Gene expression during oogenesis plays an important role in early mammalian development. The genes coding for the mouse zona pellucida are expressed during oogenesis where they are translated into three sulfated glycoproteins designated ZP1, ZP2 and ZP3. We have isolated cDNAs coding for ZP2 and ZP3 from an ovarian lambda gt11 expression library using monoclonal antibodies specific to ZP2 and ZP3. The identity of the ZP3 clone has been confirmed by a comparison of its nucleic acid sequence with the amino acid sequence of an internal ZP3 peptide. By Northern blot analysis and in situ hybridization we have shown that ZP3 is expressed uniquely in oocytes as a 1.5 kb poly(A)+ mRNA. ZP3 transcripts are not detectable in resting oocytes (15 um) but become very abundant during oocyte growth and represent 0.1-0.2% of the poly(A)+ RNA in 50 um diameter oocytes. There is a subsequent dramatic fall-off of ZP3 transcripts in the latter stages of oocyte growth (65 um) which closely parrellels the decline in zona protein synthesis. ZP3 is a single copy gene and is located on mouse chromosome 6. There appears to be neither gene amplification nor gene rearrangement to account for the tissue specific expression of ZP3, although the ZP3 locus is hypomethylated in ovarian tissue (where it is expressed) compared to somatic tissue. We have isolated genomic clones containing both the mouse and human ZP3 genes and are in the processes of defining their intron and axon structures. We are particularly interested in investigating the 5' flanking regions which may modulate the tissue specific expression of the zona genes. We have previously reported that monoclonal antibodies to the ZP2 and ZP3 were effective, longterm but eventually reversible contraceptive agents. Based on our recent ability to clone the zona genes, we are now exploring the use of synthetic zona peptides and ZP/beta-galactosidase fusion proteins as active immunogens for the development of a contraceptive vaccine.
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