Coupling of the Na+/K+-ATPase to Ankyrin B controls Na+/Ca2+ exchanger activity in cardiomyocytes.

Abstract

Ankyrin B (AnkB) is an adaptor protein that assembles Na+/K+-ATPase (NKA) and Na+/Ca2+ exchanger (NCX) in the AnkB macromolecular complex. Loss-of-function mutations in AnkB cause the AnkB syndrome in humans, characterized by ventricular arrhythmias and sudden cardiac death. It is unclear to what extent NKA binding to AnkB allows regulation of local Na+ and Ca2+ domains and hence NCX activity. To investigate the role of NKA binding to AnkB in cardiomyocytes, we synthesized a disruptor peptide (MAB peptide) and its AnkB binding ability was verified by pulldown experiments. As opposed to control, the correlation between NKA and NCX currents was abolished in adult rat ventricular myocytes dialyzed with MAB peptide, as well as in cardiomyocytes from AnkB+/- mice. Disruption of NKA from AnkB (with MAB peptide) increased NCX-sensed cytosolic Na+ concentration, reduced Ca2+ extrusion through NCX, and increased frequency of Ca2+ sparks and Ca2+ waves without concomitant increase in Ca2+ transient amplitude or SR Ca2+ load, suggesting an effect in local Ca2+ domains. Selective inhibition of the NKAα2 isoform abolished both the correlation between NKA and NCX currents and the increased rate of Ca2+ sparks and waves following NKA/AnkB disruption, suggesting that an AnkB/NKAα2/NCX domain controls Ca2+ fluxes in cardiomyocytes. NKA binding to AnkB allows ion regulation in a local domain, and acute disruption of the NKA/AnkB interaction using disruptor peptides lead to increased rate of Ca2+ sparks and waves. The functional effects were mediated through the NKAα2 isoform. Disruption of the AnkB/NKA/NCX domain could be an important pathophysiological mechanism in the AnkB syndrome.