CRL3KEAP1 complex licenses genotoxic stress-induced tumor cell death

CRL3KEAP1复合物许可基因毒性应激诱导的肿瘤细胞死亡

• 批准号: 10378636
• 负责人: Yi-Nan Gong
• 金额: $ 19.72万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10378636
• 关键词: Antioxidants; Apoptosis; Apoptosis Inhibitor; Apoptotic; BIRC4 gene; CASP8 gene; CRISPR screen; Cell Death; Cell Line; Cell Survival; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Colorectal Neoplasms; Complex; DNA Damage; Data; Genotoxic Stress; HT29 Cells; Human; Knock-out; Knowledge; Licensing; Link; Malignant Neoplasms; Mediating; Mediator of activation protein; Membrane; Modeling; Molecular; Mutate; Mutation; Necrosis; Neoplasm Metastasis; Pathway interactions; Phosphorylation; Phosphotransferases; Positioning Attribute; Predisposition; Protein Kinase; Proteins; RIPK1 gene; RIPK3 gene; Radiation therapy; Reagent; Reporting; Resistance; Signal Pathway; Signaling Molecule; System; TNF gene; TP53 gene; Testing; UV induced; base; cancer cell; cancer therapy; cell killing; chemotherapy; genome-wide; genotoxicity; insight; neoplastic cell; novel; overexpression; refractory cancer; response; screening; therapy resistant; tumor; tumor progression; ubiquitin-protein ligase

PROJECT SUMMARY/ABSTRACT This application is responsive to PA-17-449: The Interplay of Cell Death Pathways in Cancer Cell Survival and Resistance to Therapy (R21). In general, genotoxic stress triggers apoptosis if tumor cells express functional p53. Genotoxic stress can also induce p53 independent apoptosis due to the auto-depletion of XIAP and cIAP1/2, which leads to the formation of “ripoptosome” complex. Ripoptosome can stimulate caspase-8-mediated apoptosis. Mutated p53 and caspase-8 have been frequently detected in human cancers. Caspase-8 mutation or inactivation often leads to resistance to apoptosis. However, in a few cases, it renders cells highly susceptible to another form of programmed cell death, termed “necroptosis.” Kinase RIPK3 and its substrate MLKL are the critical players for necroptosis. Although several studies have indicated that DNA damage can induce necroptosis (or undefined necrosis), the molecular mechanism is largely unknown. Herein, we have used an unbiased genome-wide CRISPR knockout approach to identify new players in DNA damage-induced necroptosis. We identified Cullin3 (Cul3)-RING E3 ubiquitin Ligase (CRL3KEAP1) complex as a promising candidate that mediated DNA damage-induced necroptosis. Our working model is that CRL3KEAP1 complex needs to degrade an unidentified substrate(s) to let the progression of DNA damage-induced necroptosis, thus licensing genotoxicity-induced cell death. We will make efforts to test this hypothesis in this proposal. We will determine what the CRL3KEAP1 substrate(s) is(are) and how it(they) can halt cell death. The proposed studies will provide new mechanistic insights on DNA damage agents induced cell death. Our ongoing efforts have the potential to change how we trigger cell death in tumors, especially in those with p53, caspase-8, and/or KEAP1 mutations.

项目摘要/摘要 该应用程序响应PA-17-449：癌细胞中细胞死亡途径的相互作用 生存和对治疗的抵抗(R21)。一般来说，如果肿瘤发生，基因毒性应激会触发细胞凋亡。 细胞表达功能性的P53。基因毒性应激也可以诱导P53非依赖的细胞凋亡，这是由于 XIAP和cIAP1/2的自身耗竭导致核小体复合体的形成。 Ripoposome可刺激caspase-8介导的细胞凋亡。突变的p53和caspase-8 在人类癌症中经常被检测到。Caspase-8突变或失活通常会导致 对细胞凋亡的抵抗。然而，在少数情况下，它使细胞对另一种细胞高度敏感 程序性细胞死亡的一种形式，称为“坏死性下垂”。激酶RIPK3及其底物MLKL是 坏死性下垂的关键因素。尽管多项研究表明，DNA损伤可以 导致坏死性下垂(或不明坏死)，其分子机制很大程度上是未知的。在这里， 我们使用了一种无偏见的全基因组CRISPR基因敲除方法来识别新的参与者 DNA损伤导致的坏死性下垂。我们鉴定了Cullin3(CUL3)-环E3泛素连接酶 (CRL3KEAP1)复合体作为一种有希望的候选分子，可介导DNA损伤引起的坏死性下垂。 我们的工作模型是CRL3KEAP1复合体需要将一个未知的底物(S)降解到 让DNA损伤导致坏死性下垂的进展，从而许可遗传毒性诱导的细胞 死亡。我们将努力在这一提案中检验这一假设。我们将确定什么是 CRL3KEAP1底物(S)是(ARE)及其如何阻止细胞死亡。拟议的研究将 为DNA损伤剂诱导细胞死亡提供新的机制见解。我们正在进行的努力 有可能改变我们在肿瘤中触发细胞死亡的方式，特别是在那些携带p53基因的肿瘤中， Caspase-8和/或Keap1突变。