Transport of choline by plasma membrane vesicles from lung-derived epithelial cells.
A549 cells, a lung epithelium-derived cell line, were used as a model system to study choline transport by granular pneumocytes. Intact cells accumulated free choline against a concentration gradient by a low-affinity transport system with kinetic characteristics similar to that previously described for granular pneumocytes (Am. J. Respir. Cell Mol. Biol. 1: 455, 1989). Membrane vesicles prepared from these cells showed a 10-fold enrichment in plasma membrane marker enzymes with a vesicular H2O space of 5.7 +/- 0.05 (SE) microliters/mg protein. Vesicles showed a time- and concentration-dependent uptake of free [3H]choline in Na(+)-free medium. With 5 microM choline, choline uptake reached an apparent steady-state concentration gradient (inside/outside) of 50. 3H that was membrane associated ("bound" choline) represented approximately 5% of total uptake. In the presence of an initial gradient of NaCl, choline uptake showed an overshoot with a plateau value similar to Na(+)-free conditions; a similar effect was observed for plasma membrane vesicles from rat lung type 2 epithelial cells. The steady-state uptake of choline was inhibited at low pH (6.5) and by the presence of valinomycin or carbonyl cyanide p-tri-fluoromethoxyphenylhydrazone and was abolished when both were present. These results show that plasma membrane vesicles from A549 cells accumulate choline by binding to the membranes and by Na(+)-dependent and -independent transport mechanisms, the latter apparently reflecting a transmembrane proton gradient.
Fisher
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Institute for Environmental Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
Dodia
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Chander
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Kleinzeller
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D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002794:Choline / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors / Q000493:pharmacokinetics*; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism / Q000648:ultrastructure; D008168:Lung / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D014634:Valinomycin / Q000494:pharmacology
D014634:Valinomycin; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D002794:Choline