Relationship between Intracellular Na+ Concentration and Reduced Na+ Affinity in Na+,K+-ATPase Mutants Causing Neurological Disease

The neurological disorders familial hemiplegic migraine type 2 (FHM2), alternating hemiplegia of childhood (AHC), and rapid-onset dystonia parkinsonism (RDP) are caused by mutations of Na⁺,K⁺-ATPase α₂ and α₃ isoforms, expressed in glial and neuronal cells, respectively. Although these disorders are distinct, they overlap in phenotypical presentation. Two Na⁺,K⁺-ATPase mutations, extending the C terminus by either 28 residues (“+28” mutation) or an extra tyrosine (“+Y”), are associated with FHM2 and RDP, respectively. We describe here functional consequences of these and other neurological disease mutations as well as an extension of the C terminus only by a single alanine. The dependence of the mutational effects on the specific α isoform in which the mutation is introduced was furthermore studied. At the cellular level we have characterized the C-terminal extension mutants and other mutants, addressing the question to what extent they cause a change of the intracellular Na⁺ and K⁺ concentrations (Na⁺]_i and K⁺]_i) in COS cells. C-terminal extension mutants generally showed dramatically reduced Na⁺ affinity without disturbance of K⁺ binding, as did other RDP mutants. No phosphorylation from ATP was observed for the +28 mutation of α₂ despite a high expression level. A significant rise of Na⁺]_i and reduction of K⁺]_i was detected in cells expressing mutants with reduced Na⁺ affinity and did not require a concomitant reduction of the maximal catalytic turnover rate or expression level. Moreover, two mutations that increase Na⁺ affinity were found to reduce Na⁺]_i. It is concluded that the Na⁺ affinity of the Na⁺,K⁺-ATPase is an important determinant of Na⁺]_i.