Molecular mechanisms of chromosome organization and recombination control by the meiotic chromosome axis

项目摘要 真核生物的有性生殖涉及减数分裂中产生单倍体配子（在人类中，精子和卵细胞）， 随后两个配子融合产生二倍体后代。在减数分裂中，同源染色体识别一个 另一个通过修饰的同源重组DNA修复途径物理连接， 交换使得在减数分裂I分裂中能够精确地分离同源物以降低倍性。在大多数真核生物中， 在人类中，染色体由一种高度保守的结构（称为染色体）组成， 轴线染色体轴还募集和控制DNA切割和重组因子，以介导 交叉，并在交叉形成后在控制重组水平的关键反馈途径中被重塑。 在这里，我们建议将联合收割机生物化学、大分子结构和遗传学结合起来研究这两种S。酿酒酵母和小鼠， 确定染色体轴如何组装，组织染色体，并介导交叉形成。我们将首先 确定了S.酿酒酵母Red 1和哺乳动物SYCP 2：SYCP 3，功能相关的染色体轴 我们已经发现的“基础”蛋白共享保守的结构域结构和自组装成细丝的倾向。 接下来，我们将确定这些蛋白质如何与减数分裂粘附素复合物相互作用，以了解其结构基础。 轴介导的染色体组构。接下来，我们将剖析由S介导的相互作用网络。酿酒酵母Hop 1，a 保守的HORMAD家族成员轴蛋白和减数分裂重组的主调节器，并研究如何 这种相互作用网络随着减数分裂前期的进展而变化。Hop 1最终从染色体轴上移除， 控制重组水平的重要反馈途径，由AAA+ ATP酶Pch 2介导。我们将测试我们的 假设Pch 2直接识别特定的Hop 1构象并部分展开其HORMA结构域以介导 将其从轴上移除。最后，我们将研究结构，DNA结合特异性，和两个减数分裂的相互作用- 特定的蛋白质复合物，Msh 4：Msh 5和Zip 2：Zip 4：Spo 16，以了解它们如何稳定特定的DNA重组 中间体和协调的交叉形成与染色体轴形态的变化。 总的来说，这里提出的工作将导致染色体轴如何组装的全面分子图像， 协调交叉形成，然后随着重组的进行而分解。了解分子 染色体轴和相关因素的机制与人类健康高度相关，如减数分裂错误， 染色体分离是人类流产的主要原因，并且是“非整倍性疾病”的来源， 唐氏综合症和特纳综合症。此外，许多癌症类型显示减数分裂染色体轴的错误表达 蛋白质，包括TRIP 13，HORMAD 1和SYCP 2。更好地了解这些蛋白质在其天然 环境对于确定它们的错误表达如何导致基因组不稳定和癌症至关重要。

PROJECT SUMMARY Sexual reproduction in eukaryotes involves the generation of haploid gametes (in humans, sperm and egg cells) in meiosis, followed by the fusion of two gametes to produce diploid offspring. In meiosis, homologous chromosomes recognize one another and become physically linked through a modified homologous recombination DNA repair pathway, and the resulting crossovers enable accurate homolog segregation in the meiosis I division to reduce ploidy. In most eukaryotes including humans, chromosomes are organized as an array of chromatin loops by a highly conserved structure called the chromosome axis. The chromosome axis also recruits and controls DNA cleavage and recombination factors to mediate the formation of crossovers, and is remodeled after crossover formation in a key feedback pathway controlling recombination levels. Here, we propose to combine biochemistry, macromolecular structure, and genetics in both S. cerevisiae and the mouse to determine how the chromosome axis assembles, organizes chromosomes, and mediates crossover formation. We will first determine the structures of S. cerevisiae Red1 and mammalian SYCP2:SYCP3, functionally-related chromosome axis “foundation” proteins that we have found share a conserved domain structure and propensity to self-assemble into filaments. We will next determine how these proteins interact with meiotic cohesin complexes, to understand the structural basis for axis-mediated chromosome organization. Next, we will dissect the network of interactions mediated by S. cerevisiae Hop1, a member of the conserved HORMAD family of axis proteins and a master regulator of meiotic recombination, and study how this interaction network changes during as meiotic prophase progresses. Hop1's eventual removal from the chromosome axis, an important feedback pathway controlling recombination levels, is mediated by the AAA+ ATPase Pch2. We will test our hypothesis that Pch2 directly recognizes a specific Hop1 conformation and partially unfolds its HORMA domain to mediate its removal from the axis. Finally, we will examine the structures, DNA binding specificity, and interactions of two meiosis- specific protein complexes, Msh4:Msh5 and Zip2:Zip4:Spo16, to learn how they stabilize specific DNA recombination intermediates and coordinate crossover formation with chromosome axis morphology changes. Overall, the work proposed here will result in a comprehensive molecular picture of how the chromosome axis assembles, coordinates crossover formation, and is then disassembled as recombination proceeds. Understanding the molecular mechanisms of the chromosome axis and associated factors is highly relevant to human health, as errors in meiotic chromosome segregation are a principal cause of miscarriage in humans, and are the source of “aneuploidy disorders” like Down syndrome and Turner syndrome. Moreover, many cancer types show mis-expression of meiotic chromosome axis proteins, including TRIP13, HORMAD1, and SYCP2. A better understanding of these proteins' mechanisms in their native environment will be critical to determine how their mis-expression might lead to genome instability and cancer.