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 apparen t differences in isoform-specific kinetics of the Na,K-ATPase reported in earlier studies. Thus, tissue-specific differences in Na+ and K+ a ctivation kinetics of Na,K-ATPase activity of the same species (rat) w ere observed when the same isoform was assayed in different tissues or cells. In the case of alpha 1, alpha 1-transfected HeLa cell, rat kid ney, and axolemma membranes were compared. For alpha 3, the ouabain-in sensitive alpha 3-transfected HeLa cell (cf.Jewell, E. A., and Lingre l, J.B. (1991) J. Biol. Chem. 266, 16925-16930), pineal gland, and axo lemma (mainly alpha 3) membranes were compared. The order of apparent affinities for Na+ of alpha 1 pumps was axolemma approximate to rat al pha 1-transfected HeLa > kidney, and for K+, kidney approximate to HeL a > axolemma. For alpha 3, the order of apparent affinities for Na+ wa s pineal gland approximate to axolemma > alpha 3-transfected HeLa, an d for K+, alpha 3-transfected HeLa > axolemma approximate to pineal g land. In addition, the differences in apparent affinities for Na+ of e ither kidney alpha 1 or HeLa alpha 3 as compared to the same isoform in other tissues were even greater when the K+ concentration was incre ased. A kinetic analysis of the apparent affinities for Na+ as a funct ion of K+ concentration indicates that isoform-specific as well as tis sue-specific differences are related to the apparent affinities for bo th Na+ and K+, the latter acting as a competitive inhibitor at cytopla smic Na+ activation sites. Although the nature of the tissue-specific modulation of K+/Na+ antagonism remains unknown, an analysis of the na ture of the beta isoform associated with alpha 1 or alpha 3 using isof orm-specific immunoprecipitation indicates that the presence of distin ct 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 ins tances beta 1 is the predominant beta isoform present or associated wi th either alpha 1 or alpha 3. However, a kinetic difference in K+/Naantagonism due to distinct beta s may apply to alpha 3 of axolemma (al pha 3 beta 1) and pineal gland (alpha 3 beta 2).