J. Biol. Chem., Vol. 261, Issue 21, 9779-9786, 07, 1986
Reduced ouabain inhibition of Na,K-activated adenosine triphosphatase in cultured cell recipients of the ouabain-resistance gene
TA Pressley and IS Edelman
Using sequential DNA-mediated gene transfer, a mouse DNA sequence (ouaR)
that confers resistance to the cytotoxic effects of ouabain in cultured
cells was recently isolated. To determine the basis of this resistance, we
examined the ouabain-resistant phenotype of cells from each stage of the
transfers that led to isolation of the ouaR gene, as well as the recipient
of the isolated gene. Membranes prepared from the original DNA donor, the
two intermediate stages of the sequential gene transfer, and the final
isolated gene recipient contained at least two functionally distinct forms
of the Na,K-ATPase. In addition to a form indistinguishable in ouabain
affinity from that in the parental lines, these ouabain-resistant cells
contained a form characterized by a low affinity for the glycoside. That
the low-affinity form contributed to ouabain resistance was suggested by
the correlation between its relative abundance and the ability to limit the
increase in intracellular Na+ content when exposed to ouabain. Maintenance
of the final recipient of the isolated ouaR gene for 24 h in 10 microM
ouabain had no effect on the specific activity of the resistant form of the
Na,K-ATPase, implying that the low-affinity form was not uniquely inducible
by ouabain. These results suggest that the ouabain-resistant phenotype
conferred by ouaR is attributable to expression of a Na,K- ATPase with a
low affinity for the glycoside.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?