Leukoencephalopathy-causing CLCN2 mutations are associated with impaired Cl- channel function and trafficking

Mutations in CLCN2 have been recently identified in patients suffering from a type of leukoencephalopathy involving intramyelinic oedema. Here, we characterised most of these mutations that reduce the function of the chloride channelClC-2 and impair its plasmamembrane (PM) expression. Detailed biochemical and electrophysiological analyses of the Ala500Val mutation revealed that defective gating and increased cellular and PM turnover contributed to defectiveA500V-ClC-2 functional expression. Co-expression of the adhesionmoleculeGlialCAM, which forms a tertiary complex with ClC-2 and megalencephalic leukoencephalopathy with subcortical cysts 1 (MLC1), rescued the functional expression of the mutant by modifying its gating properties. GlialCAM also restored the PM levels of the channel by impeding its turnover at the PM. This rescue required ClC-2 localisation to cell-cell junctions, since a GlialCAMmutant with compromised junctional localisation failed to rescue the impaired stability ofmutant ClC-2 at the PM. Wild-type, but not mutant, ClC-2 was also stabilised by MLC1 overexpression. We suggest that leukodystrophy-causing CLCN2mutations reduce the functional expression of ClC-2, which is partly counteracted by GlialCAM/MLC1-mediated increase in the gating and stability of the channel.