Role of the Na,K-ATPase beta-subunit in the cellular accumulation and maturation of the enzyme as assessed by glycosylation inhibitors.
No functional role could yet be established for the glycosylated beta-subunit of the Na,K-ATPase. In this study, we describe the intracellular processing of the beta-subunit as a glycoprotein in toad bladder cells and the consequences of its structural perturbation with glycosylation inhibitors on the cellular expression of the alpha- and beta-subunits and on the structural and functional maturation of the enzyme. Controlled trypsinolysis of homogenates from pulse-labeled cells reveals that the beta-subunit is subjected to glycosylation-dependent structural rearrangements during its intracellular routing. Inhibition of correct terminal glycosylation of the beta-subunit with deoxynojirimycin or swainsonine has no effect on the trypsin sensitivity of the alpha-subunit, its ability to perform cation-dependent conformation changes or the cellular Na,K-ATPase activity. Acquisition of core-sugars is sufficient for the enzyme to assume its catalytic functions. On the other hand, complete inhibition of glycosylation with tunicamycin leads to a destabilization of both the beta- and the alpha-subunits as judged by their higher trypsin sensitivity. In addition, tunicamycin treatment results in a decrease of the amount of newly synthesized beta- and alpha-subunit indicating that a glycoprotein, possibly the beta-subunit itself, plays a role in the efficient accumulation of the alpha-subunit in the endoplasmic reticulum.
Zamofing
D
D
Institute of Pharmacology, University of Lausanne, Switzerland.
Rossier
B C
BC
Geering
K
K
D017485:1-Deoxynojirimycin; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D005944:Glucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D006023:Glycoproteins / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology; D006031:Glycosylation; D011233:Precipitin Tests; D011487:Protein Conformation / Q000187:drug effects; D011499:Protein Processing, Post-Translational; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology*; D017026:Swainsonine; D014357:Trypsin / Q000494:pharmacology; D014415:Tunicamycin / Q000494:pharmacology
D000470:Alkaloids; D006023:Glycoproteins; D014415:Tunicamycin; D017485:1-Deoxynojirimycin; D014357:Trypsin; D000254:Sodium-Potassium-Exchanging ATPase; D005944:Glucosamine; D017026:Swainsonine