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Dynamics of the Global Tyrosine Phosphorylation During Capacitation and Acquisition of the Ability to Fuse with Oocytes in Human Spermatozoa¹

Andrologic Unit, Department of Internal Medicine,³ and Department of Experimental Medicine,⁴ University of L'Aquila, 67100 L'Aquila, Italy

ABSTRACT

Phosphorylation of tyrosine residues in cellular proteins represents a major event during sperm capacitaton, but its relationship with the acquisition of sperm-fertilizing ability is still unclear. In this study we explored the relationship between the kinetics of the global tyrosine phosphorylation, monitored with a flow cytometric assay, and the acquisition of the human sperm ability to fuse with oocytes, evaluated with the progesterone-enhanced hamster egg penetration test. Sperm tyrosine phosphorylation appeared to be an early event in the capacitation process, with a 3.6-fold mean increase within 1 h of capacitation, but at this time sperm-oocyte fusion was extremely poor compared with that observed at 5 h of capacitation. Capacitation in calcium-free medium produced a 2-fold mean increase in tyrosine phosphorylation compared with that seen in complete capacitation medium both at 1 h and 5 h of capacitation, whereas sperm-oocyte fusion significantly increased only at 1 h, remaining unchanged at 5 h of capacitation. The cAMP analog, N,2-O-dibutyryladenosine 3',5'-cyclic monophosphate (dbcAMP), prevented the inhibitory effect of seminal plasma on tyrosine phosphorylation but not on sperm-oocyte fusion. In conclusion, these results suggest that the acquisition of sperm-fertilizing ability is always associated with an increase of the global tyrosine phosphorylation, but tyrosine phosphorylation does not necessarily reflect the acquisition of the sperm-fertilizing ability. Flow cytometry assay, a reliable technique to quickly quantify the global levels of the human sperm tyrosine phosphorylation, could be useful for a further elucidation of the biological meaning of this process, with the perspective of its clinical use as a measure of the sperm-fertilizing potential.

flow cytometry, human spermatozoa, sperm capacitation, sperm/oocyte fusion, tyrosine phosphorylation

¹Supported by MIUR, COFIN 2005, and COFIN 2006, Italy.

Correspondence: ²Felice Francavilla, Department of Internal Medicine, University of L'Aquila, Blocco 11, Coppito, 67100 L'Aquila, Italy. FAX: 39 0862 368342; e-mail: francavi{at}cc.univaq.it