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BOR - Papers in Press, published online ahead of print April 28, 2004.
Biol Reprod 2004, 10.1095/biolreprod.104.028779
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BIOLOGY OF REPRODUCTION 71, 661–668 (2004)
DOI: 10.1095/biolreprod.104.028779
© 2004 by the Society for the Study of Reproduction, Inc.

Ovary

Biosynthesis of the Canine Zona Pellucida Requires the Integrated Participation of Both Oocytes and Granulosa Cells1

Daniel G. Blackmore3, Lucan R. Baillie3, Janet E. Holt3, Lynda Dierkx3, R. John Aitken3,4, and Eileen A. McLaughlin2,3,4

Reproductive Science Group, Discipline of Biological Sciences3 ARC Centre of Excellence in Biotechnology and Development,4 School of Environmental & Life Sciences, University of Newcastle, Callaghan, New South Wales 2308, Australia

In the dog, attempts to localize the expression of zona pellucida (ZP) proteins during folliculogenesis have failed to demonstrate conclusively whether any or all of the zona proteins are synthesized in the oocyte or the granulosa cells. Probing of paraformaldehyde-fixed prepubertal canine ovarian tissue sections with a panel of fluorescently conjugated lectins localized the expression of glycoproteins during folliculogenesis. We confirm that six lectins (PSA, s-WGA, ECL, GSL-II, LEL, and STL) consistently labeled the ZP and adjacent granulosa cells of the developing follicle and that canine ZP expresses ß-gal(1,4)glcNAc, ß-gal(1,3)galNac, {alpha}-mannose, and terminal sialic acid residues in a developmentally specific manner. Riboprobes for canine ZPA and ZPC genes were produced and used for in situ hybridization studies of mRNA expression in canine folliculogenesis. In addition, we isolated a partial cDNA transcript from total ovarian RNA for the canine ZPB gene having a high degree of sequence identity with the felid and porcine ZPB homologues. Subsequently, the ZPA gene transcripts were localized to the cytoplasm of oocytes in primordial, primary, and early secondary follicles. We then localized expression of ZPB and ZPC gene transcripts to the granulosa cells of growing follicles, but not in squamous granulosa cells of primordial follicles or oocytes. These observations indicate that in the juvenile canine ovary, the oocyte is responsible for synthesis of the ZPA protein and directing synthesis of the ZPB and ZPC proteins by the granulosa cells and that ZP gene transcription occurs in a sequential manner during folliculogenesis.

1 Supported by an Australian Research Council SPIRT grant to R.J.A.

2 Correspondence. FAX: 61 2 4921 6923; eileen.mclaughlin{at}newcastle.edu.au






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Copyright © 2004 by the Society for the Study of Reproduction.