Functional characterization of carrier-mediated transport of uridine diphosphate N-acetylglucosamine across the endoplasmic reticulum membrane.
Uptake and metabolism of uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) by rough-endoplasmic-reticulum (rER)-derived vesicles was studied. Analysis of the molecular species, double-label experiments, cis-inhibition and trans-stimulation experiments revealed that uptake represented entry of intact UDP-GlcNAc into the microsomal lumen. The amount of vesicle-associated label at equilibrium uptake was directly proportional to the volume of the intravesicular space, and permeabilized microsomes were unable to retain UDP-GlcNAc. These findings indicate that uptake constituted effective translocation from the medium into the lumen of the vesicles. The microsomal uptake of UDP-GlcNAc met the criteria of bidirectional, carrier-mediated translocation. Transport was time and temperature dependent, saturable, selective, capable of trans-stimulation and operational against a concentration gradient. Uptake studies performed in membrane preparations that were highly enriched in either rER, smooth ER, or Golgi revealed that UDP-GlcNAc was taken up by the ER and by the Golgi apparatus.
Bossuyt
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Department of Molecular Cell Biology, Catholic University of Leuven, Belgium.
Blanckaert
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D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D001693:Biological Transport, Active / Q000187:drug effects; D002352:Carrier Proteins / Q000378:metabolism*; D002451:Cell Compartmentation; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D006460:Hemolysin Proteins / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000031:analogs & derivatives / Q000378:metabolism*
D001427:Bacterial Toxins; D002352:Carrier Proteins; D006460:Hemolysin Proteins; C021091:staphylococcal alpha-toxin; D014537:Uridine Diphosphate N-Acetylglucosamine