Reproductive Medicine Center, Department of Obstetrics and Gynecology, The First Affiliated Hospital of Anhui Medical University, Hefei, China.
Anhui Province Key Laboratory of Reproductive Health and Genetics, Anhui Medical University, Hefei, China.
J Med Genet. 2019 Feb;56(2):96-103. doi: 10.1136/jmedgenet-2018-105486. Epub 2018 Nov 10.
Male infertility is a major issue of human reproduction health. Asthenoteratospermia can impair sperm motility and cause male infertility. Asthenoteratospermia with multiple morphological abnormalities of the flagella (MMAF) presents abnormal spermatozoa with absent, bent, coiled, short and/or irregular-calibre flagella. Previous studies on MMAF reported that genetic defects in cilia-related genes (eg, , , , and ) are the major cause of MMAF. However, the known MMAF-associated genes are only responsible for approximately 30% to 50% of human cases. We further investigated the cases with MMAF in search of additional genes mutated in this condition.
We conducted whole exome sequencing in a male individual with MMAF from a consanguineous Han Chinese family. Sanger sequencing was also conducted in additional individuals with MMAF. Intriguingly, a homozygous frameshift mutation (p.Leu357Hisfs11) was identified in the gene encoding CFAP69 (cilia and flagella-associated protein 69), which is highly expressed in testis. The subsequent Sanger sequencing of the coding regions among 34 additional individuals with MMAF revealed a case with homozygous nonsense mutation (p.Trp216) of . Both of these loss-of-function mutations were not present in the human population genome data archived in the 1000 Genomes Project and ExAC databases, nor in 875 individuals of two Han Chinese control populations. Furthermore, we generated the knockout model in mouse orthologue using the CRISPR-Cas9 technology. Remarkably, male -knockout mice manifested with MMAF phenotypes.
Our experimental findings elucidate that homozygous loss-of-function mutations in can lead to asthenoteratospermia with MMAF in humans and mice.
男性不育是人类生殖健康的主要问题。弱精子症可损害精子运动能力，导致男性不育。具有鞭毛多种形态异常的弱精子症（MMAF）表现为精子鞭毛缺失、弯曲、卷曲、短小和/或不规则的精子。先前关于 MMAF 的研究报告称，纤毛相关基因（例如， ， ， ， 和 ）中的遗传缺陷是 MMAF 的主要原因。然而，已知的与 MMAF 相关的基因仅占人类病例的 30%至 50%左右。我们进一步研究了具有 MMAF 的病例，以寻找该病症中突变的其他基因。
我们对一个来自近亲汉族家庭的具有 MMAF 的男性个体进行了全外显子组测序。还对其他具有 MMAF 的个体进行了 Sanger 测序。有趣的是，在编码纤毛和鞭毛相关蛋白 69（CFAP69）的基因中发现了一个纯合移码突变（p.Leu357Hisfs11），该基因在睾丸中高度表达。随后在 34 名具有 MMAF 的个体的 编码区域进行 Sanger 测序，发现一个病例存在纯合无义突变（p.Trp216）。这两种 功能丧失突变均不存在于人类基因组数据库 1000 Genomes Project 和 ExAC 中存档的人群基因组数据中，也不存在于两个汉族对照人群中的 875 个人中。此外，我们使用 CRISPR-Cas9 技术在小鼠同源物 中生成了敲除模型。值得注意的是， 敲除雄性小鼠表现出 MMAF 表型。
我们的实验结果表明， 中的纯合功能丧失突变可导致人类和小鼠的弱精子症伴 MMAF。
