Localization of the lipid intermediate pathway of protein glycosylation in oviduct cell types.
Oviduct tissue slices were incubated with [3H]-leucine or [3H]-mannose in the presence and absence of tunicamycin, a specific inhibitor of lipid-mediated protein glycosylation. Conditions were established where tunicamycin had maximal effect on [3H]-mannose incorporation (greater than 90% inhibition) but a minimal effect on [3H]-leucine incorporation (less than 10% inhibition) into total TCA-insoluble products. Analysis of incubated tissues by SDS-polyacrylamide gel electrophoresis revealed that in the absence of tunicamycin, [3H]-mannose was incorporated into only a few proteins, of which ovalbumin represented the major radiolabeled component. Tunicamycin markedly reduced the incorporation of [3H]-mannose into ovalbumin and other oviduct glycoproteins. In contrast, analysis by SDS-polyacrylamide gel electrophoresis showed that [3H]-leucine was incorporated into a variety of proteins in the absence of tunicamycin. The radioactivity profile of some of these proteins was shifted toward lower Mr when oviduct slices were incubated in the presence of tunicamycin, with only a minimal decrease in protein labeling. Light microscopic autoradiograms of tissue incubated with [3H]-leucine in either the presence or absence of tunicamycin exhibited extensive labeling of tubular gland and epithelial cells. In the absence of tunicamycin, these cell types also become markedly labeled with [3H]-mannose; however, incorporation of label in both cell types was substantially reduced in the presence of tunicamycin. Qualitatively, labeling of tubular gland cells appeared greater than that of epithelial cells, largely due to the concentration of silver grains over the dense population of secretory vesicles in the tubular gland cells.(ABSTRACT TRUNCATED AT 250 WORDS)
J J
JJ
Lucas
C
C
Makunike
S F
SF
Basinger
S J
SJ
Fliesler
D000818:Animals; D001345:Autoradiography / Q000379:methods; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006023:Glycoproteins / Q000096:biosynthesis*; D007930:Leucine / Q000378:metabolism; D050356:Lipid Metabolism; D008358:Mannose / Q000378:metabolism; D010047:Ovalbumin / Q000096:biosynthesis; D010057:Oviducts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D014316:Tritium; D014415:Tunicamycin / Q000494:pharmacology
D006023:Glycoproteins; D014316:Tritium; D014415:Tunicamycin; D010047:Ovalbumin; D007930:Leucine; D008358:Mannose