AJP - Cell Physiology, Vol 263, Issue 6 C1250-C1257, Copyright © 1992 by American Physiological Society
Transport of choline by plasma membrane vesicles from lung-derived epithelial cells
A. B. Fisher, C. Dodia, A. Chander and A. Kleinzeller
Institute for Environmental Medicine, University of Pennsylvania School of Medicine, Philadelphia 19104.
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.
