Noda Taichi, Shinohara Hina, Kobayashi Sumire, Taira Ayumu, Oura Seiya, Tahara Duri, Tokuyasu Midori, Araki Kimi, Ikawa Masahito
Division of Reproductive Biology, Institute of Resource Development and Analysis, Kumamoto University, Kumamoto, Kumamoto, Japan.
Priority Organization for Innovation and Excellence, Kumamoto University, Kumamoto, Kumamoto, Japan.
Biol Reprod. 2024 Apr 11;110(4):750-760. doi: 10.1093/biolre/ioae008.
Sperm proteins undergo post-translational modifications during sperm transit through the epididymis to acquire fertilizing ability. We previously reported that the genomic region coding Pate family genes is key to the proteolytic processing of the sperm membrane protein ADAM3 and male fertility. This region contains nine Pate family genes (Pate5-13), and two protein-coding genes (Gm27235 and Gm5916), with a domain structure similar to Pate family genes. Therefore, in this study, we aimed to identify key factors by narrowing the genomic region. We generated three knockout (KO) mouse lines using CRISPR/Cas9: single KO mice of Pate10 expressed in the caput epididymis; deletion KO mice of six caput epididymis-enriched genes (Pate5-7, 13, Gm27235, and Gm5916) (Pate7-Gm5916 KO); and deletion KO mice of four genes expressed in the placenta and epididymis (Pate8, 9, 11, and 12) (Pate8-12 KO). We observed that the fertility of only Pate7-Gm5916 KO males was reduced, whereas the rest remained unaffected. Furthermore, when the caput epididymis-enriched genes, Pate8 and Pate10 remained in Pate7-Gm5916 KO mice were independently deleted, both KO males displayed more severe subfertility due to a decrease in mature ADAM3 and a defect in sperm migration to the oviduct. Thus, our data showed that multiple caput epididymis-enriched genes within the region coding Pate5-13 cooperatively function to ensure male fertility in mice.
精子蛋白在通过附睾的过程中会经历翻译后修饰，以获得受精能力。我们之前报道过，编码Pate家族基因的基因组区域是精子膜蛋白ADAM3蛋白水解加工和雄性生育能力的关键。该区域包含9个Pate家族基因（Pate5 - 13）和2个蛋白质编码基因（Gm27235和Gm5916），其结构域结构与Pate家族基因相似。因此，在本研究中，我们旨在通过缩小基因组区域来确定关键因素。我们使用CRISPR/Cas9技术构建了三种基因敲除（KO）小鼠品系：在附睾头表达的Pate10单基因敲除小鼠；6个附睾头富集基因（Pate5 - 7、13、Gm27235和Gm5916）的缺失敲除小鼠（Pate7 - Gm5916 KO）；以及在胎盘和附睾中表达的4个基因（Pate8、9、11和12）的缺失敲除小鼠（Pate8 - 12 KO）。我们观察到，只有Pate7 - Gm5916 KO雄性小鼠的生育能力降低，而其余小鼠未受影响。此外，当Pate7 - Gm5916 KO小鼠中剩余的附睾头富集基因Pate8和Pate10被独立敲除时，由于成熟ADAM3减少和精子向输卵管迁移缺陷，两种基因敲除雄性小鼠均表现出更严重的生育力低下。因此，我们的数据表明，编码Pate5 - 13区域内的多个附睾头富集基因协同作用以确保小鼠的雄性生育能力。
