Tissue-specific versus isoform-specific differences in cation activation kinetics of the Na,K-ATPase

The experiments described in this report reconcile some of the apparent differences in isoform-specific kinetics of the Na,K-ATPase reported in earlier studies. Thus, tissue-specific differences in Na+ and K+ activation kinetics of Na,K-ATPase activity of the same species (rat) were observed when the same isoform was assayed in different tissues or cells. In the case of alpha 1, alpha 1-transfected HeLa cell, rat kidney, and axolemma membranes were compared. For alpha 3, the ouabain-insensitive alpha 3*-transfected HeLa cell (cf.Jewell, E. A., and Lingrel, J.B. (1991) J. Biol. Chem. 266, 16925-16930), pineal gland, and axolemma (mainly alpha 3) membranes were compared. The order of apparent affinities for Na+ of alpha 1 pumps was axolemma approximate to rat alpha 1-transfected HeLa > kidney, and for K+, kidney approximate to HeLa > axolemma. For alpha 3, the order of apparent affinities for Na+ was pineal gland approximate to axolemma > alpha 3*-transfected HeLa, and for K+, alpha 3*-transfected HeLa > axolemma approximate to pineal gland. In addition, the differences in apparent affinities for Na+ of either kidney alpha 1 or HeLa alpha 3* as compared to the same isoform in other tissues were even greater when the K+ concentration was increased. A kinetic analysis of the apparent affinities for Na+ as a function of K+ concentration indicates that isoform-specific as well as tissue-specific differences are related to the apparent affinities for both Na+ and K+, the latter acting as a competitive inhibitor at cytoplasmic Na+ activation sites. Although the nature of the tissue-specific modulation of K+/Na+ antagonism remains unknown, an analysis of the nature of the beta isoform associated with alpha 1 or alpha 3 using isoform-specific immunoprecipitation indicates that the presence of distinct beta subunits does not account for differences of alpha 1 of kidney, axolemma, and HeLa, and of alpha 3 of axolemma and HeLa; in both instances beta 1 is the predominant beta isoform present or associated with either alpha 1 or alpha 3. However, a kinetic difference in K+/Na+ antagonism due to distinct beta s may apply to alpha 3 of axolemma (alpha 3 beta 1) and pineal gland (alpha 3 beta 2).