Understanding the molecular basis of checkpoint response during DNA double-strand break repair

了解 DNA 双链断裂修复过程中检查点反应的分子基础

• 批准号: MR/Y001192/1
• 负责人: Xiaodong Zhang
• 金额: $ 259.76万
• 依托单位: Imperial College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2024
• 资助国家: 英国
• 起止时间: 2024 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FY001192%2F1
• 关键词: Understanding; molecular; basis; checkpoint; response

Our genomic information is stored in DNA, which has a double helical structure and is organised into nucleosomes and chromosome. However our DNA suffers from constant assaults from both external and internal sources such as UV/ionizing radiation, chemicals from smoke and drugs as well as molecules produced from our body's normal metabolic activities. Some of the assaults lead to severe DNA damages such as a double strand break (DSB), when both DNA strands are broken and therefore cells can't use the intact strand as a template for repair. If unrepaired or misrepaired, a DSB can result in changes in our DNA that lead to cell death or permanent changes in our genes. These factors contribute to aging and other human diseases such as cancer. Fortunately our cells have developed several ways to repair DSBs, especially to ensure they are repaired before our cells duplicate and pass our genes to next generation of cells. We want to study how this is achieved. Currently we know that three very large protein molecules called DNA-PKcs, ATM and ATR are the master coordinators. They modify other protein molecules (phosphorylate them) to enable them to carry out repair and to ensure the cells are slowed down or stopped progressing to the next stage, until the repair is complete. How precisely these master coordinators carry out these complex tasks is currently unknown. We plan to study ATM and ATR, both are involved in the process that carries out DNA repair faithfully. We want to find out how they are recruited to the damaged site (in the context of nucleosomes and chromosome), how they are activated by other factors and how they then modify other protein substrates that lead to slow down or halt cell cycle progession. We will use purified molecules to assemble these complexes and to study them using cutting edge methodolgies such as cryo electron microscopy, which allow us to obtain high resolution 3-dimensional structures of molecules and their complexes. These structures, complemented by biochemical, biophysical and cellular studies, will inform us the molecular details on how they are recruited to a damaged DNA within the chromosome, how other factors (activators) change their structrues to enable them to modify their substrates and finally how they act on their substrates efficiently, especially some of them are distally located. Our work will provide crucial information on these important large molecules, fill in a critical knowledge gap on how our cell cycle progress is controlled upon a DSB, can have profound therapeutic implications. Both proteins are tumor suppressors and mutations are found in cancer patients so our mechanistic understanding will provide molecular explanation for how these mutations increase cancer development. Further, these proteins are validated drug targets and our knowledge will help with new avenues for future drug development.

我们的基因组信息存储在DNA中，DNA具有双螺旋结构，并组织成核小体和染色体。然而，我们的DNA受到来自外部和内部来源的持续攻击，如紫外线/电离辐射、烟雾和药物的化学物质以及我们身体正常代谢活动产生的分子。一些攻击会导致严重的DNA损伤，如双链断裂(DSB)，即两条DNA链都断裂，因此细胞不能使用完整的链作为修复的模板。如果没有修复或错误修复，DSB可能会导致我们的DNA发生变化，从而导致细胞死亡或我们的基因发生永久性变化。这些因素会导致衰老和其他人类疾病，如癌症。幸运的是，我们的细胞已经开发出几种修复DSB的方法，特别是确保在我们的细胞复制并将我们的基因传递给下一代细胞之前修复它们。我们想研究如何实现这一点。目前我们知道三个非常大的蛋白质分子DNA-PKcs、ATM和ATR是主协调者。它们修饰其他蛋白质分子(使它们磷酸化)，使它们能够进行修复，并确保细胞减慢或停止进入下一阶段，直到修复完成。目前尚不清楚这些主协调人如何准确执行这些复杂的任务。我们计划研究ATM和ATR，两者都参与了忠实地进行DNA修复的过程。我们想弄清楚它们是如何被招募到受损部位(在核小体和染色体的背景下)的，它们是如何被其他因素激活的，以及它们是如何修改其他蛋白质底物从而减缓或停止细胞周期进展的。我们将使用纯化的分子组装这些络合物，并使用冷冻电子显微镜等尖端方法对其进行研究，这使我们能够获得分子及其络合物的高分辨率三维结构。这些结构，辅以生化、生物物理和细胞研究，将告诉我们关于它们如何被招募到染色体内受损的DNA上的分子细节，其他因子(激活剂)如何改变它们的结构以使它们能够修改它们的底物，最后它们如何有效地作用于它们的底物，特别是其中一些位于远端。我们的工作将提供关于这些重要大分子的关键信息，填补关于我们的细胞周期进程如何受DSB控制的关键知识空白，可能具有深远的治疗意义。这两种蛋白质都是肿瘤抑制因子，而且在癌症患者中发现了突变，因此我们对机制的理解将为这些突变如何促进癌症的发展提供分子解释。此外，这些蛋白质是有效的药物靶点，我们的知识将有助于为未来的药物开发开辟新的途径。