Ion channel function is fundamental to the existence of life. In metazoans, the coordinate activities of voltagegated Na + channels underlie cellular excitability and control neuronal communication, cardiac excitationcontraction coupling, and skeletal muscle function. However, despite decades of research and linkage of Na + channel dysfunction with arrhythmia, epilepsy, and myotonia, little progress has been made toward understanding the fundamental processes that regulate this family of proteins. Here, we have identified β IV -spectrin as a multifunctional regulatory platform for Na + channels in mice. We found that β IV -spectrin targeted critical structural and regulatory proteins to excitable membranes in the heart and brain. Animal models harboring mutant β IV -spectrin alleles displayed aberrant cellular excitability and whole animal physiology. Moreover, we identified a regulatory mechanism for Na + channels, via direct phosphorylation by β IV -spectrin-targeted calcium/calmodulin-dependent kinase II (CaMKII). Collectively, our data define an unexpected but indispensable molecular platform that determines membrane excitability in the mouse heart and brain.

Background The cardiac cytoskeleton plays key roles in maintaining myocyte structural integrity in health and disease. In fact, human mutations in cardiac cytoskeletal elements are tightly linked with cardiac pathologies including myopathies, aortopathies, and dystrophies. Conversely, the link between cytoskeletal protein dysfunction in cardiac electrical activity is not well understood, and often overlooked in the cardiac arrhythmia field. Methods and Results Here, we uncover a new mechanism for the regulation of cardiac membrane excitability. We report that βII spectrin, an actin-associated molecule, is essential for the post-translational targeting and localization of critical membrane proteins in heart. βII spectrin recruits ankyrin-B to the cardiac dyad, and a novel human mutation in the ankyrin-B gene disrupts the ankyrin-B/βII spectrin interaction leading to severe human arrhythmia phenotypes. Mice lacking cardiac βII spectrin display lethal arrhythmias, aberrant electrical and calcium handling phenotypes, and abnormal expression/localization of cardiac membrane proteins. Mechanistically, βII spectrin regulates the localization of cytoskeletal and plasma membrane/sarcoplasmic reticulum protein complexes that include the Na/Ca exchanger, RyR2, ankyrin-B, actin, and αII spectrin. Finally, we observe accelerated heart failure phenotypes in βII spectrin-deficient mice. Conclusions Our findings identify βII spectrin as critical for normal myocyte electrical activity, link this molecule to human disease, and provide new insight into the mechanisms underlying cardiac myocyte biology.

The coordinated sorting of ion channels to specific plasma membrane domains is necessary for excitable cell physiology. KATP channels, assembled from pore-forming (Kir6.x) and regulatory sulfonylurea receptor subunits, are critical electrical transducers of the metabolic state of excitable tissues, including skeletal and smooth muscle, heart, brain, kidney, and pancreas. Here we show that the C-terminal domain of Kir6.2 contains a motif conferring membrane targeting in primary excitable cells. Kir6.2 lacking this motif displays aberrant channel targeting due to loss of association with the membrane adapter ankyrin-B (AnkB). Moreover, we demonstrate that this Kir6.2 C-terminal AnkB-binding motif (ABM) serves a dual role in KATP channel trafficking and membrane metabolic regulation and dysfunction in these pathways results in human excitable cell disease. Thus, the KATP channel ABM serves as a previously unrecognized bifunctional touch-point for grading KATP channel gating and membrane targeting and may play a fundamental role in controlling excitable cell metabolic regulation.K ATP channels are critical electrical transducers of the metabolic state of excitable tissues including skeletal and smooth muscle, heart, brain, kidney, and pancreas (1). Mechanistically, decreased metabolism opens K ATP channels, resulting in K ϩ efflux, membrane hyperpolarization, and suppression of action potential formation (1). Conversely, increased metabolism closes K ATP channels, resulting in membrane depolarization, stimulation of electrical activity (2), and subsequent triggering of diverse cellular responses, such as release of hormones and neurotransmitters, or muscle contraction.Given such critical roles in the regulation of electrical excitability in many cell types, it is not surprising that K ATP channel dysfunction results in disease. Human mutations in K ATP channel genes are associated with neonatal diabetes and hyperinsulinemia (3), epilepsy (4), and dilated cardiomyopathy (5). However, despite the clear importance of K ATP function for normal vertebrate physiology, little is resolved regarding the mechanisms responsible for K ATP channel membrane targeting and/or membrane organization.Here we identify a critical motif in the Kir6.2 C-terminal domain that is essential for normal Kir6.2 membrane targeting. We demonstrate that the Kir6.2 C-terminal motif interacts with the cytoskeletal adapter ankyrin-B (AnkB) and that Kir6.2 displays aberrant membrane trafficking when the motif is disrupted or in cells lacking ankyrin-B expression. We demonstrate that the Kir6.2 C-terminal motif displays an unexpected secondary role in Kir6.2 membrane function (K ATP channel activity) by altering K ATP channel ATP sensitivity. Finally, we demonstrate that a human neonatal diabetes disease mutation located in the Kir6.2 C-terminal motif results in a complex ␤ cell phenotype, likely due to the dual role of the C-terminal motif in both Kir6.2 targeting and K ATP channel membrane activity. Together, our findings define a pathway for K ATP ch...

Rationale: Cardiac membrane excitability is tightly regulated by an integrated network of membrane-associated ion channels, transporters, receptors, and signaling molecules.