Localization of Plasma Membrane Calcium ATPase 2 and 4 (PMCA2 and PMCA4) in Chicken Sperm

Abstract

Abstract Calcium ions play important roles in several cellular processes including signalling pathway, membrane fusion and cell adhesion. In sperm, it is known that intracellular calcium concentration ([Ca2+]i) regulates acrosome exocytosis and flagellar motility. [Ca2+]i is generally regulated by influx and efflux of calcium ion. Plasma membrane calcium ATPase (PMCA) pumps the [Ca2+]i excess, providing a fine-tuning of [Ca2+]i concentration for sperm functions. To date, information related PMCA family in chicken sperm is still not documented yet. Therefore, this study was subjected to characterize expression and localization of PMCA family in chicken sperm. In this study, we selected PMCA2 and PMCA4 as representative to a specific tissue and a housekeeping isoform, respectively. Immunohistochemistry of PMCA2 and PMCA4 in chicken testis showed that both transmembrane proteins are expressed at the edge adluminal of the seminiferous tubule, indicating that these proteins are present in morphologically matured sperm that completed spermatogenesis. This was confirmed by immunocytochemistry, showing that PMCA2 and PMCA4 are highly enriched in the midpiece of chicken sperm. Furthermore, sub-cellular fractionation, combined with western blotting, showed that both isoforms are exclusively found in the membrane fraction of the sperm, together suggesting both PMCA are localized to the plasma membrane overlaying midpiece.