Interplay between Cdh1/APC and Rad17 in DNA damage checkpoints and carcinogenesis

Cdh1/APC 和 Rad17 在 DNA 损伤检查点和致癌作用中的相互作用

• 批准号: 8296553
• 负责人: Yong Wan
• 金额: $ 30.19万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-05 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8296553
• 关键词: Address; Antineoplastic Agents; Attenuated; Binding; Biological; Biological Assay; CDC2 Protein Kinase; Cancerous; Cell-Free System; Cells; Chimeric Proteins; Complex; Cutaneous Melanoma; DNA Damage; DNA Repair; DNA damage checkpoint; Data; Development; Down-Regulation; Drops; Dysplasia; Excision; Exposure to; Failure; Functional disorder; Future; Genes; Genome Stability; Genomics; Genotoxic Stress; Hepatocyte Growth Factor; Human; Knowledge; Lead; Malignant - descriptor; Malignant Neoplasms; Mediating; Mediation; Microscopy; Modeling; Molecular; Mus; Pathogenesis; Pathway interactions; Phosphorylation; Play; Process; Protein Binding; Proteins; Proteolysis; Rad17 protein; Regulation; Regulation of Proteolysis; Research; Role; Signal Transduction; Skin; Specimen; Staging; System; Testing; Therapeutic; Therapeutic Intervention; Time; UV Radiation Exposure; UV induced; UV-induced melanoma; Ubiquitin; Ubiquitin-Protein Ligase Complexes; Ultraviolet Rays; base; carcinogenesis; chromatin immunoprecipitation; design; melanocyte; melanoma; mouse model; multicatalytic endopeptidase complex; mutant; novel; protein degradation; reconstruction; response; tumorigenesis; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): Failure of DNA damage checkpoint regulation can predispose a cell to become cancerous. The ubiquitin-proteosome system (UPS) is thought to play an important role in regulating responses to DNA damage and DNA repair. In attempting to further define this role, we have observed that a critical component of the regulatory machinery, Rad17, is a UV-induced and ubiquitin-dependent fast-turnover protein. We observed that temporal degradation of Rad17 by Cdh1/APC (ubiquitin-protein ligase) is involved in terminating checkpoint signaling after the completion of the checkpoint response, and that stabilization of Rad17 through functional interference by a non-degradable Rad17 mutant attenuates checkpoint termination and the removal of DNA-damaged cells. We have further observed that impaired proteolysis and persistently elevated Rad17 is a conspicuous feature of malignant melanomas, and that the level of persistence parallels the transition from normal melanocyte through an increasing degree of dysplasia and ultimately to melanoma itself. In addition, stabilized Rad17 impedes genomic integrity, which induces malignant transformation for human primary melanocytes. Our objective in the proposed study is to elucidate details underlying Rad17-mediated checkpoint dysregulation, and examine its impact to the process of melanomagenesis. We seek to assess the extent to which Rad17 persistence may play a pathogenetic role in some cancers. Based on the evidence, we hypothesize that failure of checkpoint regulation is an important facilitator of the development of melanomas and possibly other cancers as well. Our Specific Aims are the following: (1) To determine the upstream mechanism that regulates Cdh1 and further examine how the interplay between Cdh1/APC and Rad17 terminates checkpoint signaling; (2) to study how Rad17 is regulated by Cdh1/APC during the DNA damage response; and (3) to determine the impact of dysregulated Rad17 proteolysis on carcinogenesis using an established melanoma mouse model and 3D skin reconstruction model. Understanding the biological mechanisms involved in genomic integrity via Rad17 proteolytic regulation and elucidating a potential role for Rad17 proteolysis in melanoma development will be an important antecedent to future studies on the role of impaired DNA damage signaling in oncogenesis, as well as for efforts to identify potential targets for therapeutic intervention.

描述(由申请人提供)：DNA损伤检查点规则的失败会使细胞易于癌变。泛素-蛋白小体系统(UPS)被认为在调节DNA损伤和DNA修复反应中发挥重要作用。在试图进一步定义这一角色时，我们观察到调控机制的一个关键组成部分Rad17是一种紫外线诱导和泛素依赖的快速周转蛋白。我们观察到，CDH1/APC(泛素蛋白连接酶)对Rad17的瞬时降解参与了在检查点反应完成后终止检查点信号的过程，而不可降解的Rad17突变体通过功能干扰稳定了Rad17，从而减弱了检查点的终止和DNA损伤细胞的移除。我们进一步观察到，蛋白分解受损和持续升高的Rad17是恶性黑色素瘤的一个显著特征，这种持续水平与从正常黑素细胞到日益严重的异型增生并最终向黑色素瘤本身的转变平行。此外，稳定的Rad17阻碍了基因组的完整性，从而诱导了人类原代黑素细胞的恶性转化。我们在这项拟议的研究中的目标是阐明Rad17介导的检查点失调的细节，并检查其对黑色素瘤发生过程的影响。我们试图评估Rad17的持久性在某些癌症中可能在多大程度上发挥致病作用。根据证据，我们假设检查点调控的失败是黑色素瘤和其他癌症发展的重要促进因素。我们的具体目标如下：(1)确定调控CDH1的上游机制，并进一步研究CDH1/APC和Rad17之间的相互作用如何终止检查点信号；(2)研究在DNA损伤反应中，CDH1/APC如何调控Rad17；以及(3)通过建立黑色素瘤小鼠模型和3D皮肤重建模型，确定调控失调的Rad17蛋白分解对肿瘤发生的影响。了解通过Rad17蛋白降解调节参与基因组完整性的生物学机制，并阐明Rad17蛋白降解在黑色素瘤发生中的潜在作用，将是未来研究受损的DNA损伤信号在肿瘤发生中的作用以及努力确定治疗干预的潜在靶点的重要前提。