Digitalis-induced cell signaling by the sodium pump: On the relation of Src to Na+/K+-ATPase

Abstract

In addition to performing its essential transport function, the sodium pump also activates multiple cell signaling pathways in response to digitalis drugs such as ouabain. Based mainly on cell-free studies with mixtures of purified Src kinase and Na(+)/K(+)-ATPase, a well-advocated hypothesis on how ouabain initiates the activation of signaling pathways is that there is a preexisting physiological complex of inactive Src bound to the α-subunit of Na(+)/K(+)-ATPase, and that ouabain binding to this subunit disrupts the bound Src and activates it. Because of the published disagreements of the results of such cell-free experiments of two other laboratories, our aim was to attempt the resolution of these discrepancies. We reexamined the effects of ouabain, vanadate, and oligomycin on mixtures of Src, Na(+)/K(+)-ATPase, Mg(2+), and ATP as specified in prior studies; and assayed for Src-418 autophosphorylation as the measure of Src activation. In contrast to the findings of the proponents of the above hypothesis, our results showed similar effects of the three inhibitors of Na(+)/K(+)-ATPase; indicating that Src activation in such experiments is primarily due to the ATP-sparing effect of the ATPase inhibitor on the mixture of two enzymes competing for ATP. We conclude that there is no solid evidence for direct molecular interaction of Src with Na(+)/K(+)-ATPase under physiological conditions.

Do Src Kinase and Caveolin Interact Directly with Na,K-ATPase?

“…However, subsequently, it was reported that Src kinase is activated either by ouabain or by vanadate with the conclusion that Src kinase activation is dependent on the energetic state of Na,K-ATPase, in particular the ATP/ADP ratio (18). Similarly, it was reported recently that all three inhibitors (ouabain, vanadate, and oligomycin) activate Src Tyr-418 autophosphorylation, and it was concluded that Src kinase activation is due primarily to the ATP-sparing effects of the ATPase inhibitors (16). Src kinase activation was also reportedly induced by stabilization of purified renal Na,KATPase using either ouabain or BeF and AlF (11).…”

“…More recently, other studies have challenged the model of direct interaction between Src kinase and Na,K-ATPase (14,16,17,18) (see "Discussion"). Although several observations raise doubts about the direct interaction model, they do not rule out indirect interactions mediated by other proteins that might be involved in the signaling events.…”

Protein Carbonylation of an Amino Acid Residue of the Na/K‐ATPase α1 Subunit Determines Na/K‐ATPase Signaling and Sodium Transport in Renal Proximal Tubular Cells

“…In this model, the Na/K‐ATPase α1 subunit provides the ligand binding site and the associated c‐Src functions as the kinase moiety. It has also been proposed that c‐Src activation is primarily due to an ATP‐sparing effect based on a cell‐free system . While the Na/K‐ATPase inhibitors (vanadate and oligomycin) and ATP/ADP ratio regulate c‐Src activation, a possible interaction between the α1 and c‐Src was not addressed …”

Targeting Na/K-ATPase Signaling: A New Approach to Control Oxidative Stress

“…In this model, the Na/K-ATPase α1 subunit provides the ligand binding site, and associated c-Src provides the kinase moiety. A second model is that c-Src activation is primarily a consequence of an ATP-sparing effect (observed in a cell-free system) [45, 46]. A third model proposes that c-Src transiently interacts with a complex formed between the Na/K-ATPase α1 subunit and caveolin-1[47].…”