Accumulation of UDP-GlcNAc into intracellular vesicles and occurrence of a carrier-mediated transport. Study with plasma-membrane-permeabilized mouse thymocytes.
Metabolic labelling of mouse thymocytes with radioactive mannose or glucosamine leads to the formation of labelled GDP-Man and UDP-GlcNAc. Using isotonic ammonium chloride treatment which renders the plasma membrane of thymocytes permeable to sugar nucleotides, we demonstrate that, in contrast to GDP-Man, a pool of UDP-GlcNAc remains associated with the cells after plasma membrane permeabilization. These observations are confirmed in experiments in which permeabilized thymocytes are incubated with exogenous labelled GDP-Man and UDP-GlcNAc, and we show that only UDP-GlcNAc is accumulated into sealed intracellular vesicles. This accumulation is a saturable process which can be inhibited by UDP, demonstrating the occurrence of a specific carrier. This transport mechanism can be blocked by covalent attachment of a non-permeant inhibitor UDP-dialdehyde without affecting the N-acetylglucosaminyltransferase itself. The fact that this carrier-mediated transport is not inhibited by tunicamycin indicates that this translocation process of UDP-GlcNAc does not involve lipid intermediates.
R
R
Cecchelli
R
R
Cacan
A
A
Verbert
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000502:physiology; D002463:Cell Membrane Permeability; D006155:Guanosine Diphosphate Mannose / Q000378:metabolism; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism*; D051379:Mice; D013950:Thymus Gland / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism*; D014539:Uridine Diphosphate Sugars / Q000378:metabolism*
D002352:Carrier Proteins; D014539:Uridine Diphosphate Sugars; D014415:Tunicamycin; D006155:Guanosine Diphosphate Mannose; D014537:Uridine Diphosphate N-Acetylglucosamine