A Long QT Mutation Substitutes Cholesterol for Phosphatidylinositol-4,5-Bisphosphate in KCNQ1 Channel Regulation

INTRODUCTION: Phosphatidylinositol-4,5-bisphosphate (PIP(2)) is a cofactor necessary for the activity of KCNQ1 channels. Some Long QT mutations of KCNQ1, including R243H, R539W and R555C have been shown to decrease KCNQ1 interaction with PIP(2). A previous study suggested that R539W is paradoxically less sensitive to intracellular magnesium inhibition than the WT channel, despite a decreased interaction with PIP(2). In the present study, we confirm this peculiar behavior of R539W and suggest a molecular mechanism underlying it. METHODS AND RESULTS: COS-7 cells were transfected with WT or mutated KCNE1-KCNQ1 channel, and patch-clamp recordings were performed in giant-patch, permeabilized-patch or ruptured-patch configuration. Similar to other channels with a decreased PIP(2) affinity, we observed that the R243H and R555C mutations lead to an accelerated current rundown when membrane PIP(2) levels are decreasing. As opposed to R243H and R555C mutants, R539W is not more but rather less sensitive to PIP(2) decrease than the WT channel. A molecular model of a fragment of the KCNQ1 C-terminus and the membrane bilayer suggested that a potential novel interaction of R539W with cholesterol stabilizes the channel opening and hence prevents rundown upon PIP(2) depletion. We then carried out the same rundown experiments under cholesterol depletion and observed an accelerated R539W rundown that is consistent with this model. CONCLUSIONS: We show for the first time that a mutation may shift the channel interaction with PIP(2) to a preference for cholesterol. This de novo interaction wanes the sensitivity to PIP(2) variations, showing that a mutated channel with a decreased affinity to PIP(2) could paradoxically present a slowed current rundown compared to the WT channel. This suggests that caution is required when using measurements of current rundown as an indicator to compare WT and mutant channel PIP(2) sensitivity.