Mechanisms of EXO1 regulation in response to radiation-induced DNA damage

EXO1 响应辐射引起的 DNA 损伤的调节机制

• 批准号: 9926813
• 负责人: Sandeep Burma
• 金额: $ 30.49万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-11-28 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9926813
• 关键词: Back; Binding; Cell Cycle; Cell Survival; Cells; Clinical Trials; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; Double Strand Break Repair; EXO1 gene; Enzymes; Event; Excision; Exercise; G2 Phase; Genomic Instability; Genomics; Goals; Human; In Vitro; Ionizing radiation; Lesion; Maintenance; Malignant - descriptor; Malignant Neoplasms; Modification; Molecular; Nonhomologous DNA End Joining; Pathway interactions; Phase; Phosphorylation; Play; Post-Translational Protein Processing; Process; Proteins; Publishing; Radiation; Radiation Induced DNA Damage; Radiation therapy; Radiosensitization; Regimen; Regulation; Research; Resistance; Role; S Phase; Site; Testing; Therapeutic; Ubiquitin; Ubiquitination; Work; base; cancer cell; cancer therapy; experimental study; genome integrity; homologous recombination; in vivo; inhibitor/antagonist; novel; preservation; prevent; public health relevance; radiation response; recruit; repaired; response; spleen exonuclease

 DESCRIPTION (provided by applicant): Ionizing radiation (IR) remains one of the mainstays of cancer therapy. The most deleterious lesion induced by IR is the DNA double-strand break (DSB). Accurate repair of DSBs is essential for preventing loss of genomic integrity and malignant transformation. Efficient repair of DSBs also underlies the resistance of many cancers to radiation therapy. A cell must choose between two major repair pathways to fix these breaks - non- homologous end joining (NHEJ), an error-prone pathway that is operative in all phases of the cell cycle or homologous recombination (HR), an error-free pathway that is restricted to the post-replicative phases of the cell cycle. Optimal usage of these two pathways is vital for the maintenance of genomic integrity and cell survival in the face of genomic insults. The DNA end resection step of HR is a pivotal point at which correct repair pathway choice is exercised. Importantly, research from our lab and others has established that the 5' to 3' exonuclease EXO1 is a critical player in DNA end resection and repair pathway choice in human cells. While DNA end resection is currently an avidly researched topic in the field of DNA repair, the exact sequence of molecular events involving EXO1 that allows commitment to a particular repair pathway is not well worked out. Exciting new results from our lab demonstrate that EXO1 is phosphorylated by CDKs 1/2 in a cell cycle- dependent manner and by ATM/ATR in a DNA damage-dependent manner to promote DNA end resection. However, soon after DNA damage, EXO1 is SUMOylated, ubiquitinated, and rapidly targeted for degradation, presumably to prevent uncontrolled resection of DNA ends. It is important to mechanistically understand how these and other post translational modifications stimulate or restrain EXO1's functions in the cellular response to IR. Towards this goal, we propose to develop a comprehensive picture of cell cycle- and IR-dependent modifications and interacting partners of EXO1, and to mechanistically understand how these modifications promote EXO1 activation and subsequent degradation in response to IR. Based upon our preliminary results, we hypothesize that EXO1 activation and inactivation is a tightly controlled process involving phosphorylation, SUMOylation and ubiquitination events that fine-tune DNA end resection, optimize DSB repair, and preserve genomic integrity. Understanding the sequence and functions of EXO1 post- translational modifications and the exact choreography of EXO1 and its interacting partners at DSBs will be of paramount importance in developing more effective radiosensitization approaches that target the critical DNA end resection step. Specifically, we propose to: 1) Test the hypothesis that phosphorylation of EXO1 by CDKs and PI3KKs regulates DNA end resection and influences repair pathway choice, 2) Test the hypothesis that EXO1 degradation post-radiation restrains DNA end resection and preserves genomic integrity, and 3) Test the hypothesis that blocking EXO1 activation with CDK 1/2 inhibitors may be a viable strategy for therapeutically sensitizing cancers to ionizing radiation.

