MESSENGER-RNA ENCODING AN ESTROGEN-DEPENDENT OVIDUCT SECRETORY PROTEIN IN THE SHEEP IS LOCALIZED IN THE APICAL TIPS AND BASAL COMPARTMENTS OF FIMBRIA AND AMPULLA EPITHELIAL-CELLS IMPLYING TRANSLATION AT UNIQUE CYTOPLASMIC FOCI

Data from our laboratory have shown that an estrogen (E(2))-dependent Mr 90,000-92,000 protein and its mRNA are synthesized and expressed in abundant amounts at estrus from the fimbria and ampulla, not isthmus, oviduct of the sheep. Immunocytochemical studies have shown that the Mr 90,000-92,000 protein is contained in apical secretory granules of oviduct epithelial cells. The objective of this study was to determine whether the mRNA for the E(2)-dependent oviduct protein was localized and compartmentalized in similar manner. Fimbria, ampulla, and isthmus oviducts obtained from estrous ewes were flash frozen in liquid nitrogen, cryosectioned, fixed in 4% paraformaldehyde, hybridized with digoxigenin (DIG)-labeled oviduct-specific riboprobes, incubated in anti-DIG antibodies conjugated with alkaline phosphatase, and developed in color substrate. Oviduct protein-specific transcripts were localized to basal perinuclear compartments and, surprisingly, at sites distant from the nucleus in the apical cytoplasm of epithelial cells in the fimbria and ampulla. No specific reaction product was observed in the underlying mucosa or smooth muscle layers. Oviduct protein mRNA was contained predominantly in the apical cytoplasm of epithelial cells at the free margins of mucosal folds and in the basal regions of cells located at the crypts of longitudinal folds. No reaction product was present when sections of the fimbria and ampulla oviduct of estrous ewes were incubated in sense riboprobe to the oviduct protein. In addition, when sections of the isthmus oviduct obtained from estrous ewes or fimbria and ampulla oviducts from long-term ovariectomized ewes were hybridized with antisense riboprobes no specific reaction product was detected. Electron microscopy of oviduct protein mRNA containing areas revealed the presence of secretory granules, rough endoplasmic reticulum (RER) and Golgi in the apical cytoplasm, and RER in the basal regions of epithelial cells. These data show that the mRNA encoding an E(2)-dependent oviduct-specific protein is distributed in epithelial cells at perinuclear foci and at sites distant from the nucleus, which are also the sites of protein localization and protein synthesizing organelles, implying translation at unique cytoplasmic foci. (C) 1995 Wiley-Liss, Inc.