An autoradiographic analysis of N-linked glycoconjugates in embryonic salivary gland morphogenesis

The synthesis, deposition, and loss of mannose-bearing glycoconjugates during branching morphogenesis of embryonic mouse salivary glands has been evaluated. Day 13 embryonic mouse salivary glands were cultured for 44 hr, pulse labeled 4 hr with [3H]mannose, then fixed after 0, 2, 4, 8, or 24 hr of chase in nonradioactive medium, and processed for autoradiography. Light microscopic autoradiograms of sectioned rudiments reveal extensive label within the epithelium, little label over the mesenchyme, and a concentration of radioactivity at the basal surface of the epithelium. Autoradiograms of "chased" rudiments reveal a) no detectable loss of label from the epithelium or the basal epithelial surface over the first 8 hr, and b) significant label loss by 24 hr of chase at the basal epithelial surface, while moderate amounts of radioactivity remain throughout the rest of the epithelium. The [3H]bound material is insensitive to chondroitinase ABC, a glycosaminoglycan degradative enzyme, but is sensitive to tunicamycin presence in the culture medium. Earlier studies showed that embryonic mouse salivary glands cultured in medium containing tunicamycin (25 ng/ml) continued normal epithelial branching while epithelial growth was inhibited. The present autoradiographic studies of [3H]mannose-labeled rudiments demonstrate that tunicamycin causes a significant decrease in radioactivity, relative to controls. Thus, our results suggest that epithelial branching activity is independent of control levels of mannose-containing/tunicamycin-sensitive, glycoconjugate deposition.