 描述（由申请人提供）：电离辐射（IR）仍然是癌症治疗的支柱之一，IR引起的最有害的损伤是DNA双链断裂（DSB），DSB的精确修复对于防止基因组丢失至关重要。 DSB 的有效修复也是许多癌症对放射治疗产生耐药性的基础，细胞必须在两种主要修复途径之间进行选择——非同源末端连接。 (NHEJ)，一种在细胞周期的所有阶段均有效的易错途径；或同源重组 (HR)，一种仅限于细胞周期复制后阶段的无错途径。面对基因组损伤时，DNA 末端切除步骤对于维持基因组完整性和细胞存活至关重要。重要的是，我们实验室和其他人的研究已经证实这一点。这5' 至 3' 核酸外切酶 EXO1 在人类细胞 DNA 末端切除和修复途径选择中发挥着关键作用，虽然 DNA 末端切除目前是 DNA 修复领域的热门研究课题，但涉及 EXO1 的分子事件的确切序列使得这一过程成为可能。我们实验室的令人兴奋的新结果表明，EXO1 以细胞周期依赖性方式被 CDK 1/2 磷酸化，并被 DNA 中的 ATM/ATR 磷酸化。然而，在 DNA 损伤后不久，EXO1 就会被 SUMO 化、泛素化，并迅速靶向降解，这可能是为了防止 DNA 末端不受控制的切除，从机制上了解这些和其他翻译后的过程非常重要。修饰刺激或抑制 EXO1 在细胞对 IR 反应中的功能，为了实现这一目标，我们建议全面了解细胞周期和 IR 依赖性修饰以及 EXO1 的相互作用伙伴。从机制上了解这些修饰如何促进 EXO1 激活和随后响应 IR 的降解 根据我们的初步结果，我们发现 EXO1 激活和失活是一个严格控制的过程磷酸化、SUMO 化和泛素化事件，可微调 DNA 末端切除、优化 DSB。了解 EXO1 翻译后修饰的序列和功能以及 EXO1 及其在 DSB 中的相互作用伙伴的精确编排对于修复和保持基因组完整性至关重要。开发针对关键 DNA 末端切除步骤的更有效的放射增敏方法，具体来说，我们建议：1) 检验 CDK 和 PI3KK 磷酸化 EXO1 调节 DNA 末端切除并影响修复途径选择的假设，2) 检验 EXO1 的假设。辐射后降解抑制 DNA 末端切除并保持基因组完整性，并且 3) 检验用 CDK 1/2 抑制剂阻断 EXO1 激活可能是可行策略的假设治疗上使癌症对电离辐射敏感。

项目成果

A TNF-JNK-Axl-ERK signaling axis mediates primary resistance to EGFR inhibition in glioblastoma.

• DOI: 10.1038/nn.4584
• 发表时间: 2017-08
• 期刊: Nature neuroscience
• 影响因子: 25
• 作者: Guo G;Gong K;Ali S;Ali N;Shallwani S;Hatanpaa KJ;Pan E;Mickey B;Burma S;Wang DH;Kesari S;Sarkaria JN;Zhao D;Habib AA
• 通讯作者: Habib AA

MET signaling promotes DNA repair and radiation resistance in glioblastoma stem-like cells.

• DOI: 10.21037/atm.2017.01.67
• 发表时间: 2017-02
• 期刊: Annals of translational medicine
• 作者: P. Todorova;B. Mukherjee;S. Burma

EEPD1 promotes repair of oxidatively-stressed replication forks.

• DOI: 10.1093/narcan/zcac044
• 发表时间: 2023-03
• 期刊: NAR cancer
• 影响因子: 5.1

Endonuclease EEPD1 Is a Gatekeeper for Repair of Stressed Replication Forks.

• DOI: 10.1074/jbc.m116.758235
• 发表时间: 2017-02-17
• 期刊: The Journal of biological chemistry
• 作者: Kim HS;Nickoloff JA;Wu Y;Williamson EA;Sidhu GS;Reinert BL;Jaiswal AS;Srinivasan G;Patel B;Kong K;Burma S;Lee SH;Hromas RA
• 通讯作者: Hromas RA

Enhanced dependency of KRAS-mutant colorectal cancer cells on RAD51-dependent homologous recombination repair identified from genetic interactions in Saccharomyces cerevisiae.

• DOI: 10.1002/1878-0261.12040
• 发表时间: 2017-05
• 期刊: Molecular oncology
• 影响因子: 6.6
• 作者: Kalimutho M;Bain AL;Mukherjee B;Nag P;Nanayakkara DM;Harten SK;Harris JL;Subramanian GN;Sinha D;Shirasawa S;Srihari S;Burma S;Khanna KK
• 通讯作者: Khanna KK