Parallel homodimer structures of the extracellular domains of the voltage-gated sodium channel β4 subunit explain its role in cell–cell adhesion

Abstract

Voltage-gated sodium channels (VGSCs) are transmembrane proteins required for the generation of action potentials in excitable cells and essential for propagating electrical impulses along nerve cells. VGSCs are complexes of a pore-forming α subunit and auxiliary β subunits, designated as β1/β1B–β4 (encoded by SCN1B–4B, respectively), which also function in cell–cell adhesion. We previously reported the structural basis for the trans homophilic interaction of the β4 subunit, which contributes to its adhesive function. Here, using crystallographic and biochemical analyses, we show that the β4 extracellular domains directly interact with each other in a parallel manner that involves an intermolecular disulfide bond between the unpaired Cys residues (Cys58) in the loop connecting strands B and C and intermolecular hydrophobic and hydrogen-bonding interactions of the N-terminal segments (Ser30-Val35). Under reducing conditions, an N-terminally deleted β4 mutant exhibited decreased cell adhesion compared with the wild type, indicating that the β4 cis dimer contributes to the trans homophilic interaction of β4 in cell–cell adhesion. Furthermore, this mutant exhibited increased association with the α subunit, indicating that the cis dimerization of β4 affects α–β4 complex formation. These observations provide the structural basis for the parallel dimer formation of β4 in VGSCs and reveal its mechanism in cell–cell adhesion.

Supramolecular clustering of the cardiac sodium channel Nav1.5 in HEK293F cells, with and without the auxiliary β3‐subunit

“…The β3‐subunit has also been shown to promote trans ‐cell adhesion, at least under conditions of high expression . Hence, the ability of β3‐subunits to homo‐trimerize may reflect a more general tendency of Nav channel β‐subunits to self‐assemble, at least under some in vivo conditions—for example, in the absence of Nav α‐subunits …”

“…19 Hence, the ability of β3subunits to homo-trimerize may reflect a more general tendency of Nav channel β-subunits to self-assemble, at least under some in vivo conditions-for example, in the absence of Nav α-subunits. 23 On neuronal cells and cardiomyocytes, Nav channels are known to be restricted into local micro-domain regions of the plasma membrane. 24 Furthermore, it has been reported that that these channels may functionally interact under pathological conditions.…”

Cell-Adhesion Properties of β-Subunits in the Regulation of Cardiomyocyte Sodium Channels

“…Firstly, the presence of an inter-subunit disulphide bond between the Cys58 residue on each Ig domain. Secondly, a reciprocal strand-swap interaction, in which the first seven N-terminal amino-acid residues from one Ig domain interact with residues on the partner Ig domain ( Figure 8 A) [ 79 ]. N-terminal strand-swapping is a known feature of several trans -mediated CAMs that contain Ig domains, including cadherins in the adherens junctions and desmogleins in the desmosomes [ 80 , 81 , 82 ].…”

Voltage-Gated Sodium Channel β Subunits and Their Related Diseases

“…Recent evidence shows that β4 Ig domains interact in a parallel manner involving a disulfide bond between cysteine 58 and hydrophobic and hydrogen bonding interactions between residues 30 through 35. Deletion of the β4 N-terminal domain led to decreased cell adhesion and increased association with the α subunit, revealing the importance of β4 cis dimerization (Shimizu et al 2017). …”