IDENTIFICATION OF MOUSE ZP3 PROTEIN IN MAMMALIAN OOCYTES WITH ANTISERA AGAINST SYNTHETIC ZP3 PEPTIDES

The mouse zona pellucida (ZP) protein ZP3 plays an important role in t he process of fertilization by mediating sperm binding and the acrosom e reaction. ZP3 primary structures are highly conserved, as revealed b y cDNA cloning. We raised antisera against synthetic peptides that are either conserved in the structure of ZP3 from different mammalian spe cies (AS ZP3-5 and AS ZP3-6) or specific for mouse ZP3 (AS ZP3-2). In ovary sections, AS ZP3-2 revealed immunoreactivity only to mouse ZP. A S ZP3-5 and AS ZP3-6 reacted with mouse, human, rat, hamster, porcine, and bovine ZP proteins. In porcine oocytes, immunoreactive material w as highly abundant in the ooplasm. Immunoblots showed that antiserum A S ZP3-5 recognized the mouse ZP3 protein. In porcine ZP preparations, AS ZP3-5 recognized a 53-kDa ZP protein. No reaction was observed with purified porcine ZP3 alpha or with ZP3 beta. Immunofluorescence studi es revealed that AS ZP3-5 and AS ZP3-6 antibodies react with ZP of iso lated porcine and human oocytes. Our results show that antisera agains t synthetic mouse ZP3 peptides can be used as markers for the identifi cation of ZP3-like proteins in mammalian oocytes and might be useful t ools for the evaluation of ZP integrity and ZP3 function.