structures and mechanism of BRCA2 in meiotic recombination

在繁殖生物体时，需要交换和整合两组基因组。以双链断裂形式对我们的DNA进行扰动是亲本染色体交换遗传信息和发生基因重组所必需的。有趣的是，在非有性生殖（有丝分裂）细胞中，同样形式的断裂也是对DNA最严重的损害之一，因为修复失败可能导致细胞死亡，而修复错误可能导致DNA突变，从而导致癌症的发展。更值得注意的是，这两个过程使用相似的过程来修复这些断裂。在有丝分裂细胞中，这个过程被称为同源重组，它利用姐妹染色单体作为修复的模板，而在减数分裂中，这个过程被称为减数分裂重组，它利用同源染色体。在这两种过程中，他们都使用一种重组酶蛋白来包裹单链DNA的长链。这些蛋白质-DNA细丝用于寻找配对染色体上的同源DNA序列。在同源重组中，需要一个称为RAD51的重组酶，而在减数分裂重组中，需要RAD51和另一个与RAD51非常相似的称为DMC1的重组酶。这两个过程也使用一种叫做BRCA2的关键蛋白质。BRCA2是乳腺癌易感蛋白之一，BRCA2基因突变会增加患乳腺癌、卵巢癌等癌症的风险。BRCA2基因突变会降低细胞修复DNA损伤的能力，从而增加患癌症的风险，这一点目前还不完全清楚。鉴于BRCA2在减数分裂重组中的重要性，在小鼠中，BRCA2缺陷导致雄性不育也就不足为奇了。然而，尚不清楚这是否也适用于人类。因此，了解BRCA2在减数分裂重组中的确切作用以及BRCA2如何发挥其作用是有趣和重要的。此外，为什么在这个过程中需要RAD51和DMC1，以及RAD51在这个过程中的确切作用是什么，这是令人着迷的。在本提案中，我们计划研究这些重要问题。我们计划使用尖端的电子显微镜单粒子技术来研究BRCA2和DMC1的三维结构，并可能同时研究BRCA2， RAD51和DMC1。此外，我们将使用一些生化和生物物理技术来可视化和量化BRCA2影响下DMC1细丝的长度和数量。我们的研究将提供详细的机制理解BRCA2在减数分裂重组中是什么以及如何发挥这些重要作用，减数分裂重组是产生遗传多样性和繁殖的基本过程。

In reproducing organisms, two sets of genomes need to be exchanged and integrated. The perturbations to our DNA in the form of double stranded breaks are required for parental chromosomes to exchange genetic information and for genetic recombination to occur. Interestingly, the same form of break is also one of most severe types of damage to our DNA in non-sexually reproducing (mitotic) cells as failure to repair it can cause cell death while mis-repair could cause mutations in DNA that can lead to the development of cancer. Even more remarkable is that the two processes use similar processes to repair these breaks. In mitotic cells, this process is called homologous recombination where it utilizes the sister chromatid as templates for repair while in meiosis, the process is called meiotic recombination, which utilizes the homologous chromosome. In both processes, they use a recombinase protein that coats long stretches of single stranded DNA. These protein-DNA filaments are used to search for homologous DNA sequence in the partner chromosome. In homologous recombination, a single recombinase called RAD51 is required while in meiotic recombination, both RAD51 and another recombinase called DMC1, which is very similar to RAD51, are required. Both processes also use a key protein called BRCA2. BRCA2 is one of the breast cancer susceptibility proteins, and mutations in BRCA2 gene increase the risk of breast, ovarian and other cancers. It is partially understood why mutations in BRCA2 can increase the risk of cancer as it reduces the ability of cells to repair a DNA damage. Given the importance of BRCA2 in meiotic recombination, it is not surprising that, in mice, defects in BRCA2 cause male infertility. However, it is unclear if this is also true in human. It is thus intriguing and important to understand what the exact roles BRCA2 plays in meiotic recombination and how BRCA2 fulfills its roles. Furthermore, it is fascinating as why both RAD51 and DMC1 are required in this process and what the exact role of RAD51 is in this process. In this proposal, We plan to investigate these important questions. We plan to use the cutting edge electron microscopy single particle technique to study the 3-dimensional structures of BRCA2 and DMC1 and possibly BRCA2, RAD51 and DMC1 together. Furthermore, we will use a number of biochemical and biophysical techniques to visualize and quantify the length and number of DMC1 filaments under the influence of BRCA2. Our studies will provide a detailed mechanistic understanding of what and how BRCA2 plays these important roles in meiotic recombination, the essential process in generating genetic diversity and reproduction.