Tyrosine phosphorylation of a 95 kDa protein and induction of the acrosome reaction 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 capacitated spermatozoa (48.6 +/- 3.2%; P < 0.01) compared with control (no rhZP3) samples (14.8 +/- 2.1%). However, rhZP3-containing medium did not cause increased acrosomal exocytosis in non-capacitated spermatozoa (16.8 +/- 3.0%). Induction of the AR was associated with increased tyrosine phosphorylation of a 95 +/- 5 kDa epitope only in capacitated spermatozoa. A dose-dependent increase in the protein phosphorylation of a 95 kDa epitope in response to rhZP3 was detected by [gamma-32P]-ATP labelling of detergent-solubilized sperm proteins. When spermatozoa were co-incubated with monoclonal antibody 97.25 (mAb 97.25) recognizing a 95 kDa tyrosine kinase epitope, there was no rhZP3 induction of tyrosine phosphorylation of the 95 kDa protein. Such co-incubation also markedly inhibited the AR (23.9 +/- 3.1%). These results support the model that initial interaction of the fertilizing spermatozoon with ZP3 involves the tyrosine phosphorylation of a 95 kDa tyrosine kinase protein and that this requires capacitation.
Brewis
I A
IA
Department of Molecular Biology and Biotechnology, The University of Sheffield, UK.
Clayton
R
R
Browes
C E
CE
Martin
M
M
Barratt
C L
CL
Hornby
D P
DP
Moore
H D
HD
D020101:Acrosome Reaction / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015153:Blotting, Western; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D004527:Egg Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D000939:Epitopes; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D010766:Phosphorylation; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D013094:Spermatozoa / Q000187:drug effects / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*; D015044:Zona Pellucida / Q000378:metabolism*; D000072216:Zona Pellucida Glycoproteins
D000911:Antibodies, Monoclonal; D004527:Egg Proteins; D000939:Epitopes; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C000606517:ZP3 protein, human; D000072216:Zona Pellucida Glycoproteins; D014443:Tyrosine