Ankyrin-G Coordinates Intercalated Disc Signaling Platform to Regulate Cardiac Excitability In Vivo

Abstract

Rationale Nav1.5 (SCN5A) is the primary cardiac voltage-gated Nav channel. Nav1.5 is critical for cardiac excitability and conduction, and human SCN5A mutations cause sinus node dysfunction, atrial fibrillation, conductional abnormalities, and ventricular arrhythmias. Further, defects in Nav1.5 regulation are linked with malignant arrhythmias associated with human heart failure. Consequently, therapies to target select Nav1.5 properties have remained at the forefront of cardiovascular medicine. However, despite years of investigation, the fundamental pathways governing Nav1.5 membrane targeting, assembly, and regulation are still largely undefined. Objective Define the in vivo mechanisms underlying Nav1.5 membrane regulation. Methods and Results Here, we define the molecular basis of a Nav channel regulatory platform in heart. Using new cardiac-selective ankyrin-G−/− mice (cKO), we report that ankyrin-G targets Nav1.5, and its regulatory protein, calcium/calmodulin-dependent kinase II (CaMKII) to the intercalated disc. Mechanistically, βIV-spectrin is requisite for ankyrin-dependent targeting of CaMKIIδ, however βIV-spectrin is not essential for ankyrin-G expression. Ankyrin-G cKO myocytes display decreased Nav1.5 expression/membrane localization, and reduced INa associated with pronounced bradycardia, conduction abnormalities, and ventricular arrhythmia in response to Nav channel antagonists. Moreover, we report that ankyrin-G links Nav channels with broader intercalated disc signaling/structural nodes, as ankyrin-G loss results in reorganization of plakophilin-2 and lethal arrhythmias in response to beta-adrenergic stimulation. Conclusions Our findings provide the first in vivo data for the molecular pathway required for intercalated disc Nav1.5 targeting/regulation in heart. Further, these new data identify the basis of an in vivo cellular platform critical for membrane recruitment and regulation of Nav1.5.

Ankyrin G Expression Regulates Apical Delivery of the Epithelial Sodium Channel (ENaC)

“…In neurons and cardiomyocytes, AnkG is essential for assembly of the axon initial segment (23)(24)(25) and the intercalated disk (26,27), respectively, and elimination of AnkG results in mislocalized voltage-gated Na ϩ and K ϩ channels (25,28,29). In the lungs and kidney, AnkG plays a role in lateral membrane biogenesis (30 -32) and has been shown to regulate endocytosis of E-cadherin (33).…”

Targeting the Sodium Channel Na V 1.5 to Specific Membrane Compartments of Cardiac Cells

“…This article by Makara et al 20 demonstrates the crucial role of ankyrin-G in vivo in the trafficking of the Na V 1.5 channel in the heart and reveals the molecular and functional consequences that lead to cardiac physiological phenotypes when this protein is deleted exclusively in the heart. The importance of Na V 1.5 regulation via phosphorylation by CaMKII has been studied previously 10,21 ; however, Makara et al 20 in this article elaborated on this knowledge and put it in the context of a functional macromolecular complex that is influential to only 1 subset of Na V 1.5 channels, the ID Na V 1.5 pool.…”

“…In this issue of Circulation Research, Makara et al 20 accomplished this goal by investigating the effects of deletion of the ankyrin-G gene on cardiac cellular, electric, and physiological function. Using a cardiac-specific knockout mouse for the ankyrin-G gene, they observed, through an elegant series of experiments, that loss of ankyrin-G leads to a decrease in Na V 1.5 expression, localization, and sodium current, specifically at…”

“…The importance of Na V 1.5 regulation via phosphorylation by CaMKII has been studied previously 10,21 ; however, Makara et al 20 in this article elaborated on this knowledge and put it in the context of a functional macromolecular complex that is influential to only 1 subset of Na V 1.5 channels, the ID Na V 1.5 pool. This leads us back to the interesting question of the functional differences among the different pools of Na V 1.5.…”

The multi-faceted aspects of the complex cardiac Nav1.5 protein in membrane function and pathophysiology

“…Instead, the pool of Na v 1.5 at the intercalated disks interacts, in vivo, with ankirin-G that is also linked with the desmosomal protein plakophilin-2 [37]. Importantly, also the kinetic properties of the sodium channel differed between the two Na v 1.5 pools: compared with the pool of Na v at the intercalated disk, the Na v 1.5 at the lateral membrane had a smaller current amplitude, a negative shift in the inactivation kinetic and a slower recovery from inactivation [38].…”