Regulation of the sperm EGF receptor by ouabain leads to initiation of the acrosome reaction

Abstract

The sperm acrosome reaction occurs after the binding of the capacitated sperm to the egg zona pellucida. This study describes a novel mode of regulation of the sperm epidermal growth factor receptor (EGFR) under physiological conditions and its relevance to the acrosome reaction. Ouabain, a known Na/K ATPase blocker is present in the blood and in the female reproductive tract. We show here that physiological concentrations (nM) of ouabain enhance phosphorylation of EGFR on tyr-845, stimulate Ca(2+) influx and induce the acrosome reaction in sperm. These effects could be seen only in the presence of very low concentrations of EGF (0.1 ng/ml or 0.016 nM) added together with nano-molar ouabain. Phosphorylation, Ca(2+) influx, and the acrosome reaction are inhibited by an EGFR blocker, suggesting that trans-activation of the EGFR is involved. Moreover, our data revealed that protein kinase A and the family of tyrosine kinase, SRC, shown before to be involved in EGFR activation in sperm, mediate the acrosome reaction induced by ouabain. Ouabain alone (without EGF) at relatively high concentration (10microM) could enhance EGFR phosphorylation, Ca(2+) influx and acrosome reaction, and these processes were inhibited by EGFR blockers. Moreover, we show here that PKA and SRC family are involved in the activation of EGFR by 10 microM ouabain, further demonstrating that ouabain induces the acrosome reaction by a mechanism mediated by the trans-activation of EGFR. In conclusion, this study describes an interesting regulatory path of EGFR by physiological concentrations of ouabain and EGF found in the female reproductive tract. Neither of these compounds can activate the EGFR alone at such low physiological levels; however, when both are present, the interaction of ouabain with the Na/K ATPase leads to the priming of the EGFR, which undergoes its full activation by EGF.

Endogenous ouabain regulates cell viability

“…This implies that EO not only has a stimulatory effect on cell viability but also may affect stimulation of growth caused by other growth factors. Indeed, numerous studies (12,30) have shown a tight crosstalk between several growth factors, including insulin, insulin growth factor -1, and EGF with Na ϩ -K ϩ -ATPase and CS, although the mechanisms involved in these interactions are yet to be elucidated.…”

Calcium Channels in the Development, Maturation, and Function of Spermatozoa

“…Other cellular ligands can induce the AR such as progesterone (31, 55, 355, 506), ␥-aminobutyric acid (GABA) (280,355,559), glycine (301), EGF (133), ATP (339,442), acetylcholine (66), and sphingosine-1-phosphate (495). The role of many of these "alternative" agonists is not clear.…”

“…IVF), protein tyrosine kinases are present in mammalian spermatozoa, and their antagonists can inhibit ZP (317,420), progesterone (294,338,354,429,504), and thapsigargin-induced AR (156). Tyrosine kinases such as Src (detected in bovine spermatozoa) were shown to participate together with PKA and ouabain (a specific Na ϩ -K ϩ -ATPase inhibitor present in the female tract) in the transactivation of EGF receptor (EGFR) leading to [Ca 2ϩ ] i increases and the AR (133). Tyrosine kinases and phosphatases have indeed been proposed as key elements for AR.…”

Sperm Epidermal Growth Factor Receptor (EGFR) Mediates α7 Acetylcholine Receptor (AChR) Activation to Promote Fertilization

“…In two recent publications we further emphasized the role and regulation of EGFR by G protein-coupled receptors and ouabain (50,68). Hence, we FIGURE 3.…”