| Abstract: | To initiate fertilization in mice, free-swimming sperm bind to mZP3, an approximately 83-kDa glycoprotein present in the ovulated egg zona pellucida (ZP). mZP3 is located periodically along the filaments that constitute the ZP. Sperm recognize and bind to specific oligosaccharides linked to one or more of five Ser residues clustered in the carboxy-terminal one-third of the mZP3 polypeptide. When all five Ser residues are converted to nonhydroxy amino acids by site-directed mutagenesis of the mZP3 gene, an inactive form of mZP3, called mZP3ser], is secreted by embryonal carcinoma cells stably transfected with the mutated gene. Here, seven independent transgenic mouse lines were established that harbor the mutated mZP3 gene. In all lines, the mutant gene is expressed by growing oocytes and mZP3ser] is synthesized, secreted, and incorporated into the ZP. Purified mZP3ser] prepared from ovaries of transgenic mice, like mZP3ser] from transfected embryonal carcinoma cells, is inactive in sperm binding assays in vitro. On the other hand, the presence of mZP3ser] in the ZP does not significantly affect either the binding of sperm to ovulated eggs in vitro or the reproduction of the mice, i.e., the transgenic mice are fertile, breed at normal intervals, and produce litters of normal sizes. These results indicate that the number of functional sperm receptors in the ZP can be reduced by more than 50% without adversely affecting fertilization of eggs in vivo. |
