Tyrosine phosphorylation of a 95 kDa protein and induction of the acrosomereaction in human spermatozoa by recombinant human zona pellucida glycoprotein 3

Protein tyrosine phosphorylation and induction of the acrosome reaction (AR ) in non-capacitated and capacitated human spermatozoa was investigated in response to recombinant human zona pellucida glycoprotein (rhZP3) produced by Chinese hamster ovary cells transfected with a plasmid containing human ZP3 cDNA. rhZP3-containing medium promoted the AR in a high proportion of c apacitated spermatozoa (48.6 +/- 3.2%; P < 0.01) compared with control (no rhZP3) samples (14.8 +/- 2.1%). However, rhZP3-containing medium did not ca use increased acrosomal exocytosis in non-capacitated spermatozoa (16.8 +/- 3.0%). Induction of the AR was associated with increased tyrosine phosphor ylation of a 95 +/- 5 kDa epitope only in capacitated spermatozoa. A dose-d ependent increase in the protein phosphorylation of a 95 kDa epitope in res ponse to rhZP3 was detected by [gamma-P-32]-ATP labelling of detergent-solu bilized sperm proteins. When spermatozoa were co-incubated with monoclonal antibody 97.25 (mAb 97.25) recognizing a 95 kDa tyrosine kinase epitope, th ere was no rhZP3 induction of tyrosine phosphorylation of the 95 kDa protei n. Such coincubation also markedly inhibited the AR (23.9 +/- 3.1%). These results support the model that initial interaction of the fertilizing sperm atozoon with ZP3 involves the tyrosine phosphorylation of a 95 kDa tyrosine kinase protein and that this requires capacitation.