Germline mutagenesis at meiotic double-strand breaks

项目摘要/摘要 减数分裂重组对哺乳动物生殖细胞的减数分裂是必不可少的，从而对 单倍体配子的发育，即精子和卵子。重组是由数百个DNA双链启动的 链断裂(DSB)引入了由SPO11蛋白催化的全基因组。忠实传输 将基因组传递给后代需要对这些大量的DSB进行适当的修复，主要是通过 与同源基因重组。在减数分裂细胞中，DSB的形成是由ATM激酶调节的，它是 在有丝分裂细胞中是DSB的主要响应者，因此在没有ATM的情况下，减数分裂DSB 增加~10倍。 我们最近发现减数分裂DSB存在引发生殖系重排的风险，在 涉及非同源末端连接的特殊缺失和串联复制，尤其是在缺少 自动取款机。就扰乱这些热点所在的基因而言，这些事件也是重要的 作为相关的PRDM9结合位点，控制这些基因座上的重组。因此，我们的发现揭示了一个 先前隐藏的可能影响人类健康和基因组的生殖系突变潜力 进化论。在人类身上，最近对冰岛人进行的远程测序支持这种影响。这项建议旨在 旨在了解导致这些事件的机制、减数分裂DSB的事件范围以及 年龄的影响。我们假设在减数分裂DSB中可能有比我们已有的重排更多的重排 之前确认过的。因此，在第一个目标中，我们建议确定可以 产生于减数分裂双链断裂，包括远距离缺失、重复和染色体易位。在……里面 第二个目的，我们考察可能影响缺失形成的因素。我们关注的是DNA的作用 结束处理分两步进行，即SPO11去除和结束处理，以及重组。此外，我们还解决了 双链断裂附近形成的间隙是否是同源重组的底物以及父本的影响 减数分裂DSB重排事件的年龄。

Project Summary/Abstract Meiotic recombination is essential for the reductional cell division in mammalian germ cells and thus for the development of haploid gametes, i.e., sperm and eggs. Recombination is initiated by hundreds of DNA double- strand breaks (DSBs) introduced genome-wide that are catalyzed by the SPO11 protein. Faithful transmission of the genome to subsequent generations requires proper repair of these numerous DSBs, primarily through recombination with the homolog. DSB formation is regulated in meiotic cells by the ATM kinase, which is known to be a primary responder to DSBs in mitotic cells, such that in the absence of ATM, meiotic DSBs increase ~10-fold. We recently discovered that meiotic DSBs are at risk for provoking germline rearrangements, in particular deletions and tandem duplications involving nonhomologous end-joining, especially in the absence of ATM. These events are consequential in terms of disrupting the genes in which these hotspots occur as well as the associated PRDM9 binding sites that govern recombination at those loci. Thus, our findings reveal a previously hidden potential for germline mutagenesis that is likely to affect human health and genome evolution. In humans, recent long-range sequencing of Icelanders supports this impact. This proposal pursues aims to understand the mechanisms that give rise to these events, the range of events at meiotic DSBs, and the effect of age. We hypothesize that other rearrangements are possible at meiotic DSBs than what we have previously identified. Thus, in the first aim, we propose to determine the range of mutagenic outcomes that can arise from meiotic DSBs, including long-range deletions and duplications and chromosomal translocations. In the second aim, we examine factors that may impact the formation of deletions. We focus on the effect of DNA end processing at two steps, SPO11 removal and end processing, and recombination. Further, we address whether gaps formed at nearby DSBs are substrates for homologous recombination and the impact of paternal age in the rearrangement events at meiotic DSBs.