Molecular analysis of ouabain-resistant mutants of the mouse lymphoma cell line L5178Y.
We have carried out molecular and biochemical analyses on several spontaneous and mutagen-induced ouabain-resistant mutants from the mouse lymphoma cell line L5178Y. Mutant cells were much more resistant than wild-type cells to the toxic effects of ouabain. The gross structures and copy numbers of two ouabain-resistant genes were unaltered in the mutants, as was the expression of the Na,K-ATPase gene. Uptake of 86Rb+ was more resistant to ouabain in the mutants than in wild-type cells. There was no evidence for an inducible uptake mechanism. The Na,K-ATPase activity in extracts of the mutants was, in all cases examined, more resistant than wild-type cells to the inhibitory action of 10(-3) M ouabain. At 10(-5) M ouabain, the Na,K-ATPase activity in three out of eleven mutants was actually more sensitive to ouabain than in wild-type cells. These data suggest that the ouabain-resistance of the mutants probably results from single base-changes in the alpha subunit of the Na,K-ATPase gene, rather than from amplification or overexpression of ouabain-resistance genes. The latter types of alterations have been described for ouabain-resistant lines derived in other laboratories.
Muriel
W J
WJ
MRC Cell Mutation Unit, University of Sussex, Falmer, Brighton, UK.
Cole
J
J
Lehmann
A R
AR
D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D002999:Clone Cells; D004351:Drug Resistance / Q000235:genetics; D007940:Leukemia L5178 / Q000235:genetics* / Q000378:metabolism; D007942:Leukemia, Experimental / Q000235:genetics*; D051379:Mice; D009154:Mutation; D009693:Nucleic Acid Hybridization; D010042:Ouabain / Q000494:pharmacology*; D011188:Potassium / Q000378:metabolism; D012414:Rubidium Radioisotopes; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics
D012414:Rubidium Radioisotopes; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium