Abstract

Proper oocyte maturation is a prerequisite for successful reproduction and requires the resumption of meiosis to the metaphase II stage (MII). In bovine oocytes, nuclear maturation has been shown to occur in in vitro maturing cumulus-enclosed oocytes (COCs) in the absence of transcription, but their developmental capacity is reduced compared to transcriptionally competent COCs. To assess the impact of transcription during in vitro maturation of bovine COCs on the quantitative oocyte proteome, a holistic nano-LC–MS/MS analysis of germinal vesicle oocytes and MII oocytes matured with or without addition of the transcription inhibitor actinomycin D (ActD) was carried out. Analyzing eight biological replicates for each of the three groups, a total of 2018 proteins was identified. These could be clearly classified into proteins depending or not depending on transcription during oocyte maturation. Proteins whose abundance increased after maturation irrespective of transcription inhibition - and hence independent of transcription - were related to the cell cycle, reflecting the progression of meiosis, and to cellular component organization, which is crucial for cytoplasmic maturation. In contrast, transcription-dependent proteins were associated with cell–cell adhesion and translation. Since a high rate of protein synthesis in oocytes has been shown to correlate with their developmental competence, oocyte maturation in transcriptionally impaired COCs is apparently disturbed. Our experiments reveal that impaired transcription during in vitro maturation of COCs has a substantial effect on specific components of the oocyte proteome, and that transcription is required for specific classes of oocyte proteins predominantly involved in translation.

中文翻译：

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体外成熟过程中转录抑制对牛卵母细胞蛋白质组的影响。

摘要

适当的卵母细胞成熟是成功繁殖的先决条件，需要将减数分裂恢复到中期 II 期 (MII)。在牛卵母细胞中，核成熟已显示在体​​外成熟的卵丘封闭卵母细胞 (COC) 中在没有转录的情况下发生，但与具有转录能力的 COC 相比，它们的发育能力降低。为了评估牛 COC 体外成熟过程中转录对定量卵母细胞蛋白质组的影响，对添加或不添加转录抑制剂放线菌素 D (ActD) 成熟的胚泡卵母细胞和 MII 卵母细胞进行整体纳米 LC-MS/MS 分析进行了。分析三组中每组的八个生物学重复，共鉴定出 2018 种蛋白质。这些可以根据卵母细胞成熟过程中的转录清楚地分为取决于或不取决于转录的蛋白质。成熟后丰度增加的蛋白质与转录抑制无关 - 因此独立于转录 - 与细胞周期有关，反映减数分裂的进展，以及细胞成分组织，这对细胞质成熟至关重要。相反，转录依赖性蛋白与细胞间粘附和翻译有关。由于卵母细胞中蛋白质合成的高速率已被证明与其发育能力相关，因此转录受损的 COC 中的卵母细胞成熟显然受到干扰。