Nine genes abundantly expressed in the epididymis are not essential for male fecundity in mice
Nine genes abundantly expressed in the epididymis are not essential for male fecundity in mice
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DOI：
10.1111/andr.12621
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发表时间：
2019-09-01
期刊：
影响因子：
4.5
通讯作者：
Ikawa, M.
中科院分区：
文献类型：
--
作者：
Noda, T.;Sakurai, N.;Ikawa, M.
精子在通过附睾的过程中具有受精能力。由于来自近端附睾尾部的精子可以使卵子受精，因此精子成熟需要来自附睾头和附睾体的蛋白质。目的基因芯片分析表明，超过17,000个基因在附睾中表达，然而，这些基因的表达仅限于附睾。为了分析附睾富集的基因在体内的功能，我们产生了敲除（KO）突变的9个基因，大量表达在附睾的头和体区。使用CRISPR/Cas9系统产生KO小鼠。苏木精-伊红染色观察附睾组织学变化。将KO雄性与野生型雌性关在一起3-6个月以检查生育力。结果我们产生了在Pate 1、Pate 2或Pate 3中具有插入缺失突变的个体突变小鼠系。我们还独立地删除了Clps 12、Epp 13和Rnase 13的编码区。最后，缺失编码Gm 1110、Glb 112和Glb 113的150 kb区域，以产生三重KO小鼠系。所有KO系的附睾组织学和精子形态与对照雄性相当。与这些KO雄性交配的雌性动物产下的幼仔数量与对照雄性动物相当。讨论和结论我们发现9个基因在附睾头和附睾体中高表达，它们与精子功能和男性生殖力有关。CRISPR/Cas9介导的KO小鼠的产生加速了附睾富集基因在生殖中的潜在功能的筛选。
Background Spermatozoa become competent for fertilization during transit through the epididymis. As spermatozoa from the proximal caudal epididymis can fertilize eggs, proteins from the caput and corpus epididymis are required for sperm maturation. Objectives Microarray analysis identified that more than 17,000 genes are expressed in the epididymis; however, few of these genes demonstrate expression restricted to the epididymis. To analyze epididymis-enriched gene function in vivo, we generated knockout (KO) mutations in nine genes that are abundantly expressed in the caput and corpus region of the epididymis. Materials and methods KO mice were generated using the CRISPR/Cas9 system. The histology of the epididymis was observed with hematoxylin and eosin staining. KO males were caged with wild-type females for 3-6 months to check fertility. Results We generated individual mutant mouse lines having indel mutations in Pate1, Pate2, or Pate3. We also deleted the coding regions of Clpsl2, Epp13, and Rnase13, independently. Finally, the 150 kb region encoding Gm1110, Glb1l2, and Glb1l3 was deleted to generate a triple KO mouse line. Histology of the epididymis and sperm morphology of all KO lines were comparable to control males. The females mated with these KO males delivered pups at comparable numbers as control males. Discussion and conclusion We revealed that nine genes abundantly expressed in the caput and corpus epididymis are dispensable for sperm function and male fecundity. CRISPR/Cas9-mediated KO mice generation accelerates the screening of epididymis-enriched genes for potential functions in reproduction.