Spatial and temporal control of meiosis

卵子激活是成熟卵子被触发完成减数分裂，同时经历关键的发育变化，为随后的胚胎发育做准备的过程。CORT基因编码减数分裂特异的后期促进复合体激活物(APC/C)，几乎参与卵子激活的各个方面。少数其他基因也与卵子激活有关，包括CanB2磷酸酶和CycB3-CDK1。我们研究的一个主要目标是确定这些基因如何相互作用来促进减数分裂的完成。我们还有兴趣发现这些细胞周期调节器如何控制与卵子激活有关的关键发育过程。目的1.果蝇减数分裂过程中APC/C的调控在目标1中，我们使用质谱仪(MS)来鉴定CORT和Fzy APC/C复合体的亚基，并鉴定随着EGG激活而变化的翻译后修饰。我们将通过比较野生型和突变型Cort和Fzy基因的MS图谱来确定CanB2和CycB3如何调节减数分裂中的APC/C。为了补充这种生化方法，我们使用缺失筛查来探索整个基因组，以寻找增强或抑制CycB3获得或功能丧失的等位基因。同样在目标1中，我们将继续我们令人惊讶的发现，即APC/CFzr的抑制剂Rca1在减数分裂中具有APC/C激活作用。*目标2.确定染色体在减数分裂中如何分离。我们发现，果蝇减数分裂中的染色体分离需要蛋白酶、分离酶及其抑制剂Securin的APC/C介导的降解。这一目标的一个主要目标是识别分离目标。同样在目标2中，我们评估了CORT、CanB2、CycB3和Rca1在手臂凝聚力释放中的作用，并继续我们的发现，CycB-CDK1在减数分裂中参与了染色体分离。我们还确定了在减数分裂中分离酶是否受局部控制，可能是由Securin或CycB控制。*目的3.确定APC/CCort在性别决定中的作用。我们发现了APC/C在性别决定中的一个新角色。我们将测试我们的模型，即APC/CCort针对性别决定的关键调节因子Tra进行破坏。我们认为，CORT在雌性生殖系中起作用，以防止母体Tra将胚胎交给女性命运。我们认为CORT的这一作用在昆虫物种中尤其重要，因为在昆虫物种中，TrA是性别决定的主要调节者。我们将以MedFly为模型直接进行测试。*该研究项目将直接培养3名博士和20名本科生。所有人都将接受广泛的分子、遗传学、生物化学和生物成像技术方面的高水平培训。所有学生都将接受科学方法的培训，他们将发展他们设计和解释实验的能力，他们将学习写作和展示他们的工作。学生将非常适合从事研究、教学和工业方面的工作。

Egg activation is the process by which a mature egg is triggered to complete meiosis and at the same time to undergo critical developmental changes that prepare the egg for subsequent embryonic development. The cort gene, which encodes a meiosis-specific activator of the Anaphase Promoting Complex (APC/C), is involved in almost all aspects of egg activation. A small number of other genes have also been implicated in egg activation, including the CanB2 phosphatase and CycB3-Cdk1. A major goal of our research is to determine how these genes interact to promote the completion of meiosis. We are also interested in discovering how these cell cycle regulators also control critical developmental processes that are linked to egg activation. ******Aim 1. Regulation of the APC/C in Drosophila meiosis. In Aim 1 we use mass-spectrometry (MS) to identify subunits of Cort and Fzy APC/C complexes and identify post-translational modifications that change with egg activation. We will determine how CanB2 and CycB3 regulate the APC/C in meiosis by comparing MS profiles of Cort and Fzy in wild type versus mutants for these genes. To complement this biochemical approach, we employ a deficiency screen to probe the entire genome for genes that enhance or suppress cycB3 gain or loss-of-function alleles. Also in Aim 1 we will follow up on our surprising finding that Rca1, an inhibitor of APC/CFzr, has an APC/C activating role in meiosis. ******Aim 2. Determine how chromosomes segregate in meiosis. We found that chromosome segregation in Drosophila meiosis requires the protease, Separase, and the APC/C-mediated degradation of its inhibitor Securin. A major goal in this Aim is to identify the Separase target. Also in Aim 2 we evaluate the role of Cort, CanB2, CycB3 and Rca1 in the release of arm cohesion, and we follow up on our finding that CycB-Cdk1 plays a role in chromosome segregation in meiosis. We also determine if Separase is controlled locally in meiosis, possibly by Securin or CycB. ******Aim 3. Determine role of APC/CCort in sex determination. We uncovered a novel role for the APC/C in sex determination. We will test our model that APC/CCort targets a key regulator of sex determination, Tra for destruction. We propose that Cort acts in female germline to prevent maternal Tra from committing the embryo to a female fate. We propose that this role for Cort is particularly important in insect species in which Tra is the master regulator of sex determination. We will directly test this using the medfly as a model.******This research program will directly train 3 PhD and 20 undergraduate students. All will receive a high level of training in a broad range of molecular, genetics, biochemistry, and biological imaging techniques. All students will be trained in scientific method, they will develop their ability to design and interpret experiments and they will learn to write and present their work. Students will be highly qualified for jobs in research, teaching and industry.