RAT ZONA-PELLUCIDA GLYCOPROTEINS - MOLECULAR-CLONING AND CHARACTERIZATION OF THE 3 MAJOR COMPONENTS

The zona pellucida (ZP), the extracellular glycocalyx that surrounds t he oocyte, is well known to mediate homologous gamete interaction. In a previous study from our laboratories, we reported the qualitative ch aracterization of the rat ZP. The ZP in this species, like the mouse, hamster, and human, was found to contain three glycoproteins, namely r ZP1, rZP2, and rZP3 (Araki et al. [1992] Biol Reprod 46:912-919). In t his study, cDNAs encoding whole rat ZP major components have been isol ated and characterized, A rat ovary cDNA library was screened with the mouse ZP3 and ZP2 cDNA probes, respectively. For rZP1 cDNA cloning, c DNAs generated using reverse transcriptase-polymerase chain reaction a nd rapid amplification of 5' and 3' cDNA ends, were isolated and seque nced. The rZP3 cDNA showed 1338 bp with a coding region containing 127 2 bp, that translates into 424 amino acids. The total translation of r ZP3 peptide has a molecular weight of 45,820, containing six potential N-glycosylation sites and 75 Ser/Thr residues, possible O-glycosylati on sites. The amino acid sequence derived from the cDNA sequence share s high sequence homologies to mouse (90%), hamster (78%), and human (6 5%) ZP3 (ZPC) glycoproteins, indicating that the vat and mouse ZP3 hav e quite a conserved amino acid sequence, including the potential glyco sylation sites. The total transcript of the rZP2 was 2154 nucleotides and the largest open reading frame was 695 amino acids. This would tra nslate into a protein of 78.4 kDa. In the case of rZP1, the cDNA clone consisted of 1960 bp, and the coding region contained 1851 bp transla ting into 617 amino acids. Significant homologies were observed betwee n rZP2 and ZPA family from various mammalian species. The rZP1 also sh owed a sequence homology to mouse ZP1, known as a mouse orthologue of ZPB family, suggesting that the rZP2 and rZP1 are members of ZPA and Z PB families, respectively The message distributions for each zona comp onents were limited within the ovary and the signal was detectable in the growing oocytes. The present results will further our understandin g of the structure of rat zona components and lead to a better underst anding of species-specificity observed during sperm-egg interaction. M ol. Reprod. Dev. 51:454-467, 1998. (C) 1998 Wiley-Liss, Inc.