Capacitation dependent activation of tyrosine phosphorylation generates two sperm head plasma membrane proteins with high primary binding affinity for the zona pellucida.
The recognition and binding of sperm cells to the zona pellucida (the extracellular matrix of the oocyte) are essential for fertilization and are believed to be species specific. Freshly ejaculated sperm cells do not bind to the zona pellucida. Physiologically this interaction is initiated after sperm activation in the female genital tract (capacitation) via a yet unknown mechanism, resulting in the binding of a receptor in the apical sperm plasma membrane to the zona pellucida. In order to mimic this biochemically, we isolated zona pellucida fragments from gilt ovaries to prepare an affinity column with the intact zona pellucida structure and loaded this column with solubilized apical plasma membranes of boar sperm cells before and after in vitro capacitation. With this technique we demonstrated that two plasma membrane proteins of capacitated boar sperm cells showed high affinity for zona pellucida fragments. Further analysis showed that these proteins were tyrosine phosphorylated. Plasma membrane proteins from freshly ejaculated sperm cells did not exhibit any zona pellucida binding proteins, likely because these proteins were not tyrosine phosphorylated.
Flesch
F M
FM
Institute of Biomembranes, Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
Wijnand
E
E
van de Lest
C H
CH
Colenbrander
B
B
van Golde
L M
LM
Gadella
B M
BM
D020101:Acrosome Reaction; D000818:Animals; D015153:Blotting, Western; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D004789:Enzyme Activation; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008858:Microscopy, Phase-Contrast; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D011485:Protein Binding; D013075:Sperm Capacitation; D013077:Sperm Head / Q000378:metabolism*; D013552:Swine; D015044:Zona Pellucida / Q000378:metabolism*
D008565:Membrane Proteins; D019000:Phosphotyrosine