Institute of Reproductive Medicine, Department of Biomedical Science, College of Life Science, CHA University, Pangyo-Ro 335, Bundang-gu, Seongnam-si, Gyeonggi-do, 13488, Korea.
Sci Rep. 2019 Jul 17;9(1):10343. doi: 10.1038/s41598-019-46459-3.
Previously, we found that the silencing of growth arrest-specific gene 6 (Gas6) expression in oocytes impairs cytoplasmic maturation through mitochondrial overactivation with concurrent failure of pronuclear formation after fertilization. In this study, we report that Gas6 regulates mitophagy and safeguards mitochondrial activity by regulating mitophagy-related genes essential to the complete competency of oocytes. Based on RNA-Seq and RT-PCR analysis, in Gas6-silenced MII oocytes, expressions of mitophagy-related genes were decreased in Gas6-silenced MII oocytes, while mitochondrial proteins and Ptpn11, the downstream target of Gas6, was increased. Interestingly, GAS6 depletion induced remarkable MTOR activation. Gas6-depleted MII oocytes exhibited mitochondrial accumulation and aggregation caused by mitophagy inhibition. Gas6-depleted MII oocytes had a markedly lower mtDNA copy number. Rapamycin treatment rescued mitophagy, blocked the increase in MTOR and phosphorylated-MTOR, and increased the mitophagy-related gene expression in Gas6-depleted MII oocytes. After treatment with Mdivi-1, a mitochondrial division/mitophagy inhibitor, all oocytes matured and these MII oocytes showed mitochondrial accumulation but reduced Gas6 expression and failure of fertilization, showing phenomena very similar to the direct targeting of Gas6 by RNAi. Taken together, we conclude that the Gas6 signaling plays a crucial role in control of oocytes cytoplasmic maturation by modulating the dynamics and activity of oocyte mitochondria.
先前，我们发现卵母细胞中生长停滞特异性基因 6（Gas6）表达的沉默通过线粒体过度激活，同时受精后原核形成失败，从而损害细胞质成熟。在这项研究中，我们报告 Gas6 通过调节卵母细胞完全成熟所必需的与线粒体自噬相关的基因来调节线粒体自噬并保护线粒体活性。基于 RNA-Seq 和 RT-PCR 分析，在 Gas6 沉默的 MII 卵母细胞中，线粒体自噬相关基因的表达在 Gas6 沉默的 MII 卵母细胞中降低，而线粒体蛋白和 Gas6 的下游靶标 Ptpn11 增加。有趣的是，Gas6 耗竭诱导了显著的 MTOR 激活。Gas6 耗尽的 MII 卵母细胞表现出由于线粒体自噬抑制而导致的线粒体积累和聚集。Gas6 耗尽的 MII 卵母细胞的 mtDNA 拷贝数明显较低。雷帕霉素处理挽救了线粒体自噬，阻止了 MTOR 和磷酸化-MTOR 的增加，并增加了 Gas6 耗尽的 MII 卵母细胞中与线粒体自噬相关的基因表达。在用线粒体分裂/线粒体自噬抑制剂 Mdivi-1 处理后，所有卵母细胞成熟，这些 MII 卵母细胞显示线粒体积累，但 Gas6 表达减少和受精失败，表现出与 Gas6 通过 RNAi 直接靶向非常相似的现象。总之，我们得出结论，Gas6 信号通过调节卵母细胞线粒体的动态和活性在控制卵母细胞细胞质成熟中起关键作用。
