The Na,K-ATPase α4 isoform from humans has distinct enzymatic properties and is important for sperm motility

In the rat, the Na,K-ATPase alpha 4 isoform exhibits unique enzymatic characteristics and is important for sperm motility. In this work, we studied expression, localization and function of alpha 4 in human spermatozoa. We show two catalytically active Na,K-ATPase alpha polypeptides with different ouabain affinity and identified expression of alpha 1, alpha 4, beta 1 and beta 3 isoforms in the gametes. In addition, human sperm presented two Na,K-ATPases composed of alpha 4, alpha 4 beta 1 and alpha 4 beta 3. Kinetic analysis of these isozymes produced in insect cells showed that, compared with human alpha 1 beta 1, alpha 4 beta 1 and alpha 4 beta 3 exhibit higher Na+ and lower K+ affinity and higher sensitivity to ouabain. These particular enzymatic properties suggested a role for alpha 4 in sperm function. Using computer-assisted sperm analysis (CASA), we found that ouabain inhibition of alpha 4 significantly decreased percentage sperm motility. In contrast, ouabain did not affect linearity of forward progression, amplitude of lateral head displacement, beat cross frequency and sperm straight-line, curvilinear or average path velocities. This suggests a primary role of alpha 4 in flagellar motility. Accordingly, we found alpha 4 in the sperm tail, predominating in the mid-piece of the flagellum. Therefore, similar to the rat ortholog, human Na,K-ATPase alpha 4 isoform has a distinct activity that is essential for sperm function.