背景:早期胚胎停滞和破碎(EEAF)是女性不孕的常见原因,但是遗传原因仍然很大程度上是未知的。CIP2A编码PP2A的细胞抑制剂,在有丝分裂和小鼠卵母细胞减数分裂中起着至关重要的作用。
方法:进行外显子组测序和Sanger测序以鉴定EEAF患者的候选致病基因。通过Westernblotting在培养的细胞系和人卵母细胞中评估并确认了CIP2A变体的致病性,半定量RT-PCR,TUNEL染色,和荧光定位分析。
结果:我们确定了CIP2A(c.1510C>T,p.L504F)是来自近亲家族的人类EEAF中的新型候选致病基因。L504在进化过程中是高度保守的。TheCIP2A变体(c.1510C>T,p.L504F)降低了mutantCIP2A蛋白的表达水平,导致突变CIP2A蛋白的异常聚集和细胞凋亡。患者的卵母细胞和早期胚胎中发生CIP2A蛋白的异常聚集和染色体分散。我们通过在人卵母细胞中敲低CIP2A进一步复制了患者表型。此外,CIP2A缺乏导致磷酸化ERK1/2水平降低。
结论:我们首先发现CIP2A功能缺失变异与EEAF表征的女性不孕症相关。我们的发现表明CIP2A基因在人类卵母细胞和早期胚胎发育中的独特性和重要性。
背景:这项工作得到了国家重点研究发展计划(2023YFC2706302)的支持,国家自然科学基金(81000079,81170165,81870959),HUST学术前沿青年队(2016QYTD02),和华中科技大学的重点研究,同济医院(2022A20)。
BACKGROUND: Early embryonic arrest and fragmentation (EEAF) is a common cause of female infertility, but the genetic causes remain to be largely unknown.
CIP2A encodes the cellular inhibitor of PP2A, playing a crucial role in mitosis and mouse oocyte meiosis.
METHODS: Exome sequencing and Sanger sequencing were performed to identify candidate causative genes in patients with EEAF. The pathogenicity of the CIP2A variant was assessed and confirmed in cultured cell lines and human oocytes through Western blotting, semi-quantitative RT-PCR, TUNEL staining, and fluorescence localization analysis.
RESULTS: We identified CIP2A (c.1510C > T, p.L504F) as a novel disease-causing gene in human EEAF from a consanguineous family. L504 is highly conserved throughout evolution. The
CIP2A variant (c.1510C > T, p.L504F) reduced the expression level of the mutant
CIP2A protein, leading to the abnormal aggregation of mutant
CIP2A protein and cell apoptosis. Abnormal aggregation of CIP2A protein and chromosomal dispersion occurred in the patient\'s oocytes and early embryos. We further replicated the patient phenotype by knockdown
CIP2A in human oocytes. Additionally,
CIP2A deficiency resulted in decreased levels of phosphorylated ERK1/2.
CONCLUSIONS: We first found that the CIP2A loss-of-function variant associate with female infertility characterized by EEAF. Our findings suggest the uniqueness and importance of CIP2A gene in human oocyte and early embryo development.
BACKGROUND: This work was supported by National Key Research and Development Program of China (2023YFC2706302), the National Natural Science Foundation of China (81000079, 81170165, and 81870959), the HUST Academic Frontier Youth Team (2016QYTD02), and the Key Research of Huazhong University of Science and Technology, Tongji Hospital (2022A20).