A NOVEL SIGNAL-TRANSDUCTION CASCADE IN CAPACITATING HUMAN SPERMATOZOACHARACTERIZED BY A REDOX-REGULATED, CAMP-MEDIATED INDUCTION OF TYROSINE PHOSPHORYLATION

Capacitation is a priming event that renders mammalian spermatozoa res ponsive to signals originating from the cumulus-oocyte complex. The at tainment of a capacitated state is dependent upon an increase in tyros ine phosphorylation and results in the acquisition of responsiveness t o physiological agonists such as progesterone and ZP3. In this study w e have shown that this capacitation-dependent increase in tyrosine pho sphorylation is controlled by a unique redox-regulated, cAMP-mediated, signal transduction cascade. Either stimulation of reactive oxygen sp ecies generation or elevation of intracellular cAMP induced increases in phosphotyrosine expression by human spermatozoa and enhanced their responsiveness to progesterone, Ultimate convergence of the redox-and cAMP-regulated pathways was indicated by the ability of the protein ki nase A inhibitor, H89, to block both modes of signal transduction. Fur thermore, the fact that the redox-regulated pathway could be silenced by catalase, while this enzyme had no effect on the cAMP-mediated resp onse, indicated that oxidant generation must lie upstream from cAMP in the reaction sequence. In keeping with this conclusion, a functional association was demonstrated between the redox status of human spermat ozoa and their cAMP content. The continuous production of reactive oxy gen species was also shown to be necessary for the protein kinase A-ty rosine phosphorylation axis to remain functional. If the generation of oxidising conditions during capacitation was prevented with 2-mercapt oethanol, 2-deoxyglucose or the flavoprotein inhibitor, diphenylene io donium, then cAMP could no longer trigger tyrosine phosphorylation. Th ese data support a model for human sperm capacitation as a redox-regul ated process, involving a unique sequence of interactive events includ ing reactive oxygen species production, elevation of intracellular cAM P, stimulation of protein kinase A and the induction of tyrosine phosp horylation. This is the first report of such a signal transduction cas cade and may have implications for the functional significance of reac tive oxygen metabolites in other cell types.