Investigating the Role of CDK-2 in Meiotic Recombination

有性繁殖生物在减数分裂过程中依靠适当的染色体分离来确保 产生具有完整遗传互补的配子。在减数分裂前期I，染色体对 并经历染色体交叉，即两个同源基因之间的遗传信息交换 染色体。这一过程导致在同系物之间形成物理连接，并使每个 在减数分裂过程中分离的染色体对I.交叉形成中的缺陷可能是灾难性的，导致 非整倍体和诸如年龄相关性不孕、流产和唐氏综合症等疾病。尽管 尽管这一过程很重要，但管理交叉形成的机制仍然知之甚少。目标是 这个项目的目的是确定如何在减数分裂过程中指定交叉，以实现对 遗传信息。最近在线虫中的一项遗传筛查发现了一种类似周期蛋白的COSA-1，它是 对于处理减数分裂DNA双链断裂成交叉是必不可少的。这一发现随后被 然后鉴定了其哺乳动物直系同源基因CNTD1，该基因在交换中具有保守的作用 队形。然而，目前还没有发现CDK是COSA-1/CNTD1的结合伙伴，以及这些细胞周期蛋白是如何与细胞周期蛋白结合的。 就像指定交叉的蛋白质一样，目前还不清楚。最近，我发现线虫的同源物 CDK2(CDK-2)定位于交叉位点，增加了CDK2与COSA-2合作的可能性。 1/CNTD1，并作为一种活性激酶促进交叉形成。支持这一想法的哺乳动物 长期以来，CDK2在交叉点和端粒都被观察到。然而，由于其端粒功能，CDK2 基因敲除小鼠表现出严重的同源配对和突触缺陷，这是减数分裂的先决条件。 重组。因此，CDK-2在交叉部位的作用仍未得到检验。在这里，我建议使用C。 以elegans为模型系统研究CDK2在减数分裂重组中的保守功能。不像 哺乳动物的CDK2、CDK-2对于同源配对和突触是不必要的，但对于同源配对和突触是特殊需要的 交叉编队。在目标1中，我将使用生长素诱导的降解系统来确定 重组机制上CDK-2耗竭的超分辨显微镜研究。我还会决定是否 CDK-2和COSA-1利用纯化的组分形成活性激酶复合体。在目标2中，我将建立 CDK2通过两种候选的减数分裂底物识别其减数分裂底物来指定交叉的机制- 基于和不偏不倚的化学遗传方法。然后我将确定CDK2的功能意义 通过在线虫中进行定向突变的靶标。总体而言，这项工作的结果将阐明保守的 指定交叉的调控机制，将广泛适用于高等真核生物。约书亚 霍普金斯大学提供最先进的研究环境，研究生成功的良好记录， 庞大的地区性科学支持网络和广泛的职业发展计划，使其独一无二 适合我提出的研究和职业目标的成功。

Sexually reproducing organisms rely on proper chromosome segregation during meiosis to ensure the production of gametes with the complete genetic complement. During meiotic prophase I, chromosomes pair and undergo chromosomal crossover, an exchange of genetic information between two homologous chromosomes. This process leads to the formation of physical linkages between the homologs and enables each chromosome pair to separate during meiosis I. Defects in crossover formation can be disastrous, leading to aneuploidy and conditions such as age-related infertility, miscarriages, and Down Syndrome. Despite the importance of this process, the mechanisms governing crossover formation remain poorly understood. The goal of this project is to determine how crossovers are designated during meiosis to achieve faithful transmission of genetic information. A recent genetic screen in C. elegans has identified a cyclin-like protein COSA-1 that is essential for processing meiotic DNA double-strand breaks into crossovers. This finding was subsequently followed by the identification of its mammalian ortholog CNTD1, which has conserved roles in crossover formation. However, no CDK has been identified as a binding partner of COSA-1/CNTD1, and how these cyclin- like proteins designate crossovers is not known. Recently, I have discovered that the C. elegans homolog of CDK2 (CDK-2) localizes to the sites of crossovers, raising the possibility that CDK2 might partner with COSA- 1/CNTD1 and function as an active kinase to promote crossover formation. Supporting this idea, mammalian CDK2 has long been observed at crossovers as well as telomeres. However, due to its telomeric function, CDK2 knockout mice exhibit severe defects in homolog pairing and synapsis, which are prerequisite to meiotic recombination. Therefore, the role of CDK-2 at crossover sites has remained untested. Here I propose to use C. elegans as a model system to investigate the conserved function of CDK2 in meiotic recombination. Unlike the mammalian CDK2, CDK-2 is dispensable for homolog pairing and synapsis, but is specifically required for crossover formation. In Aim 1 I will employ the auxin-inducible degradation system to determine the effect of CDK-2 depletion on recombination machineries by super-resolution microscopy. I will also determine whether CDK-2 and COSA-1 form an active kinase complex using purified components. In Aim 2 I will establish the mechanisms by which CDK2 designates crossovers by identifying its meiotic substrates through both candidate- based and unbiased chemical genetic approaches. I will then determine the functional significance of CDK2 targets through targeted mutagenesis in C. elegans. Overall, the results of this work will elucidate the conserved regulatory mechanisms that designate crossovers and will be broadly applicable to higher eukaryotes. Johns Hopkins University offers a state of-the-art research environment, strong records of post-graduate success, a vast regional scientific support network, and a wide array of career development programs that make it uniquely suited for success of my proposed research and career goals.