Understanding DNA break repair pathway choice regulation by the cNHEJ inhibitor CYREN

了解 cNHEJ 抑制剂 CYREN 的 DNA 断裂修复途径选择调节

基本信息

批准号：
10397557
负责人：
Jan Karlseder
金额：
$ 43.13万
依托单位：
SALK INSTITUTE FOR BIOLOGICAL STUDIES
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-06-06 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10397557
关键词：
Affect Alleles Alternative Splicing Amino Acids Applications Grants Binding Cell Cycle Cell Cycle Deregulation Cell Cycle Regulation Cell Cycle Stage Cell Survival Cell physiology Cells Complex DNA DNA Damage DNA Double Strand Break DNA Repair DNA Repair Pathway Dominant-Negative Mutation Double Strand Break Repair Ensure Essential Amino Acids Event Excision Exposure to G2 Phase G22P1 gene Generations Genome Genome Stability Genomic Instability Genotoxic Stress Goals Human Human Genome Infection Invaded Lead Learning Lesion Lysine Maintenance Malignant Neoplasms Maps Metabolism Modeling Modification Mutagens Mutate Nature Neoplastic Cell Transformation Nonhomologous DNA End Joining Open Reading Frames Pathway interactions Pharmaceutical Preparations Play Point Mutation Predisposition Proteasome Inhibition Protein Isoforms Proteins Regulation Role S phase Sister Chromatid Site System Therapeutic Intervention Translating XRCC5 gene base brca gene cancer cell design experimental study genetic information genome integrity genotoxicity homologous recombination inhibitor mutant novel overexpression p53-binding protein 1 polypeptide preservation prevent public health relevance repaired telomere

项目摘要

Project Summary DNA double stranded breaks (DSBs) are deleterious lesions that require rapid repair to avoid the loss of genetic information, genomic instability, neoplastic transformation and cancer formation. In human systems, the two predominant pathways for double stranded DNA break repair are canonical non-homologous end joining (cNHEJ) and homologous recombination (HR). The cNHEJ machinery recognizes breaks, indiscriminately joins them independent of sequence context and is therefore considered error prone and potentially genotoxic. HR relies on resection of the 5’ strand with the generation of single stranded 3’ overhangs, which invade homologous sister chromatids to promote error-free break repair. Choice between HR and cNHEJ depends primarily on the cell cycle stage and the nature of the break. During G1 of the cell cycle HR is inhibited by RIF1 and 53BP1, which prevent the required BRCA1/2 complex assembly and end resection for HR initiation. During S and G2, when sister chromatids are available as a template for HR, both cNHEJ and HR pathways can be employed and compete to repair DSBs. End resection is activated by CtIP in S and G2 phases and promotes HR, but it is unclear how the abundant and efficient cNHEJ machinery is suppressed in S and G2 to allow resection at break sites and commencement of HR, thereby ensuring error- free repair of lesions to preserve genome integrity. CYREN (Cell cYcle REgulator of NHEJ) was originally identified in a screen for potential modulators of retroviral infection. Later, an alternatively spliced isoform of CYREN, CYREN-2, was found as short open reading frame translated polypeptide and shown to interact with the Ku70/80 heterodimer, pointing at a potential role in cNHEJ. Here it is proposed to investigate the discovery that CYREN modulates the cell cycle dynamics of cNHEJ and that the small protein is a direct cell cycle regulator of cNHEJ. In three specific aims it is proposed to investigate the mechanism of how CYREN inhibits cNHEJ through the CYREN interaction with the Ku heterodimer complex (AIM 1), how CYREN is cell cycle regulated and controls the cell cycle regulation of DSB repair pathway choice (AIM2) and finally what the effects of CYREN deletion and overexpression are, how the deregulation of cell cycle control of cNHEJ influences genome maintenance and genome instability and whether cells that lack CYREN are sensitive to DNA damage causing agents (AIM 3). In summary, this grant proposal focuses on CYREN, a novel regulator of DNA repair pathway choice, the mechanism of how CYREN is regulated and controls cNHEJ, whether lack of CYREN causes genome instability and whether CYREN targeting can be exploited to sensitize cancer cells to treatment with genotoxic agents.

项目概要 DNA 双链断裂 (DSB) 是有害的损伤，需要快速修复以避免丢失在人类系统中，遗传信息、基因组不稳定性、肿瘤转化和癌症形成。双链 DNA 断裂修复的两个主要途径是典型的非同源末端连接 (cNHEJ) 和同源重组 (HR) cNHEJ 机制可识别断裂、不加区别地连接它们，与序列上下文无关，因此被认为容易出错，并且 HR 依赖于切除 5' 链并生成单链 3' 链。突出，侵入同源姐妹染色单体以促进无错误的断裂修复。 HR 和 cNHEJ 主要取决于细胞周期阶段和细胞 G1 期间的断裂性质。 HR 周期被 RIF1 和 53BP1 抑制，从而阻止所需的 BRCA1/2 复合物组装和终止在 S 和 G2 期间，当姐妹染色单体可用作 HR 模板时，进行切除。 cNHEJ 和 HR 通路可被 CtIP 激活并竞争修复 DSB。 S和G2阶段并促进HR，但尚不清楚cNHEJ机制如何丰富和高效在 S 和 G2 中被抑制，以允许在断裂部位进行切除并开始 HR，从而确保错误 CYREN（NHEJ 细胞周期调节器）最初是对病变进行自由修复以保持基因组完整性。后来在筛选逆转录病毒感染的潜在调节剂时发现了一种选择性剪接亚型。 CYREN，CYREN-2，被发现是短开放阅读框翻译的多肽，并显示与 Ku70/80 异二聚体，指出了 cNHEJ 中的潜在作用，建议对这一发现进行研究。 CYREN 调节 cNHEJ 的细胞周期动力学，并且小蛋白是直接的细胞周期在三个具体目标中，建议研究 CYREN 如何抑制的机制。 cNHEJ 通过 CYREN 与 Ku 异二聚体复合物 (AIM 1) 相互作用，CYREN 是如何影响细胞周期的调节和控制 DSB 修复途径选择 (AIM2) 的细胞周期调节，最后是什么 CYREN 缺失和过度表达的影响是，cNHEJ 细胞周期控制的失调如何影响基因组维护和基因组不稳定以及缺乏 CYREN 的细胞是否对 DNA 损伤引起剂 (AIM 3) 总之，本拨款提案重点关注 CYREN，一种新型调节剂。 DNA修复途径的选择、CYREN如何调节和控制cNHEJ的机制、是否缺乏 CYREN 导致基因组不稳定以及 CYREN 靶向是否可用于使癌细胞敏感使用基因毒性药物进行治疗。