KCNJ2 mutation causes an adrenergic-dependent rectification abnormality with calcium sensitivity and ventricular arrhythmia

Abstract

Background KCNJ2 mutations are associated with a variety of inherited arrhythmia syndromes including CPVT3. Objective Detailed cellular and mechanistic characterization of the clinically recognized KCNJ2 mutation R67Q. Methods Kir2.1 current density was measured using the whole-cell voltage clamp technique from COS-1 cells transiently transfected with WT-Kir2.1 and/or R67Q-Kir2.1. Catecholamine activity was simulated with PKA stimulating cocktail exposure. Phosphorylation deficient mutants, S425N-Kir2.1 and S425N-Kir2.1/R67Q-S425N-Kir2.1, were used in a separate set of experiments. HA- or Myc-Tag-WT-Kir2.1 or HA-Tag-R67Q-Kir2.1 were used for confocal imaging. Results A 33 year old presented with a CPVT-like clinical phenotype and was found to have KCNJ2 missense mutation R67Q. Treatment with nadolol and flecainide resulted in complete suppression of arrhythmias and symptom resolution. Under baseline conditions, R67Q-Kir2.1 expressed alone did not produce IK1 while cells co-expressing WT-Kir2.1 and R67Q-Kir2.1 showed rectification index (RI) similar to WT-Kir2.1. After PKA stimulation, R67Q-Kir2.1/WT-Kir2.1 failed to increase peak outward current density; WT-Kir2.1 increased 46% (n=5) while R67Q-Kir2.1/WT-Kir2.1 decreased 6% (n=6), p=0.002. Rectification properties in R67Q-Kir2.1/WT-Kir2.1 demonstrated sensitivity to calcium with decreased RI in high-calcium pipette solution (RI 20.3 ± 4.1%) compared to low-calcium (RI 36.5 ± 5.7%) (p< 0.05). Immunostaining of WT-Kir2.1 and R67Q-Kir2.1 individually and together showed a normal membrane expression pattern and co-localization by Pearson’s correlation coefficient. Conclusion R67Q-Kir2.1 is associated with an adrenergic-dependent clinical and cellular phenotype with rectification abnormality enhanced by increased calcium. These findings are a significant advancement of our knowledge and understanding of phenotype-genotype relationship of arrhythmia syndromes related to KCNJ2 mutations.

Exercise training prevents ventricular tachycardia in CPVT1 due to reduced CaMKII-dependent arrhythmogenic Ca2+release

“…3 The very rare autosomal recessive variant CPVT2 results from mutations in the cardiac calsequestrin 2 gene, 4 while rare mutations responsible for CPVT3 were found in KCNJ2 leading to inward-rectifier potassium channel dysfunction. 5 RyR2 gain-of-function in CPVT1 disrupts normal control of excitation -contraction coupling, resulting in abnormally increased Ca 2+ leak from the sarcoplasmic reticulum (SR) in diastole. 6 Diastolic Ca 2+ leak originating from only one RyR2 cluster may lead to abnormal activation of neighbouring RyR2 clusters and thereby trigger regenerative, cell-wide Ca 2+ release, i.e.…”

Heterogeneity of clinical syndromes related to loss of function mutations in KCNJ2

“…Subsequent testing demonstrated exercise-induced arrhythmias that included salvos of polymorphic and bidirectional ventricular tachycardia. Both V227F-Kir2.1 and R67Q-Kir2.1 demonstrated adrenergic dependent loss of function in heterologous cells [28,29].…”

“…To date, two KCNJ2 mutations associated with CPVT3 have been characterized, V227F and R67Q, by our group [28,29]. The patients identified with CPVT3 were female and presented with exertion/ emotion related near-syncope and syncope.…”

“…It is clear that there has been considerable overlap between mutations that have been associated with ATS1 and CPVT3 [11,29]. However, supporting two clinical entities includes both the distinct clinical presentation and cellular physiology.…”

“…An N-terminus mutation that alters Kir2.1 sensitivity to PIP2 has been shown to exaggerate the inhibition of Kir2.1 by a divalent cation, magnesium [31]. Therefore, one hypothesis for adrenergic modulated channel function may be enhanced inhibition in the presence of increased cellular calcium [29].…”