卵母細胞における染色体ダイナミクス制御と母体の加齢効果

卵母细胞染色体动态的控制和母亲衰老的影响

• 批准号: 16F16088
• 负责人: Carlton Peter
• 金额: $ 1.47万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for JSPS Fellows
• 财政年份: 2016
• 资助国家: 日本
• 起止时间: 2016-04-22 至 2019-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16F16088/
• 关键词: PPH-4.1 suppressor; fecundity; EMS mutagenesis; PPH-4.1 suppressors; PPH-4; immunoprecipitation (IP)

The highly conserved Serine/Threonine protein phosphatase PP4 homolog, PPH-4.1 in C. elegans, is required to regulate meiosis. The pph-4.1 null mutants Δ pph-4.1 show very low fecundity. The goal of our project is to identify the interactors of PPH-4.1, furthermore, to understand the molecular mechanisms of how PPH-4.1 regulate the fecundity of worms. We performed genetic screen to look for PPH-4.1 interactors, potentially its suppressors. Overall, 10470 genomes were screened and 73 potential pph-4.1 suppressor strains were isolated. After whole genome sequencing analysis, we found there is a mutation causing dct-17 coding sequence changed. To dampen the function of DCT-17, we also used RNA interference (RNAi) method to resemble EMS introduced dct-17 mutation. Our RNAi experiement showed consistent results, reduction of dct-17 mRNA by RNAi bacteria feeding to Δ pph-4.1 mutants dramatically improve the fecundity of the mutants. This result indicate dct-17 might be pph-4.1’s suppressor in regulating meiosis. DCT-17 is a predicted pyrophosphatase. Pyrophosphatase is an enzyme that catalyzes the conversion of one molecule of pyrophosphate to two phosphate ions. In Δ pph-4.1 mutants, PPH-4.1’s substrates are hyperphosphorylated, resulting in very low fecundity. Our result suggested dampening the function of dct-17 resulted in the reduction of the total phosphate, thereby, change the requirement of phosphorylation events during meiosis, leading to improvement of the fecundity in pph-4.1 null mutants. Our result might also shed light on how to improve human’s fecundity.

线虫中高度保守的丝氨酸/苏氨酸蛋白磷酸酶PP4同源物PPH-4.1是调节减数分裂所必需的。Pph-4.1缺失突变体ΔPph-4.1表现出极低的繁殖力。本项目的目标是确定PPH-4.1的相互作用因子，进而了解PPH-4.1调节虫子繁殖力的分子机制。我们进行了基因筛查，寻找PPH-4.1的相互作用因子，可能是它的抑制因子。总共筛选了10470个基因组，分离出73株潜在的PPH4.1抑制株。经全基因组测序分析，发现有突变导致DCT-17编码序列发生改变。为了抑制DCT-17的功能，我们还使用了RNA干扰(RNAi)的方法来模拟EMS引入的DCT-17突变。我们的RNAi实验结果一致，RNAi细菌对ΔPph-4.1突变体的RNAi降低了DCT-17mRNA的表达，显著提高了突变体的繁殖力。这一结果表明DCT-17可能是PPh-4.1‘S调控减数分裂的抑制剂。DCT-17是一种预测的焦磷酸酶。焦磷酸酶是一种催化一个焦磷酸盐分子转化为两个磷酸根离子的酶。在ΔPph-4.1突变体中，Pph-4.1‘S底物过度磷酸化，导致繁殖力非常低。我们的结果表明，抑制DCT-17的功能导致总磷的减少，从而改变了减数分裂过程中对磷酸化事件的需求，从而提高了PPH-4.1缺失突变体的繁殖力。我们的结果也可能为如何提高人类的繁殖力提供帮助。