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Na+/K+-ATPase; sproradic and familial hemiplegic migraine; pump currents; voltage dependence; surface expression; protein stability; Xenopus oocytes; HEK 293FT cells
Abstract:
Mutations in three different genes have been implicated in familial hemiplegic migraine (FHM), two of them code for neuronal voltage-gated cation channels, CACNA1A and SCN1A, while the third encodes ATP1A2, the α 2-isoform of the Na+/K+-ATPase's catalytic subunit, thus classifying FHM as an ion channel / ion transporter disorder. The Na+/K+-ATPase maintains the physiological gradients for Na+ and K+ ions and is therefore critical for the activity of ion channels and transporters involved in neurotransmitter uptake or Ca2+ signaling. Diverse functional abnormalities have been identified for disease-linked ATP1A2 mutations, which reach far beyond simple loss-of-function. We have shown recently that ATP1A2 mutations frequently lead to changes in the enzyme's voltage-dependent properties, kinetics, or apparent cation affinities. Here, we present functional data on a so far uncharacterized set of ATP1A2 mutations (G301R, R908Q, and P979L) upon expression in Xenopus oocytes and HEK293FT cells, and provide evidence for a novel pathophysiological mechanism. Whereas the G301R mutant was inactive, no functional changes were observed for mutants R908Q and P979L in the oocyte expression system. However, the R908Q mutant was less effectively expressed in the plasma membrane of oocytes, making it the first missense mutation to result in defective plasma membrane targeting. Notably, the P979L mutant exhibited the same cellular expression profile as the wild-type protein, both in Xenopus oocytes and in transfected HEK293FT cells grown at 28{degree sign}C, but much less P979L protein was found upon cell growth at 37{degree sign}C, showing for the first time that temperature-sensitive effects on protein stability can underlie ATP1A2 loss-of-function.
