexpression of c-Myc occurs in most human cancers. Thus, its level and activity must be tightly regulated during normal cell homeostasis. The ubiquitination-proteasome system plays a key role in controlling c-Myc levels and activity. c-Myc normally undergoes rapid ubiquitin-dependent proteolysis, but it is transiently stabilized by key phosphorylation events in response to growth signals. Phosphorylation of Serine 62 (S62) stabilizes c-Myc, whereas phosphorylation of Threonine 58 (T58) promotes c-Myc ubiquitination by the SCFFbw7 ubiquitin ligase and proteasomal degradation, mainly in the nucleolus. Like other post-translational modifications, ubiquitination can be reversed by the action of deubiquitinating enzymes (DUBs). While several ubiquitin ligases have been identified for c-Myc, only one DUB, USP28, has been reported to target c-Myc. We have recently discovered that the nucleolar deubiquitinating enzyme USP36 is a novel c-Myc regulator. USP36 binds to c-Myc and deubiquitinates c-Myc in cells and in vitro. Overexpression of wild-type USP36, but not its catalytic-inactive C131A mutant, stabilizes c-Myc and enhances c-Myc-driven transcription. Knockdown of USP36 reduces c-Myc levels and drastically suppresses cell proliferation. Importantly, USP36 interacts with the nucleolar Fbw7γ and abolishes Fbw7γ-mediated c-Myc degradation. In contrast, USP28 antagonizes Fbw7-mediated c-Myc degradation. Since the bulk of c-Myc is degraded in the nucleolus, our discovery leads to the novel hypothesis that USP36 functions as a crucial regulator of c-Myc by deubiquitinating c-Myc in the nucleolus. Interestingly, we found that USP36 itself is a c-Myc target gene, suggesting that USP36 and c-Myc form a positive feed-forward regulatory loop. To gain further insight into the role of USP36 in the regulation of c-Myc protein stability, activity and oncogenicity, we will investigate the molecular and biochemical mechanisms underlying the regulation of c-Myc by USP36 in Aim 1, including how USP36 interplays with Fbw7γ to regulate c-Myc in the nucleolus, whether it interplays with USP28 in the dynamic control of c-Myc ubiquitination, and the importance of c-Myc-USP36 feed-forward regulation. We will elucidate the functional consequences of USP36 regulation of c-Myc in cells in Aim 2 by analyzing whether USP36 regulates c-Myc binding and turnover at target gene promoters, whether it promotes c-Myc-dependent ribosome biogenesis, and whether it promotes c-Myc's oncogenic potential in cells and in vivo. Finally, we will elucidate whether USP36 is a therapeutic target using cell based and mouse models as proposed in Aim 3, including the investigation of USP36 deregulation in human breast cancers, whether deletion of USP36 inhibits c-Myc-driven mammary tumorigenesis in mice, and high-throughput screening of small molecule inhibitors for USP36. Achieving these goals will provide critical insight into how c-Myc is properly regulated by dynamic ubiquitination and deubiquitination, how deregulation of this dynamic contributes to tumorigenesis, and how USP36 can be targeted in human cancers.

描述（由适用提供）：C-Myc癌蛋白对于正常细胞生长和增殖至关重要。但是，在大多数人类癌症中，C-MYC的过表达发生。这是在正常细胞稳态期间必须严格调节其水平和活性。泛素化 - 蛋白酶体系统在控制C-MYC水平和活动中起关键作用。 C-Myc通常经历泛素依赖性蛋白水解的快速，但由于响应生长信号而被关键的磷酸化事件暂时稳定。丝氨酸62（S62）的磷酸化稳定C-MYC，而苏氨酸58（T58）的磷酸化促进了SCFFFBW7泛素连接酶和蛋白酶体降解的C-Myc泛素化，主要是在核中。与其他翻译后修饰一样，泛素化酶（DUB）的作用可以逆转。虽然已经确定了用于C-MYC的几种泛素连接酶，但据报道只有一个DUB USP28靶向C-Myc。我们最近发现，核去泛素化酶USP36是一种新型的C-MYC调节剂。 USP36与C-MYC结合，并在细胞和体外去泛素化C-MYC。野生型USP36的过表达，而不是其催化性C131a突变体稳定C-MYC并增强C-MYC驱动的转录。 USP36的敲低可降低C-MYC水平并大大抑制细胞增殖。重要的是，USP36与核仁FBW7γ相互作用，并废除了FBW7γ介导的C-MYC降解。相比之下，USP28拮抗FBW7介导的C-MYC降解。由于大部分C-MYC在核仁中降解，因此我们的发现导致了新的假设，即USP36通过在核Olus中的c-Myc通过去泛素化C-MYC作为C-MYC的关键调节剂。有趣的是，我们发现USP36本身是一个C-MYC靶基因，这表明USP36和C-MYC形成了正向前馈调节环。为了进一步了解USP36在调节C-MYC蛋白稳定性，活性和致癌性中的作用，我们将研究AIM 1中USP36调节C-Myc的基础的分子和生化机制，包括USP36与FBW7γ中的C-MYC在核OLUS中的调节方式，包括USP36与核OLUS的互动如何相互控制。泛素化以及C-MYC-USP36馈送法规的重要性。我们将通过分析USP36是否调节靶基因启动子的C-MYC结合和周转率，是否促进C-MYC依赖性核糖体生物发生以及它是否促进C-MYC在细胞中的Oncenitigent中是否促进C-MYC依赖性的核糖体生物发生，通过分析USP36调节C-MYC的结合和在目标基因启动子处的C-MYC结合和周转，通过分析USP36调节C-MYC的结合和转离，我们将阐明USP36调节C-MYC在AIM 2中的功能后果。最后，我们将阐明USP36是否是AIM 3中提出的基于细胞和小鼠模型的治疗靶标，包括研究人乳腺癌中对USP36进行管制的调查，USP36的删除是否抑制C-MYC驱动的乳腺肿瘤在小鼠中的乳腺肿瘤，以及对USP36的小型分子筛选的高直发筛选。实现这些目标将为C-MYC如何通过动态的泛素化和去泛素化的正确调节，这种动态导致肿瘤发生促进以及如何将USP36靶向人类癌症。