科研之友
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摘要
The alpha 4 polypeptide is a testis-specific isoform of the catalytic subunit of the Na,K-ATPase, which is essential for sperm motility and fertility. In the present study, we have investigated the regulation of activity of the a4 isoform and the relevance of this event for sperm capacitation. We have performed this by taking advantage of the selective high affinity of alpha 4 for the inhibitor ouabain. Our results show that ouabain-sensitive hydrolysis of ATP and uptake of Rb-86, corresponding to the enzymatic and ion transport activities of alpha 4, respectively, increased during sperm capacitation in a time-dependent manner. Specific labeling of alpha 4 with the fluorescent indicator bodipy-ouabain and immunoblot analysis of biotinylated and streptavidin-precipitated sperm plasma membrane proteins indicated a capacitation- and time-dependent rise in levels of active alpha 4 isoform at the sperm surface. Ouabain inhibition of alpha 4 blocked the increase in total sperm motility and the hyperactive motility pattern characteristic of sperm capacitation. Moreover, interference of alpha 4 activity with ouabain partially prevented the intracellular decrease in Na+ and the. plasma membrane hyperpolarization that typically accompany sperm capacitation. In contrast, ouabain inhibition of alpha 4 did not affect the spontaneous sperm acrosomal reaction following capacitation. Together, these results demonstrate that Na,K-ATPase alpha 4 activity is up-regulated during sperm capacitation through mechanisms that involve both increases in molecular activity and levels of alpha 4 at the sperm plasma membrane. This increase in alpha 4 activity helps maintain the changes in motility that are associated with sperm capacitation, emphasizing the biologic relevance of the Na,K-ATPase isoform in sustaining sperm function.
- 出版日期2012-10
