PubMed · 352623
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C R Austin. 1978. Patterns in metazoan fertilization.. https://doi.org/10.1016/s0070-2153(08)60591-8
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A 95-kDa mouse sperm protein has been previously identified as a putative receptor involved in the sperm-egg interactions that lead to fertilization. The ligand for this receptor is the zona pellucida glycoprotein ZP3. This constituent of the oocyte-specific extracellular matrix mediates not only sperm binding to the zona but also triggers acrosomal exocytosis. The latter, also termed the acrosome reaction, is a key regulatory event upon which fertilization is absolutely dependent. Previously, we showed that the 95-kDa protein that binds ZP3 is a substrate for tyrosine kinase, and its phosphotyrosine content increases after sperm-zona pellucida binding. Here, we show the presence of protein tyrosine kinase activity in sperm plasma membranes and in electroeluted 95-kDa protein. The tyrosine kinase activity of the isolated protein is stimulated by solubilized zona pellucida and inhibited by tyrphostin RG-50864, a membrane-permeable tyrosine kinase inhibitor. Furthermore, tyrphostin inhibits zona-triggered acrosomal exocytosis in a dose-dependent manner. These findings indicate that the 95-kDa protein participates in a critical regulatory event of gamete interaction; moreover, our experiments suggest that sperm protein tyrosine kinase may be an excellent target for the control of fertility.
Monoclonal antibodies (mAbs) against sperm cells are currently being used in an effort to define spermatozoal antigens involved in the fertilization process. We have produced a number of anti-human sperm mAbs by immunization of female mice with the 100,000 x g supernatant of octylglycoside-solubilized washed human sperm. From a panel of mAbs, 1 antibody, AG7, was selected and characterized due to its fertilization-inhibiting characteristics. MAb AG7 defines a sperm acrosome antigen-1 (SAA-1) located in the acrosomal region of human sperm as evaluated by indirect immunofluorescence. Staining of life sperm cells indicated that the antigen is present on the sperm surface. SAA-1 was also found on sperm of several other mammalian species, implying evolutionary conservation of the antigen. SAA-1 was first observed on testicular sperm and can be followed through epididymal transit, ejaculation, and capacitation. When applied in a mouse in vitro fertilization assay, mAb AG7 inhibits fertilization by greater than 95%, and inhibition is dose dependent, with half-maximal inhibition at 0.8 micrograms/ml. The block to fertilization could not be attributed to sperm agglutination, inhibition of motility, interference with adhesion to the zona pellucida, or inhibition of fusion with the oocyte membrane. MAb AG7 was demonstrated to inhibit calcium influx in spermatozoa in vitro (measured using the fluorescent indicator fura 2), a prerequisite for the acrosome reaction. Initial biochemical characterization of the antigen suggests it is proteinlike in nature, with a molecular weight of approximately 220 kD. The results suggest that SAA-1, identified by mAb AG7, is a sperm antigen crucially involved in the fertilization process, possibly an atypical steroid receptor or ion channel located within the sperm plasma membrane.
The presence of intranuclear and acrosomal lectin binding sites in spermatids and spermatozoa of the mosquito Culex quinquefasciatus was analysed. Direct and indirect lectin-gold techniques were used on LR White-embedded cells. The nuclear compartment was the structure most intensely labelled. Early spermatid nucleus showed moderate labelling for peanut agglutinin (PNA), Griffonia simplicifolia IB4 (GS-IB4) and Ricinus communis agglutinin (RCA), and light labelling for the other lectins tested. The sperm nucleus was intensely labelled by all lectins. The acrosome, an enzyme-containing structure, was labelled by some lectins. The anterior acrosomal region was labelled by PNA, while the proximal acrosomal region was labelled by PNA and G. simplicifolia II (GS II) lectins, and showed the presence of fucose residues with the use of Ulex europaeus I (UEA-I) lectin. The spermatozoa stored in the spermatheca showed the same pattern of labelling as that observed in spermatozoa localized in testis and seminal vesicles for all lectins tested. Carbohydrate residues in the nuclear compartment may be involved with the process of chromatin condensation. In the acrosomal region these residues may play a role in the process of sperm-oocyte interaction.