Extracellular Kir2.1 C122Y Mutant Upsets Kir2.1-PIP2 Bonds and Is Arrhythmogenic in Andersen-Tawil Syndrome.

BACKGROUND: Andersen-Tawil syndrome type 1 is a rare heritable disease caused by mutations in the gene coding the strong inwardly rectifying K+ channel Kir2.1. The extracellular Cys (cysteine)122 -to-Cys154 disulfide bond in the channel structure is crucial for proper folding but has not been associated with correct channel function at the membrane. We evaluated whether a human mutation at the Cys122 -to-Cys154 disulfide bridge leads to Kir2.1 channel dysfunction and arrhythmias by reorganizing the overall Kir2.1 channel structure and destabilizing its open state.

METHODS AND RESULTS: We identified a Kir2.1 loss-of-function Cys122 mutation (c.366 A>T; p.Cys122Tyr) in an Andersen-Tawil syndrome type 1 family. We generated a cardiac-specific mouse model expressing the Kir2.1C122Y that recapitulated the abnormal ECG features of Andersen-Tawil syndrome type 1 independently of sex, including corrected QT prolongation, conduction defects, and increased arrhythmia susceptibility. Isolated Kir2.1C122Y cardiomyocytes showed significantly reduced inwardly rectifier K+ (IK1 ) and inward Na+ (INa ) current densities independently of normal trafficking to and localization at the sarcolemma and the sarcoplasmic reticulum. However, molecular dynamics predicted that the C122Y mutation provoked a conformational change over the 2000-ns simulation, characterized by a greater loss of hydrogen bonds between Kir2.1 and phosphatidylinositol 4,5-bisphosphate than WT. Therefore, the phosphatidylinositol 4,5-bisphosphate-binding pocket was destabilized, resulting in a lower conductance state compared with WT. Accordingly, on inside-out patch clamping, the C122Y mutation significantly blunted Kir2.1 sensitivity to increasing phosphatidylinositol 4,5-bisphosphate concentrations. In addition, the Kir2.1C122Y mutation resulted in channelosome degradation, demonstrating temporal instability of both Kir2.1 and NaV 1.5 proteins.

CONCLUSIONS: The extracellular Cys122 -to-Cys154 disulfide bond in the tridimensional Kir2.1 channel structure is essential for the channel function. We demonstrate that breaking disulfide bonds in the extracellular domain disrupts phosphatidylinositol 4,5-bisphosphate-dependent regulation, leading to channel dysfunction and defects in Kir2.1 energetic stability. The mutation also alters functional expression of the NaV 1.5 channel and ultimately leads to conduction disturbances and life-threatening arrhythmia characteristic of Andersen-Tawil syndrome type 1.