Department of Veterinary & Animal Sciences, University of Massachusetts-Amherst, Amherst, Massachusetts, USA.
Department of Molecular and Cellular Biology Graduate Program, University of Massachusetts-Amherst, Amherst, Massachusetts, USA.
Mol Reprod Dev. 2021 Nov;88(11):718-730. doi: 10.1002/mrd.23539. Epub 2021 Oct 8.
We have previously shown that members of the family of testis-specific serine/threonine kinases (TSSKs) are post-meiotically expressed in testicular germ cells and in mature sperm in mammals. The restricted post-meiotic expression of TSSKs as well as the importance of phosphorylation in signaling processes strongly suggest that TSSKs have an important role in germ cell differentiation and/or sperm function. This prediction has been supported by the reported sterile phenotype of the TSSK6 knock-out (KO) mice and of the double TSSK1/TSSK2 KO. The aim of this study was to develop KO mouse models of TSSK3 and to validate this kinase as a target for the development of a male contraceptive. We used CRISPR/Cas9 technology to generate the TSSK3 KO allele on B6D2F1 background mice. Male heterozygous pups were used to establish three independent TSSK3 KO lines. After natural mating of TSSK3 KO males, females that presented a plug (indicative of mating) were monitored for the following 24 days and no pregnancies or pups were found. Sperm numbers were drastically reduced in all three KO lines and, remarkably, round spermatids were detected in the cauda epididymis of KO mice. From the small population of sperm recovered, severe morphology defects were detected. Our results indicate an essential role of TSSK3 in spermiogenesis and support this kinase as a suitable candidate for the development of novel nonhormonal male contraceptives.
我们之前已经表明，睾丸特异性丝氨酸/苏氨酸激酶（TSSK）家族的成员在哺乳动物的睾丸生殖细胞和成熟精子中都有减数分裂后表达。TSSK 的减数分裂后表达受限以及磷酸化在信号转导过程中的重要性强烈表明，TSSK 在生殖细胞分化和/或精子功能中具有重要作用。TSSK6 KO（敲除）小鼠和 TSSK1/TSSK2 双 KO 的不育表型已经支持了这一预测。本研究的目的是开发 TSSK3 KO 小鼠模型，并验证该激酶是开发男性避孕药的靶标。我们使用 CRISPR/Cas9 技术在 B6D2F1 背景小鼠中生成 TSSK3 KO 等位基因。雄性杂合子幼鼠用于建立三条独立的 TSSK3 KO 系。在 TSSK3 KO 雄性自然交配后，出现栓（交配迹象）的雌性被监测接下来的 24 天，没有发现怀孕或幼仔。所有三条 KO 系中的精子数量都明显减少，而且，令人惊讶的是，KO 小鼠的附睾尾部可以检测到圆形精子细胞。从小部分回收的精子中，检测到严重的形态缺陷。我们的结果表明 TSSK3 在精子发生中具有重要作用，并支持该激酶作为开发新型非激素男性避孕药的合适候选物。
