Mechanisms of noncovalent β subunit regulation of Na _V channel gating

Abstract

Voltage-gated Na _V channels are modulated by two different noncovalent accessory subunits: β1 and β3. Zhu et al. present data showing that β1 and β3 cause distinct effects on channel gating because they interact with Na _V channels at different locations. β3 regulates the voltage sensor in domain III, whereas β1 regulates the one in domain IV.

Abstract

Voltage-gated Na ⁺ (Na _V) channels comprise a macromolecular complex whose components tailor channel function. Key components are the non-covalently bound β1 and β3 subunits that regulate channel gating, expression, and pharmacology. Here, we probe the molecular basis of this regulation by applying voltage clamp fluorometry to measure how the β subunits affect the conformational dynamics of the cardiac Na _V channel (Na _V1.5) voltage-sensing domains (VSDs). The pore-forming Na _V1.5 α subunit contains four domains (DI–DIV), each with a VSD. Our results show that β1 regulates Na _V1.5 by modulating the DIV-VSD, whereas β3 alters channel kinetics mainly through DIII-VSD interaction. Introduction of a quenching tryptophan into the extracellular region of the β3 transmembrane segment inverted the DIII-VSD fluorescence. Additionally, a fluorophore tethered to β3 at the same position produced voltage-dependent fluorescence dynamics strongly resembling those of the DIII-VSD. Together, these results provide compelling evidence that β3 binds proximally to the DIII-VSD. Molecular-level differences in β1 and β3 interaction with the α subunit lead to distinct activation and inactivation recovery kinetics, significantly affecting Na _V channel regulation of cell excitability.