Deleterious variants in RNF111 impair female fertility and induce premature ovarian insufficiency in humans and mice

Premature ovarian insufficiency (POI) is a heterogeneous female disorder characterized by the loss of ovarian function before the age of 40. It represents a significant detriment to female fertility. However, the known POI-causative genes currently account for only a fraction of cases. To elucidate the genetic factors underlying POI, we conducted whole-exome sequencing on a family with three fertile POI patients and identified a deleterious missense variant in RNF111. In a subsequent replication study involving 1030 POI patients, this variant was not only confirmed but also accompanied by the discovery of three additional predicted deleterious RNF111 variants. These variants collectively account for eight cases, representing 0.78% of the study cohort. A further study involving 500 patients with diminished ovarian reserves also identified two additional RNF111 variants. Notably, RNF111 encodes an E3-ubiquitin ligase with a regulatory role in the TGF-/BMP signaling pathway. Our analysis revealed that RNF111/RNF111 is predominantly expressed in the oocytes of mice, monkeys, and humans. To further investigate the functional implications of RNF111 variants, we generated two mouse models: one with a heterozygous missense mutation (Rnf111+/M) and another with a heterozygous null mutation (Rnf111+/). Both mouse models exhibited impaired female fertility, characterized by reduced litter sizes and small ovarian reserve. Additionally, RNA-seq and quantitative proteomics analysis unveiled that Rnf111 haploinsufficiency led to dysregulation in female gonad development and negative regulation of the BMP signaling pathway within mouse ovaries. In conclusion, our findings strongly suggest that monoallelic deleterious variants in RNF111 can impair female fertility and induce POI in both humans and mice. To investigate the functional implication of monoallelic deleterious variants in RNF111 to premature ovarian insufficiency, we generated two mouse models using CRISPR-Cas9. To further investigate the functional alterations within the mutant mouse ovaries, We then performed bulk RNA-seq analyses on ovaries from wild-type and Rnf111+/- mice.早发性卵巢功能不全（Premature ovarian insufficiency, POI）是一种异质性女性疾病，以40岁前出现卵巢功能丧失为特征，对女性生育力造成严重损害。目前已知的POI致病基因仅能解释少数病例。为阐明POI背后的遗传因素，我们对一个包含3名POI患者的家系开展全外显子组测序，在RNF111基因中鉴定出1个有害错义变异。在后续纳入1030名POI患者的验证队列研究中，该变异不仅得到证实，还额外发现3个预测为有害的RNF111变异。上述变异共计覆盖8例病例，占研究队列的0.78%。在另一项纳入500名卵巢储备功能下降患者的研究中，同样鉴定出2个额外的RNF111变异。值得注意的是，RNF111编码一种E3泛素连接酶，在转化生长因子-β/骨形态发生蛋白（TGF-β/BMP）信号通路中发挥调控作用。我们的分析显示，RNF111在小鼠、猴子和人类的卵母细胞中均呈高表达。为进一步探究RNF111变异的功能意义，我们构建了两种小鼠模型：一种携带杂合错义突变（Rnf111+/M），另一种携带杂合无效突变（Rnf111+/）。两种模型均表现出雌性生育能力受损，具体表现为产仔数减少、卵巢储备降低。此外，批量RNA测序与定量蛋白质组学分析揭示，Rnf111单倍剂量不足会导致小鼠卵巢内雌性性腺发育相关通路失调，并负向调控BMP信号通路。综上，我们的研究结果强烈表明，RNF111基因的单等位有害变异可损伤雌性生育能力，并在人类和小鼠中诱发POI。为探究RNF111单等位有害变异与早发性卵巢功能不全的功能关联，我们利用CRISPR-Cas9技术构建了两种小鼠模型。为深入分析突变小鼠卵巢内的分子改变，我们对野生型与Rnf111+/-小鼠的卵巢开展了批量RNA测序分析。