Induction of system A amino acid transport through long-term treatment with ouabain: correlation with increased (Na+/K+)-ATPase activity.
Mouse embryo fibroblast cells (C3H-10T1/2) and the methylcholanthrene-transformed derivative (MCA-10T1/2) were treated with basal modified Eagle's medium (BME) containing 10% fetal bovine serum and varying concentrations of ouabain ranging from 0.05 mM to 0.7 mM for 16 h in culture. After replacing the ouabain-containing medium with Earl's balanced salts solution, System A amino acid transport activity increased from approximately 40 to 500 pmol AIB accumulated.mg protein-1.min-1 in the C3H-10T1/2 cells and from approximately 300 to 700 pmol AIB accumulated.mg protein-1.min-1 in the MCA-10T1/2 cells. The (Na+/K+)-ATPase pump activity also increased from approximately 12 to 46 nmol Rb+ accumulated.mg protein-1.min-1 in the normal cells and from approximately 20 to 42 nmol Rb+ accumulated.mg protein-1.min-1 in the transformed cells. System A and the (Na+/K+)ATPase activity were maximally increased at approximately 0.4-0.6 mM ouabain in the normal cells in contrast to the transformed cells which were maximally stimulated at a concentration of approximately 0.2 mM ouabain. This treatment with ouabain increased the [Na+]i/[K+]i as measured by atomic absorption spectroscopy, and thereby decreased the Na+ and K+ electrochemical gradients. Our data show that the internal ion gradients inverted at a lower concentration of ouabain in the transformed cells compared to the normal cells. The ouabain-induced increase in pump and System A activity shown here was used as a tool to further investigate the coordinated ion transport regulation in the control of cell growth.
Schenerman
M A
MA
Section of Biochemistry, Molecular, and Cell Biology, Cornell University, Ithaca, New York 14853.
Leister
K J
KJ
Trachtenberg
D K
DK
Racker
E
E
D000596:Amino Acids / Q000378:metabolism*; D000621:Aminoisobutyric Acids / Q000378:metabolism; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002478:Cells, Cultured; D007700:Kinetics; D051379:Mice; D010042:Ouabain / Q000494:pharmacology*; D011188:Potassium / Q000378:metabolism; D012413:Rubidium / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D000596:Amino Acids; D000621:Aminoisobutyric Acids; C100049:2-aminoisobutyric acid; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium; D011188:Potassium