McDermott Jeffrey, Sánchez Gladis, Nangia Ajay K, Blanco Gustavo
Department of Molecular and Integrative Physiology, Department of Urology and Institute for Reproductive Health and Regenerative Medicine, University of Kansas Medical Center, Kansas City, Kansas.
Mol Reprod Dev. 2015 Mar;82(3):167-81. doi: 10.1002/mrd.22454. Epub 2015 Jan 14.
Most of our knowledge on the biological role of the testis-specific Na,K-ATPase alpha 4 isoform derives from studies performed in non-human species. Here, we studied the function of human Na,K-ATPase alpha 4 after its expression in transgenic mice. Using a bacterial artificial chromosome (BAC) construct containing the human ATP1A4 gene locus, we obtained expression of the human α4 transgene specifically in mouse sperm testis and, in the sperm flagellum. The expressed human alpha 4 was active, and compared to wild-type sperm, those from transgenic mice displayed higher Na,K-ATPase alpha 4 activity and greater binding of fluorescently labeled ouabain, which is typical of the alpha 4 isoform. The expression and activity of endogenous alpha 4 and the other Na,K-ATPase alpha isoform present in sperm, alpha 1, remained unchanged. Male mice expressing the human ATP1A4 transgene exhibited similar testis size and morphology, normal sperm number and shape, and no changes in overall fertility compared to wild-type mice. Sperm carrying the human transgene exhibited enhanced total motility and an increase in multiple parameters of sperm movement, including higher sperm hyperactive motility. In contrast, no statistically significant changes in sperm membrane potential, protein tyrosine phosphorylation, or spontaneous acrosome reaction were found between wild-type and transgenic mice. Altogether, these results provide new genetic evidence for an important role of human Na,K-ATPase alpha 4 in sperm motility and hyperactivation, and establishes a new animal model for future studies of this isoform.
我们对睾丸特异性钠钾ATP酶α4亚型生物学作用的大部分认识都来自于在非人类物种中进行的研究。在此，我们研究了人类钠钾ATP酶α4在转基因小鼠中表达后的功能。利用包含人类ATP1A4基因座的细菌人工染色体（BAC）构建体，我们在小鼠精子睾丸以及精子鞭毛中特异性地获得了人类α4转基因的表达。所表达的人类α4具有活性，与野生型精子相比，转基因小鼠的精子显示出更高的钠钾ATP酶α4活性以及与荧光标记哇巴因更强的结合，这是α4亚型的典型特征。精子中存在的内源性α4以及其他钠钾ATP酶α亚型（α1）的表达和活性保持不变。与野生型小鼠相比，表达人类ATP1A4转基因的雄性小鼠睾丸大小和形态相似，精子数量和形状正常，总体生育能力无变化。携带人类转基因的精子表现出总运动能力增强以及精子运动多个参数增加，包括更高的精子超激活运动能力。相比之下，野生型和转基因小鼠之间在精子膜电位、蛋白质酪氨酸磷酸化或自发顶体反应方面未发现统计学上的显著变化。总之，这些结果为人类钠钾ATP酶α4在精子运动和超激活中的重要作用提供了新的遗传学证据，并建立了一个用于该亚型未来研究的新动物模型。
