Towards systematic analysis of cardiac voltage-gated sodium channel beta subunit function

Abstract Voltage-gated sodium channels (NaVs) maintain the electrical cadence in cardiac muscle tissues by selectively controlling the rapid inward passage of sodium. The NaV complex is comprised of a 260-kDa pore- forming α-subunit (encoded primarily by NaV1.5 in heart) that partners with β -subunits (β1-β4) comprised of a single transmembrane segment and exofacial immunoglobulin (Ig) fold. These β-subunits belong to a larger family of β/MPZ proteins that includes other single pass Ig proteins such as MPZ(P0). Defects in sodium channel function resulting from inherited mutations in either the α or β subunits are established causes of human disease, and are associated with sudden infant death, atrial fibrillation, reperfusion and ischemia injury, arrhythmia in the failing heart, epilepsy, and a variety of pain-causing syndromes. Other forms of heart disease that develop later in life and that are exacerbated by obesity are also characterized by altered sodium channel activity, in particular the inability to quickly and completely inactivate during the course of the cardiac action potential. The β-subunits have been proposed to regulate essentially every aspect of the pore-forming α subunit; including protein complex trafficking and turn-over, voltage-dependent function, and pharmacology. However, the molecular bases for these wide-ranging effects are poorly resolved primarily because the ‘gold standard’ heterologous cells that are used for ion channel characterization and high-throughput screening exhibit near ubiquitous expression of β-subunits, and their near relatives. As such, the variability amongst expression systems has stymied systematic study of β-subunit function. This in turn has prevented translational studies to examine the effect of various β subunit disease-associated mutants as well as efforts to identify drugs that may specifically modify specific α/β NaV complexes. We recently generated a CRISPR-modified human haploid cell line that lacks multiple members of the β/MPZ family including β1-β4. Electrophysiological experiments with these cells have revealed new emergent properties of NaV1.5 in the absence and presence of β subunits. These data provide the proof-of-concept that mammalian cells lacking the β/MPZ family will provide a powerful and needed way to specifically study many aspects of particular NaV α/β complexes, allowing better fundamental structure/function studies, better understanding of how disease-causing mutations in both α and β subunits cause pathology, and more precise tools for drug screening and drug safety profile testing.