Binding Between a Highly Conserved Na,K‐ATPase Domain and the InsP3R Protects from Serum‐Deprivation Triggered Apoptosis

Mounting evidence suggests that the ion pump, Na,K‐ATPase, can, in the presence of ouabain, act as a signal transducer. A prominent binding‐motif, linking the Na,K‐ATPase to intracellular signaling effectors, has however not yet been identified. Here we report that the N‐terminal tail of the Na,K‐ATPase catalytic α‐subunit (αNT‐t) binds directly to the N‐terminus of the inositol 1,4,5‐trisphosphate receptor (InsP 3 R). Three amino acid residues, LKK, conserved in most species and most α‐isoforms, are essential for the binding to occur. In wild‐type cells, low concentrations of ouabain trigger low frequency calcium oscillations that activate NF‐κB and protect from apoptosis. Over‐expression of αNT‐t in COS‐7 cells resulted in a dramatic reduction of the number of cells responding with calcium oscillations (38.9% ± 8.9), compared to control cells (63.2%± 4.7). In contrast, cells over‐expressing αNT‐t LKK responded in the same way as control cells (73.5% ± 7.9 and 73.6% ± 4.8 respectively). Primary culture of rat proximal tubule cells over‐expressing αNT‐t LKK displayed NF‐κB activation and protection from apoptosis when treated with ouabain. The activation of NF‐κB and the protection from apoptosis was abolished in cells over‐expressing αNT‐t. Thus we demonstrate a binding‐motif linking Na,K‐ATPase to InsP3R that may be of vital importance for cell signaling.