Balancing Dissolution and Resolution / Finding a Solution

平衡溶解和溶解/寻找解决方案

• 批准号: MR/X018547/1
• 负责人: Wojciech Niedzwiedz
• 金额: $ 69.18万
• 依托单位: Institute of Cancer Research
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FX018547%2F1
• 关键词: Balancing; Dissolution; Resolution; Finding; Solution

During every cell division, the genetic code must be accurately copied and separated between two daughter cells. Faithful division of the genome is vital to prevent damage during cell division. The processes which govern this process are conserved throughout evolution. Furthermore, there is a greater imperative for the accuracy of these processes in complex multicellular organisms, such as humans, who rely on the cooperation between functional organs and tissues. Errors in these mechanisms in the early stages of development may lead to dysfunction or prove lethal to the organism. Despite this, our current understanding of the fundamental processes that untangle and separate the duplicated genome remains poor. Cells that cannot resolve entangled DNA develop 'chromatin bridges' and these bridges may lead to the loss or rearrangement of genomic information leading to changes in the number or structure of chromosomes (chromosomal instability). Critical in safeguarding chromosomal stability is the multi-functional protein TOPBP1, often highly expressed in cancers and essential to survival, which holds many important protein interactions throughout the cell cycle. Importantly, loss of TOPBP1 leads to dysfunction in cell cycle progression and also the stability of the genome. Recently, we determined that TOPBP1 holds interactions in dividing cells with the multi-protein complexes the 'BTRR' and 'SMX 'complexes. These protein complexes are associated with the repair of persistent DNA replication and DNA repair intermediates that intertwine sister-chromatids in late stages of the cell cycle, as such these complexes enable the separation of the chromatin between daughter cells and prevent genome damage. As such, the present project aims to characterise the role of TOPBP1 in the regulation of BTRR and SMX functions, at precise stages throughout the cell cycle and also at specific positions within the genome. This is to determine if these interactions are spatially or temporally distinct or local competition is regulated by TOPBP1. To achieve this we will use previously generated cell line models that are defective for interactions between TOPBP1 and SMX or BTRR to aid in delineating their functional role. These cell lines will be examined with use of state of the art microscopy approaches, to precisely track changes in the recruitment of components of the BTRR and SMX complexes and also characterise unique consequences to chromosomal stability at specific stages in the cell cycle and to specific positions in the genome. Further to this, we will explore the signalling mechanisms that regulate the disentanglement of chromatin and also safeguard chromosomal stability. This highly collaborative project will work with the Chan laboratory to employ high resolution microscopy approaches to visualise DNA entanglements that persist into the final stages of the cell cycle, as so called 'chromatin bridges and with the Pearl laboratory we will employ in vitro biology approaches to aid in the identification of the precise interaction surfaces of key proteins involved in the signalling and regulation of the TOPBP1, SMX and BTRR functions to facilitate disentanglement of the intertwined genome. Continued collaboration with the Choudhary laboratory will aid in the assessment of changes in TOPBP1 protein interactions and signalling events that may be involved in a common mechanism of chromosomal disjunction. To translate the fundamental biological findings of these studies to potential benefit to patients we will also determine if disrupting these processes improves the effectiveness of established clinically relevant anti-cancer therapies.This innovative research project aims to provide new insight into the key processes that govern cell cycle progression, DNA repair and cell division, providing new scope for the development of novel rationale for the treatment of cancer and other genetic diseases.

在每次细胞分裂过程中，遗传密码必须在两个子细胞之间准确复制和分离。基因组的忠实分裂对于防止细胞分裂期间的损伤至关重要。控制这个过程的过程在整个进化过程中是保守的。此外，在复杂的多细胞生物体中，如人类，这些过程的准确性更加迫切，他们依赖于功能器官和组织之间的合作。在发育的早期阶段，这些机制的错误可能导致功能障碍或证明对生物体是致命的。尽管如此，我们目前对解开和分离重复基因组的基本过程的理解仍然很差。不能解析缠结的DNA的细胞形成“染色质桥”，这些桥可能导致基因组信息的丢失或重排，从而导致染色体数量或结构的变化（染色体不稳定性）。保护染色体稳定性的关键是多功能蛋白质TOPBP1，它通常在癌症中高度表达，对生存至关重要，在整个细胞周期中具有许多重要的蛋白质相互作用。重要的是，TOPBP1的缺失导致细胞周期进程的功能障碍以及基因组的稳定性。最近，我们确定TOPBP1在分裂细胞中与多蛋白复合物“BTRR”和“SMX”复合物相互作用。这些蛋白质复合物与持续DNA复制和DNA修复中间体的修复相关，所述DNA修复中间体在细胞周期的后期阶段干扰姐妹染色单体，因此这些复合物能够分离子细胞之间的染色质并防止基因组损伤。因此，本项目旨在确定TOPBP1在整个细胞周期的精确阶段以及基因组内的特定位置调节BTRR和SMX功能中的作用。这是为了确定这些相互作用是否在空间或时间上不同，或者局部竞争是否受TOPBP1调节。为了实现这一目标，我们将使用先前生成的细胞系模型，这些模型在TOPBP1与SMX或BTRR之间的相互作用方面存在缺陷，以帮助描述它们的功能作用。将使用最先进的显微镜方法检查这些细胞系，以精确跟踪BTRR和SMX复合物组分募集的变化，并观察细胞周期特定阶段染色体稳定性和基因组特定位置的独特结果。除此之外，我们还将探索调节染色质解开并保护染色体稳定性的信号传导机制。这个高度合作的项目将与Chan实验室合作，采用高分辨率显微镜方法来可视化持续到细胞周期最后阶段的DNA缠结，即所谓的“染色质桥”，并与Pearl实验室合作，我们将采用体外生物学方法来帮助识别参与TOPBP1信号传导和调节的关键蛋白质的精确相互作用表面，SMX和BTRR的功能是促进缠结基因组的解开。与Choudhary实验室的持续合作将有助于评估TOPBP1蛋白相互作用和信号事件的变化，这些变化可能涉及染色体分离的共同机制。为了将这些研究的基本生物学发现转化为对患者的潜在益处，我们还将确定破坏这些过程是否会提高已建立的临床相关抗癌疗法的有效性。这个创新的研究项目旨在为控制细胞周期进程，DNA修复和细胞分裂的关键过程提供新的见解，为开发治疗癌症和其他遗传疾病的新原理提供了新的范围。