转录因子AP-2-α(Tfap2a)是一种重要的序列特异性DNA结合蛋白,可以通过与诱导型病毒和细胞增强子元件合作调节多个基因的转录。在这个实验中,表达式,本地化,并在小鼠卵母细胞成熟过程中研究了Tfap2a的功能。通过显微注射Myc-Tfap2amRNA到卵质中的过表达,免疫荧光,用免疫印迹法研究Tfap2a在小鼠卵母细胞减数分裂中的作用。根据我们的结果,Tfap2a在小鼠卵母细胞成熟中起着至关重要的作用。患有2型糖尿病的小鼠的GV卵母细胞中的Tfap2a水平显著增加。Tfap2a分布在卵质和核质中,随着减数分裂恢复的进展,其水平逐渐增加。Tfap2a的过表达使染色质松弛,加速生发囊泡分解(GVBD),并在体外成熟后14小时阻断了第一次极体挤出。中期板的宽度在中期I阶段增加,过表达的Tfap2a在卵母细胞中破坏了中期II期的纺锤体和染色体组织。此外,Tfap2a过表达显著提高了小鼠GV卵母细胞中p300的表达。此外,泛组蛋白赖氨酸乙酰化水平(PanKac),组蛋白H4赖氨酸12乙酰化(H4K12ac),和H4赖氨酸16乙酰化(H4K16ac),以及泛组蛋白赖氨酸乳酸化(PanKla),组蛋白H3lysine18乳酸化(H3K18la),和H4lysine12乳酸化(H4K12la),Tfap2a过表达后GV卵母细胞均增加。总的来说,Tfap2a过表达上调p300,增加组蛋白乙酰化和乳酸化水平,阻碍纺锤体组装和染色体排列,并最终阻碍小鼠卵母细胞减数分裂。
Transcription factor AP-2-alpha (Tfap2a) is an important sequence-specific DNA-binding protein that can regulate the transcription of multiple genes by collaborating with inducible viral and cellular enhancer elements. In this experiment, the expression, localization, and functions of Tfap2a were investigated in mouse oocytes during maturation. Overexpression via microinjection of Myc-Tfap2a mRNA into the ooplasm, immunofluorescence, and immunoblotting were used to study the role of Tfap2a in mouse oocyte meiosis. According to our results, Tfap2a plays a vital role in mouse oocyte maturation. Levels of Tfap2a in GV oocytes of mice suffering from type 2 diabetes increased considerably. Tfap2a was distributed in both the ooplasm and nucleoplasm, and its level gradually increased as meiosis resumption progressed. The overexpression of Tfap2a loosened the chromatin, accelerated germinal vesicle breakdown (GVBD), and blocked the first polar body extrusion 14 h after maturation in vitro. The width of the metaphase plate at metaphase I stage increased, and the spindle and chromosome organization at metaphase II stage were disrupted in the oocytes by overexpressed Tfap2a. Furthermore, Tfap2a overexpression dramatically boosted the expression of p300 in mouse GV oocytes. Additionally, the levels of pan histone lysine acetylation (Pan Kac), histone H4 lysine 12 acetylation (H4K12ac), and H4 lysine 16 acetylation (H4K16ac), as well as pan histone lysine lactylation (Pan Kla), histone H3 lysine18 lactylation (H3K18la), and H4 lysine12 lactylation (H4K12la), were all increased in GV oocytes after Tfap2a overexpression. Collectively, Tfap2a overexpression upregulated p300, increased the levels of histone acetylation and lactylation, impeded spindle assembly and chromosome alignment, and ultimately hindered mouse oocyte meiosis.