Coordination of ATR Signaling for Genetic Quality Control, Silencing, and DNA Repair During Meiosis

减数分裂期间遗传质量控制、沉默和 DNA 修复的 ATR 信号协调

基本信息

批准号：
10172957
负责人：
Marcus Smolka
金额：
$ 42.19万
依托单位：
CORNELL UNIVERSITY
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：
2018-09-13 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10172957
关键词：
ATR checkpoint CHEK1 gene CHEK2 gene Cell Death Cells Chromatin Chromosome Pairing Chromosomes Clinical Closure by clamp Complement Complex Congenital Abnormality DNA DNA Damage DNA Repair Defect Development Double Strand Break Repair Ensure Event Failure Gene Silencing Generations Genetic Genetic Determinism Genome Germ Cells Goals Growth Haploidy Infertility Knowledge Lead Maintenance Malignant Neoplasms Mammals Mass Spectrum Analysis Mediating Meiosis Meiotic Prophase I Mitotic Molecular Monitor Mus Mutant Strains Mice Mutation Oocytes Output Pathway interactions Phenotype Phosphotransferases Process Production Proteins Proteomics Quality Control RAD9A gene Regulation Reproduction Role Scaffolding Protein Signal Transduction Somatic Cell Spermatocytes Spermatogenesis Structure TOPBP1 Gene Testis Time Work base cancer therapy design experimental study genetic analysis genome editing innovation insight mouse model mutant novel paralogous gene phosphoproteomics prevent recombinational repair repair function repaired response sex sperm cell virtual

项目摘要

Infertility and birth defects often arise due to improper genetic quality control during meiosis. To ensure the production of gametes without genetic defects, the genome of meiocytes is monitored by a set of evolutionarily conserved kinases, known as checkpoint kinases. These kinases sense damage to DNA or problems in chromosome pairing and, upon activation, can block meiotic progression and induce cell death. But the action of checkpoint kinases is not solely utilized as a quality control mechanism. Programmed double strand break (DSB) formation is an abundant and essential event for normal meiotic progression, and checkpoint kinases are required to coordinate key events in recombination repair, crossover regulation and transcriptional silencing. How meiotic checkpoint signaling is regulated is not understood, especially in mammals. In particular, little is known about how checkpoint kinases can act both as essential regulators of normal meiotic progression as well as effectors of quality control mechanisms that lead to cell death. This knowledge gap poses a major barrier for understanding the determinants of genetic quality control and how mis-regulation of checkpoint signaling may aberrantly block meiotic progression and promote infertility. The same pathways also mediate fundamental DNA repair and checkpoint functions in mitotic cells, are sometimes deregulated in cancers, and are being targeted clinically as an emerging strategy for cancer treatment. This proposal applies innovative approaches to overcome long-standing barriers for the study of meiotic checkpoint signaling in mammals. The proposed studies focus on the essential checkpoint kinase ATR, which is important for DSB repair and transcriptional silencing of unsynapsed chromatin during meiosis and is regulated in part by the scaffolding protein TOPBP1 and other upstream regulators such as the RAD9A-RAD1-HUS1 complex. Cutting edge approaches for genome editing in the mouse will be used to generate rationally designed separation-of- function mouse mutants with the goal of revealing novel checkpoint regulatory mechanisms operative in meiosis. To guide and complement genetic and functional experiments, mass spectrometry analysis of testis extracts will be used for quantitative and unbiased characterization of checkpoint signaling in spermatocytes. Collectively, these studies are expected to reveal how meiotic ATR signaling is coordinated to achieve structure-specific signaling outputs in response to unrepaired DSBs or chromosome asynapsis, without inducing cell death. Beyond providing fundamental insights into the actions of genome maintenance pathways that function in virtually all cells, the results from this work will carry important implications related to the molecular origins of infertility and birth defects as well as the impact of ATR inhibition in clinical settings.

由于减数分裂过程中遗传质量控制不当，常常会出现不孕症和出生缺陷。为了确保产生没有遗传缺陷的配子，性母细胞的基因组由一组进化上保守的激酶（称为检查点激酶）监控。这些激酶感知 DNA 损伤或染色体配对问题，一旦激活，可以阻止减数分裂进程并诱导细胞死亡。但检查点激酶的作用不仅仅被用作质量控制机制。程序性双链断裂 (DSB) 形成是正常减数分裂进程中丰富且重要的事件，检查点激酶需要协调重组修复、交叉调节和转录沉默中的关键事件。减数分裂检查点信号传导是如何调节的尚不清楚，尤其是在哺乳动物中。特别是，人们对检查点激酶如何充当正常减数分裂进程的重要调节剂以及导致细胞死亡的质量控制机制的效应器的作用知之甚少。这种知识差距为理解遗传质量控制的决定因素以及检查点信号传导的错误调节如何异常阻止减数分裂进展并促进不孕构成了主要障碍。相同的通路还介导有丝分裂细胞中的基本 DNA 修复和检查点功能，有时在癌症中失调，并作为一种新兴的癌症治疗策略在临床上受到关注。该提案应用创新方法来克服哺乳动物减数分裂检查点信号研究中长期存在的障碍。拟议的研究重点关注必需的检查点激酶 ATR，它对于减数分裂过程中 DSB 修复和非突触染色质的转录沉默非常重要，并且部分受到支架蛋白 TOPBP1 和其他上游调节因子（例如 RAD9A-RAD1-HUS1 复合物）的调节。小鼠基因组编辑的尖端方法将用于产生合理设计的功能分离小鼠突变体，目的是揭示减数分裂中起作用的新检查点调节机制。为了指导和补充遗传和功能实验，睾丸提取物的质谱分析将用于定量和公正地表征精母细胞中检查点信号传导。总的来说，这些研究有望揭示减数分裂 ATR 信号如何协调，以实现结构特异性信号输出，以响应未修复的 DSB 或染色体突触，而不诱导细胞死亡。除了提供对几乎在所有细胞中发挥作用的基因组维持途径的作用的基本见解外，这项工作的结果还将对不孕症和出生缺陷的分子起源以及 ATR 抑制在临床环境中的影响产生重要影响。