Control and impact of meiotic DNA resection on recombination and genome stability

同源重组(HR)是一个被广泛研究的过程，因为它对生育、生物多样性和基因组稳定具有重要意义。在动物、植物和真菌(如发芽酵母模式生物酿酒酵母)中，每个细胞都含有每个染色体的两个副本：一个来自母亲，一个来自父亲。在称为减数分裂的特殊细胞分裂过程中，HR发生在这些染色体对之间，产生配子(卵子和精子)，每个配子包含父母基因的独特组合。减数分裂HR是由分布在所有亲本染色体上的大量DNA断裂引起的。减数分裂细胞利用HR修复这些DNA断裂，产生所谓的“交叉”(Cos)和“非交叉”(NCOs)，这两种方式对个体配子的遗传变异都是重要的，因此对生物多样性和作物育种是必不可少的。COS对于减数分裂过程中准确的染色体分离也是必不可少的，因为它可以防止非整倍体(一种基因组不稳定的类型，其特征是配子的染色体数目不正确)，这是不孕不育和唐氏综合症等遗传疾病的常见原因。HR也是修复自发或由环境诱导的DNA错误的关键，HR缺陷可导致遗传不稳定，从而引发癌症发展。HR的一个核心步骤是各种酶对DNA断裂的处理，这些酶降解DNA双螺旋的两条DNA链中的一条，产生单链DNA(SsDNA)的短区域。这个过程被称为“切除”。这些单链DNA区域被进化上保守的修复酶(称为RecA、RAD51和Dmc1)用来搜索基因组中其他地方存在的类似DNA序列。这些相似的序列被用作复制和修复DNA断裂的模板。切除的起始步骤(参与并委托由HR修复的DNA末端)具有很好的特征，但关闭切除的机制以及切除长度、基因组稳定性和遗传变异之间的联系尚不清楚。尽管有许多令人信服的迹象表明，切除长度将影响基因组稳定性和遗传变异，但仍是如此。例如，在恶性肿瘤全基因组测序中发现的突变簇与切除过程中出现的单链DNA上的DNA损伤有关。此外，我们未发表的工作表明，非调控切除是与减数分裂过程中异常重组相关的主要风险因素之一，产生不平衡的染色体重排，从而降低育性。在这项建议中，我们将在萌芽酵母模式生物中使用先进的全基因组技术来研究DNA切除终止是如何在基因组的所有位置受到调控的。DNA断裂激活检查点通路，延迟细胞分裂，直到DNA修复。我们将研究切除是如何受到检查点途径的调节的，以及染色体的结构特征如何影响基因组中不同位置的切除过程。重要的是，我们将使用先进的遗传学方法来研究切除范围的变化如何影响HR的机制，特别是对基因组稳定性和遗传变异的影响。我们的工作--发现科学--将对理解切除是否、如何以及为什么切除以及更广泛地说HR控制生育、生物多样性、基因组稳定性和癌症具有基本的重要性。

Homologous recombination (HR) is a widely studied process because of its fundamental importance for fertility, biodiversity, and genome stability.In animals, plants and fungi (such as the budding yeast model organism Saccharomyces cerevisiae) every cell contains two copies of each chromosome: one inherited from the mother and one from the father. During the specialised cell division called "meiosis", HR occurs between these chromosome pairs producing gametes (eggs and sperm) that each contains a unique combination of the parental genes. Meiotic HR is initiated by numerous DNA breaks spread across all the parental chromosomes. Meiotic cells use HR to repair these DNA breaks to create so-called "crossovers" (COs) and "non-crossovers" (NCOs), both of which are important for genetic variation in individual gametes, and therefore essential for biodiversity and crop breeding. COs are also essential for accurate chromosome segregation during meiosis, by preventing aneuploidy (a type of genome instability characterised by gametes with an incorrect number of chromosomes), which is a common cause of infertility and genetic disorders such as Down Syndrome. HR is also critical to repair errors in the DNA arising spontaneously or induced by the environment, and HR defects can confer genetic instability, which can then trigger cancer development.A central step of HR is the processing of DNA breaks by various enzymes that degrade one of the two DNA strands of the DNA double helix, generating short regions of single-stranded DNA (ssDNA). This process is referred to as "resection". These ssDNA regions are used by evolutionarily conserved repair enzymes (called RecA, Rad51 and Dmc1) to search for similar DNA sequences present elsewhere in the genome. These similar sequences are used as templates from which to copy, and thereby repair, the DNA breaks.The initiation step of resection (engaging and commiting the DNA end to be repaired by HR) is well characterised, but the mechanisms that switch off resection, and the link between resection length, genome stability and genetic variation is poorly understood. This is despite a number of compelling indications that resection length will impact genome stability and genetic variation. For example, clusters of mutations identified in whole-genome sequencing of malignant tumors are linked to DNA damage occurring on the ssDNA that arises during resection. Furthermore, our unpublished work indicates that deregulated resection is one of the major risk factors associated with aberrant recombination during meiosis, generating imbalanced chromosome rearrangements that reduce fertility.In this proposal we will use advanced whole-genome technologies in the budding yeast model organism to examine how DNA resection termination is regulated at all locations in the genome. DNA breaks activate checkpoint pathways that delay cell division until the DNA is repaired. We will study how resection is regulated by the checkpoint pathway, and how structural features of the chromosome affect the resection process at different locations across the genome. Importantly, we will use advanced genetic methods to examine how changes in the extent of resection influence the mechanism of HR, with particular interest in the impact on genome stability and genetic variation.Our work, "discovery science", will be of fundamental importance to understand if, how and why resection, and more broadly HR, control fertility, biodiversity, genome stability and cancer.