Cardiac Kir2.1 and NaV1.5 Channels Traffic Together to the Sarcolemma to Control Excitability.
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2018
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Abstract
Rationale: In cardiomyocytes, NaV1.5 and Kir2.1 channels interact dynamically as part of membrane bound
macromolecular complexes.
Objective: The objective of this study was to test whether NaV1.5 and Kir2.1 preassemble during early forward
trafficking and travel together to common membrane microdomains.
Methods and Results: In patch-clamp experiments, coexpression of trafficking-deficient mutants Kir2.1Δ314-315 or
Kir2.1R44A/R46A with wild-type (WT) NaV1.5WT in heterologous cells reduced inward sodium current compared with
NaV1.5WT alone or coexpressed with Kir2.1WT. In cell surface biotinylation experiments, expression of Kir2.1Δ314-315
reduced NaV1.5 channel surface expression. Glycosylation analysis suggested that NaV1.5WT and Kir2.1WT channels
associate early in their biosynthetic pathway, and fluorescence recovery after photobleaching experiments
demonstrated that coexpression with Kir2.1 increased cytoplasmic mobility of NaV1.5WT, and vice versa, whereas
coexpression with Kir2.1Δ314-315 reduced mobility of both channels. Viral gene transfer of Kir2.1Δ314-315 in adult rat
ventricular myocytes and human induced pluripotent stem cell–derived cardiomyocytes reduced inward rectifier
potassium current and inward sodium current, maximum diastolic potential and action potential depolarization
rate, and increased action potential duration. On immunostaining, the AP1 (adaptor protein complex 1) colocalized
with NaV1.5WT and Kir2.1WT within areas corresponding to t-tubules and intercalated discs. Like Kir2.1WT, NaV1.5WT
coimmunoprecipitated with AP1. Site-directed mutagenesis revealed that NaV1.5WT channels interact with AP1
through the NaV1.5Y1810 residue, suggesting that, like for Kir2.1WT, AP1 can mark NaV1.5 channels for incorporation
into clathrin-coated vesicles at the trans-Golgi. Silencing the AP1 ϒ-adaptin subunit in human induced pluripotent
stem cell–derived cardiomyocytes reduced inward rectifier potassium current, inward sodium current, and
maximum diastolic potential and impaired rate-dependent action potential duration adaptation.
Conclusions: The NaV1.5-Kir2.1 macromolecular complex pre-assembles early in the forward trafficking pathway.
Therefore, disruption of Kir2.1 trafficking in cardiomyocytes affects trafficking of NaV1.5, which may have
important implications in the mechanisms of arrhythmias in inheritable cardiac diseases. (Circ Res. 2018;122:1501-
1516. DOI: 10.1161/CIRCRESAHA.117.311872.)