The regulation of chromosome replication by DBF4-Dependent Kinase

DBF4依赖性激酶对染色体复制的调节

• 批准号: 2883205
• 金额: --
• 依托单位: University of Dundee
• 依托单位国家: 英国
• 项目类别: Studentship
• 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=studentship-2883205
• 关键词: regulation; chromosome; replication; DBF4; Dependent

Defects in DNA replication during the process of cell division can lead to long-term problems, including loss of genome integrity, irreparable DNA damage, cell cycle exit and senescence. The project will address how genome integrity is ensured during cell division, in particular how chromosomal DNA replication is controlled and coordinated with other cell cycle events. An important regulator of DNA replication is the DBF4-Dependent Kinase (DDK) which controls replication initiation and coordinates it with other cell division events such as sister chromatid cohesion. In the last few years our lab has made some important advances in understanding the function of DDK and how it regulates DNA replication (see Volpi et al, Open Biol. 11: 210121; Alver et al, Cell Reports 8: 2508-2520; and Poh et al, Open Biology 4: 130138). The project will build on this work to understand in more detail the way that DDK regulates the activity of key replication initiation proteins and coordinates this with chromosome cohesion. The research will primarily use Xenopus (frog) egg extracts that support all of the key nuclear events of the cell division cycle in vitro, which provide an unparalleled system to study genome integrity at a biochemical level. Results obtained from the Xenopus system have been shown to be widely applicable to other vertebrate cell types. Published results show that in the Xenopus system, DDK is required for both replication initiation and the establishment of sister chromatid cohesion, though the precise biochemical mechanisms are not well understood. We have an extensive range of reagents to study DDKs and their potential substrates, including specific antibodies for immunoprecipitation and immunodepletion as well as highly specific DDK small molecule inhibitors. We will use these reagents in combination with the world-leading mass spectrometry facility in Dundee (http://proteomics.lifesci.dundee.ac.uk) to identify novel DDK substrates that are involved in regulating DNA replication and the establishment of sister chromatid cohesion. We will use this information to build hypotheses about how these processes are coordinated and then test them in the Xenopus cell-free system. We will aim to take the key conclusions obtained from the in vitro work and examine whether they are conserved in human tissue cells.

细胞分裂过程中DNA复制的缺陷会导致长期的问题，包括基因组完整性的丧失、不可修复的DNA损伤、细胞周期退出和衰老。该项目将解决如何在细胞分裂过程中确保基因组完整性，特别是如何控制染色体DNA复制并与其他细胞周期事件协调。DNA复制的一个重要调节因子是DBF 4依赖性激酶（DDK），它控制复制起始并与其他细胞分裂事件（如姐妹染色单体凝聚）协调。在过去的几年中，我们的实验室在理解DDK的功能以及它如何调节DNA复制方面取得了一些重要的进展（参见Volpi等人，Open Biol.11：210 - 121; Alver等人，Cell Reports 8：2508-2520;以及Poh等人，Open Biology 4：130 - 138）。该项目将建立在这项工作的基础上，更详细地了解DDK调节关键复制起始蛋白活性的方式，并与染色体凝聚力协调。该研究将主要使用非洲爪蟾（青蛙）卵提取物，这些卵提取物支持体外细胞分裂周期的所有关键核事件，这为在生物化学水平上研究基因组完整性提供了无与伦比的系统。从非洲爪蟾系统获得的结果已被证明是广泛适用于其他脊椎动物细胞类型。已发表的结果表明，在非洲爪蟾系统中，DDK是复制启动和建立姐妹染色单体凝聚力所必需的，尽管精确的生化机制还不清楚。我们拥有广泛的试剂来研究DDK及其潜在底物，包括用于免疫沉淀和免疫耗竭的特异性抗体以及高度特异性的DDK小分子抑制剂。我们将使用这些试剂与世界领先的邓迪质谱设施（http：//www.example.com）相结合，以确定参与调节DNA复制和建立姐妹染色单体凝聚力的新型DDK底物。proteomics.lifesci.dundee.ac.uk我们将利用这些信息来建立关于这些过程是如何协调的假设，然后在非洲爪蟾无细胞系统中进行测试。我们的目标是从体外工作中获得的关键结论，并检查它们是否在人体组织细胞中保守。