Dissecting BRCA1-PALB2 Activity in DNA Repair and Development

剖析 BRCA1-PALB2 在 DNA 修复和发育中的活性

• 批准号: 10388570
• 负责人: Neil Johnson
• 金额: $ 22.9万
• 依托单位: RESEARCH INST OF FOX CHASE CAN CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-05 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10388570
• 关键词: Alleles; Award; BRCA1 gene; Cell Nucleus; Chromatin; Confocal Microscopy; DNA Damage; DNA Double Strand Break; DNA Repair; Development; Equipment; Fanconi' s Anemia; Funding; Goals; Health; Human; Knowledge; Malignant neoplasm of ovary; Mechanics; Microscopy; Mutation; Nanostructures; Nuclear; PALB2 gene; Pathway interactions; Patients; Proteins; Resolution; Site; Time; homologous recombination; improved; insight; malignant breast neoplasm; mutation carrier; parent grant; recruit; repaired

PROJECT SUMMARY/ABSTRACT This submission is for a supplement award for the parent grant R01GM135293: “Dissecting BRCA1-PALB2 activity in DNA repair and development”. The parent grant is currently in its first year, making this the perfect time to upgrade our equipment. The localization of proteins to double stranded DNA breaks (DSBs) can be quantified by the presence of nuclear foci. The number of foci within a nucleus is often indicative of whether a particular protein or repair pathway is functional. A typical focus appears as a homogenous sphere under conventional widefield or confocal microscopy. The information provided does not go beyond whether foci are present or absent. In contrast, super-resolution (SR) microscopy provides high-resolution imagining, which can reveal detailed insights into the DSB-flanking chromatin topology. The ability to perform SR microscopy would greatly enhance the knowledge that can be gained from the current R01-funded project. Not only in understanding the effects of mutations on foci formation, new insights into the mechanics of the DNA damage repair machinery and the interplay between chromatin topology and DNA repair can be revealed. In the parent grant, we aim to uncover the mechanisms by which BRCA1 recruits PALB2 to sites of DNA damage. We proposed to examine the effects of BRCA1 and PALB2 mutations on DNA repair foci. However, SR microscopy has the potential to reveal so much more, beyond simply the presence or absence of foci, such as insights into the nanostructure of BRCA1-PALB2, whether there are separate or overlapping nanodomains, spatial differences, and the BRCA1-PALB2 topology relative to additional homologous recombination (HR) proteins. Moreover, the effects of BRCA1 and PALB2 patient mutations have previously not been examined for their effects on DSB-chromatin topology and nanostructure. The use of SR microscopy will provide super-resolution insight, and ultimately improve our understanding of how BRCA1 and PALB2 mutations impact human health.

项目概要/摘要 本提交内容是为了获得家长补助金 R01GM135293 的补充奖励：“解剖 BRCA1-PALB2 DNA 修复和发育活动”。家长资助目前正处于第一年，这使其成为完美的 是时候升级我们的装备了。蛋白质对双链 DNA 断裂 (DSB) 的定位可以是 通过核焦点的存在来量化。核内焦点的数量通常表明是否存在 特定的蛋白质或修复途径是有功能的。典型的焦点在以下情况下显示为同质球体： 传统的宽视野或共焦显微镜。所提供的信息不超出焦点是否是 存在或不存在。相比之下，超分辨率（SR）显微镜提供高分辨率成像，可以 揭示 DSB 侧翼染色质拓扑的详细见解。执行 SR 显微镜的能力将 大大增强了可以从当前 R01 资助的项目中获得的知识。不仅在 了解突变对病灶形成的影响，对 DNA 损伤机制的新​​见解 修复机制以及染色质拓扑和 DNA 修复之间的相互作用可以被揭示。在父级中 授予，我们的目标是揭示 BRCA1 将 PALB2 招募到 DNA 损伤位点的机制。我们 提议检查 BRCA1 和 PALB2 突变对 DNA 修复灶的影响。然而，SR 显微镜 除了焦点的存在或不存在之外，还有可能揭示更多的东西，例如对 BRCA1-PALB2 的纳米结构，无论是单独的还是重叠的纳米域，空间 差异，以及相对于其他同源重组 (HR) 蛋白的 BRCA1-PALB2 拓扑。 此外，BRCA1 和 PALB2 患者突变的影响此前尚未得到检验。 对 DSB-染色质拓扑和纳米结构的影响。 SR显微镜的使用将提供超分辨率 洞察力，最终提高我们对 BRCA1 和 PALB2 突变如何影响人类健康的理解。