Division of Reproduction and Genetics, The First Affiliated Hospital of University of Science and Technology of China, School of Basic Medical Sciences, Division of Life Sciences and Medicine, Biomedical Sciences and Health Laboratory of Anhui Province, University of Science and Technology of China, Hefei, 230027, China.
Department of Biotechnology, COMSATS University Islamabad, Abbottabad Campus, Abbottabad, 22060, Pakistan.
J Assist Reprod Genet. 2024 Jan;41(1):109-120. doi: 10.1007/s10815-023-02966-x. Epub 2023 Oct 13.
Asthenozoospermia is an important cause of male infertility, and the most serious type is characterized by multiple morphological abnormalities of the sperm flagella (MMAF). However, the precise etiology of MMAF remains unknown. In the current study, we recruited a consanguineous Pakistani family with two infertile brothers suffering from primary infertility due to MMAF without obvious signs of PCD.
We performed whole-exome sequencing on DNAs of the patients, their parents, and a fertile brother and identified the homozygous missense variant (c.1490C > G (p.P497R) in NPHP4 as the candidate mutation for male infertility in this family.
Sanger sequencing confirmed that this mutation recessively co-segregated with the MMAF in this family. In silico analysis revealed that the mutation site is conserved across different species, and the identified mutation also causes abnormalities in the structure and hydrophobic interactions of the NPHP4 protein. Different bioinformatics tools predict that NPHP4 mutation is pathogenic. Furthermore, Papanicolaou staining and scanning electron microscopy of sperm revealed that affected individuals displayed typical MMAF phenotype with a high percentage of coiled, bent, short, absent, and/or irregular flagella. Transmission electron microscopy images of the patient's spermatozoa revealed significant anomalies in the sperm flagella with the absence of a central pair of microtubules (9 + 0) in every section scored.
Taken together, these results show that the homozygous missense mutation in NPHP4 is associated with MMAF.
弱精症是男性不育的一个重要原因，其中最严重的类型表现为精子鞭毛多形态异常（MMAF）。然而，MMAF 的确切病因仍不清楚。在本研究中，我们招募了一个巴基斯坦的近亲家庭，其中两个不育的兄弟因 MMAF 而患有原发性不育，没有明显的 PCD 迹象。
我们对患者、其父母和一个生育能力正常的兄弟的 DNA 进行了全外显子组测序，鉴定出 NPHP4 中的纯合错义突变（c.1490C>G(p.P497R）为该家系男性不育的候选突变。
Sanger 测序证实该突变在这个家庭中与 MMAF 呈隐性共分离。序列比对分析表明突变位点在不同物种中保守，鉴定出的突变也导致了 NPHP4 蛋白结构和疏水性相互作用的异常。不同的生物信息学工具预测 NPHP4 突变是致病性的。此外，精子巴氏染色和扫描电镜显示，受影响的个体表现出典型的 MMAF 表型，卷曲、弯曲、短、缺失和/或不规则的鞭毛比例很高。患者精子的透射电镜图像显示，精子鞭毛的中央微管（9+0）每一节都明显缺失。
综上所述，这些结果表明 NPHP4 中的纯合错义突变与 MMAF 相关。
