Project 3: Chromatin modifiers of BRCA-related DNA repair Pathways

项目3：BRCA相关DNA修复途径的染色质修饰剂

• 批准号: 10599902
• 负责人: Shridar Ganesan
• 金额: $ 38.7万
• 依托单位: RUTGERS BIOMEDICAL AND HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-05-01 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10599902
• 关键词: Acetylation; Address; Affect; BMI1 gene; BRCA1 gene; BRCA2 gene; Binding; Cell Survival; Cells; Cellular biology; Chromatin; Chromatin Remodeling Factor; Clinical; Computational Biology; DNA; DNA Damage; DNA Repair; DNA Repair Pathway; Defect; Development; Drug Targeting; Engineering; Event; Gene Silencing; Genetic Transcription; Histones; Human; Impairment; Malignant Neoplasms; Mediating; Methods; Methyltransferase; Modification; Molecular Biology; Mus; Mutation; PALB2 gene; Pathway interactions; Phenotype; Phosphorylation; Play; Post-Translational Protein Processing; Proteins; Regulation; Repair Complex; Role; Serine; Signal Pathway; Signal Transduction; Site; System; Therapeutically Targetable; Transcriptional Regulation; Transferase; Ubiquitination; chromatin modification; clinically relevant; drug development; gene repression; genome editing; genomic locus; histone methylation; histone modification; inhibitor; innovation; insight; knock-down; member; mutant; p53-binding protein 1; recruit; repaired; response; small molecule; small molecule inhibitor; synthetic lethal interaction; targeted treatment; treatment response; tumorigenesis

PROJECT SUMMARY/ABSTRACT: Induction of a DNA break initiates DNA damage signaling pathways that lead to coordinated and dynamic modification of the local chromatin landscape, including regulated changes in post-translational modification of histones flanking the site of DNA damage. These chromatin changes are required to recruit repair complexes to the site of damage, and repress transcription of genes flanking the break during repair. We recently found that euchromatic histone methyl transferase 2 (EHMT2, also known as G9a) plays a crucial role in regulating DNA repair. EHMT2 is rapidly recruited to sites of DNA breaks in a manner dependent on its phosphorylation by ATM at serine 569. Interestingly, the methyl-transferase activity of EHMT2 is required for the early, H2AX- independent recruitment of DNA repair factors including RNF8, 53BP1 and BRCA1; We have also recently found that EHMT2 activity is required for normal regulation of key features of ATM-mediated induction of transcriptional silencing near a DNA break. This includes regulation and recruitment of key silencing factors such as BMI1 and CDYL. These findings suggest that EHMT2 has a key role in regulating both DNA repair efficiency and modulating transcriptional regulation near a DNA break. As small molecule inhibitors of EHMT2 are now being developed for clinical use, EHMT2 may be one of the components of the DNA repair pathway that may be therapeutically targetable. We hypothesize that EHMT2 activity during the DNA damage response is 1) required to mediate local transcriptional silencing by regulating both histone and non-histone substrates 2) enables the early, H2AX independent, recruitment of DNA repair factors to sites of DNA damage through the action of key binding partners and substrates and 3) is critical for cell survival in the presence of specific cancer-associated DNA repair defects. These hypotheses will be investigated through the following set of specific aims. Aim 1. Determine the role of EHMT2 in DNA-damage-induced transcriptional silencing. Aim 2: Determine role of EHMT2 in H2AX-indpendent recruitment of DNA repair factors. Aim 3: Define synthetic lethal interactions of EHMT2 in DNA repair and chromatin regulation. To address these aims we will use carefully interrogate the effect of EHMT2 loss and inhibition on multiple aspects of DNA repair and local chromatin landscape near a DNA break. This will be done in cell and mouse systems where induction of DNA damage is engineered at specific genomic locations, and the effect of DNA damage on local chromatin landscape and recruitment of DNA repair factors can be systematically interrogated. We will also determine how EHMT2 inhibition using small molecule catalytic inhibitors perturbs repair pathway and survival in both wt cells and cells harboring specific, clinically relevant mutations in DNA repair pathways, including ATM, BRCA1, PALB2 and BRCA2. By this approach we will gain new insight into the fundamental role of EHMT2 in regulating DNA repair, and develop synthetic lethal strategies to target cancers harboring specific defects in DNA repair or chromatin regulation with EHMT2 inhibitors.

项目摘要/摘要： DNA断裂的诱导启动DNA损伤信号通路，导致协调和动态 修改局部染色质景观，包括调节后翻译后修饰的变化 组蛋白侧面DNA损伤部位。这些染色质更改需要招募维修复合物 到损害部位，并阻止修复过程中断裂的基因转录。我们最近发现 正式组蛋白甲基转移酶2（EHMT2，也称为G9A）在调节中起着至关重要的作用 DNA修复。 EHMT2迅速以取决于其磷酸化的方式招募到DNA断裂部位 通过丝氨酸569的ATM。有趣的是，EHMT2的甲基转移酶活性是早期H2AX-需要 独立募集DNA修复因子，包括RNF8，53BP1和BRCA1；我们最近也有 发现正常调节ATM介导的关键特征需要EHMT2活性 DNA断裂附近的转录沉默。这包括关键沉默因素的监管和招募 例如BMI1和CDYL。这些发现表明EHMT2在调节两个DNA修复中具有关键作用 DNA断裂附近的效率和调节转录调节。作为EHMT2的小分子抑制剂 现在正在开发用于临床使用，EHMT2可能是DNA修复途径的组成部分之一 这可能是治疗目标的。我们假设在DNA损伤响应中EHMT2活性 是1）通过调节组蛋白和非历史程序底物2）需要调节局部转录沉默。 使早期的H2AX独立，将DNA修复因子募集到DNA损伤部位 关键结合伴侣和底物的作用以及3）对于在特定的存在下对细胞存活至关重要 癌症相关的DNA修复缺陷。这些假设将通过以下一组进行调查 具体目标。 AIM 1。确定EHMT2在DNA损伤诱导的转录沉默中的作用。目标2： 确定EHMT2在H2AX-Indpentent募集DNA修复因子中的作用。目标3：定义合成致命 EHMT2在DNA修复和染色质调节中的相互作用。为了解决这些目标，我们将仔细使用 询问EHMT2损失和抑制对DNA修复和局部染色质多个方面的影响 DNA断裂附近的景观。这将在诱导DNA损伤的细胞和小鼠系统中完成 在特定的基因组位置设计，以及DNA损伤对局部染色质景观和 可以系统地询问DNA修复因子的募集。我们还将确定如何EHMT2 使用小分子催化抑制剂的抑制作用perturbs修复途径和WT细胞中的存活率 在DNA修复途径中具有特定临床相关突变的细胞，包括ATM，BRCA1，PALB2 和BRCA2。通过这种方法，我们将获得有关EHMT2在调节DNA中的基本作用的新见解 修复并制定合成的致命策略，以针对具有DNA修复中具有特定缺陷的癌症或 用EHMT2抑制剂调节染色质。