Elucidating the mechanisms that govern replication restart efficiency and fidelity

在细胞分裂之前，它必须复制它的遗传物质，以便每个新的子细胞都能获得一个副本。DNA复制的这个过程是由被称为复制体的复杂的、多组分的蛋白质机器执行的，这些复制体在被称为复制起点的染色体位置组装到DNA上。这些机器解开DNA双螺旋，形成“叉形”结构，复制发生在叉形结构上。在细胞周期中，沿着每条染色体的多个起始点“起火”，每个起始点都释放出一对“复制叉子”，它们以相反的方向远离它们的起始点。当来自相邻来源的叉子合并时，DNA复制完成。即使是一对叉子未能合并，也会导致一个未复制的DNA区域，这可能导致DNA断裂，并最终导致导致癌症等疾病的基因突变。许多结合染色体DNA的蛋白质通过物理上阻碍复制叉的进程而威胁到复制的成功完成。当复制叉遇到这样的障碍时，它可能会经历一个称为叉状崩溃的过程，在这个过程中，它的复制体部分或完全从DNA上分离出来。在叉子崩溃后，细胞似乎利用两种不同的机制来确保DNA复制的完成：1)相反叉子的被动DNA复制，这可能是通过点燃附近通常处于休眠状态的起始点来促进的；以及2)通过一个鲜为人知的依赖于催化DNA重组的蛋白质的过程重新启动失败的叉子。有趣的是，重新启动复制本身可能会导致有害的突变和染色体重排，因此可能会受到严格的监管，以限制其有害影响。为了更好地了解细胞如何处理失败的复制分叉，我们的目标是确定影响复制重新启动的效率和保真度的因素。我们还旨在发现休眠的复制起点是否会对单个折叠的复制分叉做出反应，以及这是一个主动还是被动的过程。通过破译这些基本过程，我们将更好地了解癌症等疾病的分子基础，这些疾病的发展和进展往往植根于DNA复制过程中出现的问题。最终，从这个项目中获得的知识将为未来开发临床使用的新诊断和治疗方法奠定基础。

Before a cell divides it has to replicate its genetic material so that each new daughter cell can receive a copy. This process of DNA replication is performed by complex, multi-component protein machines known as replisomes, which are assembled onto DNA at chromosomal sites known as replication origins. These machines unwind the DNA double helix to form "fork" structures, at which the copying occurs. During the cell cycle, multiple origins along every chromosome "fire", each releasing a pair of "replication forks" that travel in opposite directions away from their origin. DNA replication is completed when forks from adjacent origins merge. The failure of even a single pair of forks to merge results in a region of unreplicated DNA that can lead to DNA breakage, and ultimately genetic mutations that cause diseases such as cancer. The many proteins that bind chromosomal DNA threaten successful completion of replication by physically impeding the progress of the replication fork. When a replication fork encounters such a barrier, it can undergo a process termed fork collapse in which its replisome partially or completely dissociates from the DNA. Following fork collapse, cells seem to make use of two different mechanisms to ensure completion of DNA replication: 1) passive DNA replication by an opposing fork, which may be promoted by the firing of normally dormant origins nearby; and 2) restart of the failed fork by a poorly understood process that depends on proteins that catalyse DNA recombination. Intriguingly, restarting replication can itself cause harmful mutations and chromosomal rearrangements and, therefore, is presumably strictly regulated to limit its deleterious effects. To better understand how cells deal with failed replication forks, we aim to identify the factors that affect the efficiency and fidelity of replication restart. We also aim to discover whether dormant replication origins fire in response to a single collapsed replication fork, and whether this is an active or passive process. By deciphering these fundamental processes, we will gain a better understanding of the molecular basis of diseases, such as cancer, whose development and progression are often rooted in problems that arise during DNA replication. Ultimately, the knowledge obtained from this project will underpin future endeavours to develop new diagnostics and therapeutics for clinical use.