未修复的DNA损伤在卵母细胞中的持续存在是有害的,并可能导致遗传畸变,流产,和不孕症。RPA,ssDNA结合复合物,对各种DNA相关过程至关重要。在这里,我们报道了RPA在减数分裂重组后的出生后卵母细胞发育过程中的DNA损伤修复中起着新的作用。探讨RPA在卵子发生过程中的作用,我们灭活RPA1(复制蛋白A1),异源三聚体RPA复合物的最大亚基,特异性地在卵母细胞中使用两种种系特异性Cre驱动剂(Ddx4-Cre和Zp3-Cre)。我们发现RPA1的耗尽导致RPA复合物的分解,RPA1缺陷型卵母细胞中RPA2和RPA3的缺失证明了这一点。引人注目的是,在缺乏RPA1的GV期卵母细胞中发生严重的DNA损伤。卵母细胞中RPA的缺失引发了典型的DNA损伤反应机制和途径,例如激活ATM,ATR,DNA-PK,和p53。此外,RPA缺乏导致卵母细胞中期Ⅰ和中期Ⅱ期染色体错位,这与RPA1缺陷卵母细胞中参与细胞骨架组织的基因转录水平的改变一致。卵母细胞中缺乏RPA复合物会严重损害卵泡发生,并导致卵母细胞数量和女性不育的显着减少。我们的结果表明,RPA在哺乳动物卵泡发育过程中的DNA损伤修复中起着意想不到的作用。
Persistence of unrepaired DNA damage in oocytes is detrimental and may cause genetic aberrations, miscarriage, and infertility. RPA, an ssDNA-binding complex, is essential for various DNA-related processes. Here we report that RPA plays a novel role in DNA damage repair during postnatal oocyte development after meiotic recombination. To investigate the role of RPA during oogenesis, we inactivated RPA1 (replication protein A1), the largest subunit of the heterotrimeric RPA complex, specifically in oocytes using two germline-specific Cre drivers (Ddx4-Cre and Zp3-Cre). We find that depletion of RPA1 leads to the disassembly of the RPA complex, as evidenced by the absence of RPA2 and RPA3 in RPA1-deficient oocytes. Strikingly, severe DNA damage occurs in RPA1-deficient GV-stage oocytes. Loss of RPA in oocytes triggered the canonical DNA damage response mechanisms and pathways, such as activation of ATM, ATR, DNA-PK, and p53. In addition, the RPA deficiency causes chromosome misalignment at metaphase I and metaphase II stages of oocytes, which is consistent with altered transcript levels of genes involved in cytoskeleton organization in RPA1-deficient oocytes. Absence of the RPA complex in oocytes severely impairs folliculogenesis and leads to a significant reduction in oocyte number and female infertility. Our results demonstrate that RPA plays an unexpected role in DNA damage repair during mammalian folliculogenesis.