Sodium Channel β Subunits Mediate Homophilic Cell Adhesion and Recruit Ankyrin to Points of Cell-Cell Contact

Abstract

Sodium channels isolated from mammalian brain are composed of ␣, ␤1, and ␤2 subunits. The auxiliary ␤ subunits do not form the ion conducting pore, yet play important roles in channel modulation and plasma membrane expression. ␤1 and ␤2 are transmembrane proteins with one extracellular V-set immunoglobulin (Ig) protein domain. It has been shown recently that ␤1 and ␤2 interact with the extracellular matrix proteins tenascin-C and tenascin-R. In the present study we show that rat brain ␤1 and ␤2, but not ␣IIA, subunits interact in a trans-homophilic fashion, resulting in recruitment of the cytoskeletal protein ankyrin to sites of cell-cell contact in transfected Drosophila S2 cells. Whereas ␣IIA subunits expressed alone do not cause cellular aggregation, ␤ subunits co-expressed with ␣IIA retain the ability to adhere and recruit ankyrin. Truncated ␤ subunits lacking cytoplasmic domains interact homophilically to produce cell aggregation but do not recruit ankyrin. Thus, the cytoplasmic domains of ␤1 and ␤2 are required for cytoskeletal interactions. It is hypothesized that sodium channel ␤ subunits serve as a critical communication link between the extracellular and intracellular environments of the neuron and may play a role in sodium channel placement at nodes of Ranvier.Control of the cell surface density and localization of voltagegated sodium channels are critical aspects of neuronal function. This is especially important at nodes of Ranvier of myelinated axons where high densities of sodium channels are needed for rapid and reliable saltatory conduction (1, 2). Sodium channels from mammalian brain are heterotrimeric structures composed of a central pore forming ␣ subunit and two auxiliary subunits, ␤1 and ␤2 (3, 4). Though the ␤ subunits do not form the ion conducting pore, they play critical roles in channel gating, voltage dependence of activation and inactivation, and channel plasma membrane expression levels (5-8). ␤1 and ␤2 contain Ig-like extracellular domains and are members of the V-set of the Ig superfamily, which includes many cell adhesion molecules (CAMs) 1 (9). ␤1 and ␤2 have recently been shown to interact with the extracellular matrix proteins tenascin-C and tenascin-R (10, 11). Transfected cells expressing ␤1 or ␤2 subunits initially bind to and then are repelled from a tenascin-R substrate (11). Purified sodium channels or the bacterially expressed ␤2 subunit extracellular domain bind tenascin-C and tenascin-R in an enzyme-linked immunosorbent type biochemical assay (10). These results suggested that sodium channel ␤ subunits function as CAMs. CAMs of the L1 family also bind directly to cytoskeletal elements such as ankyrin (12,13,(15)(16)(17). Sodium channels have been shown to colocalize with ankyrin G and spectrin at axon initial segments and nodes of Ranvier, and there is some evidence to show that sodium channels bind ankyrin directly in vitro (18 -22). However, it is not known to which sodium channel subunit ankyrin binds. Because ␤1 and ␤2 are structurally and functionally homologous ...

Sodium channel Scn1b null mice exhibit prolonged QT and RR intervals

“…Cytoskeletal disruption in cardiac myocytes induces sodium channels to exhibit increased I NaP [43]. This is also consistent with deletion of β1, a cell adhesion molecule that links sodium channels to the cytoskeleton via ankyrin [9].…”

“…β subunits are multi-functional molecules that regulate channel cell-surface expression levels and modulate channel function, affecting channel kinetics and voltage-dependence in vitro [8]. β subunits also function in vitro as homophilic and/or heterophilic cell adhesion molecules that recruit cytoskeletal ankyrin following homophilic cell adhesion [9][10][11].…”

Heterophilic Interactions of Sodium Channel β1 Subunits with Axonal and Glial Cell Adhesion Molecules

“…Antibodies and Constructs-Rabbit polyclonal antisera to sodium channel ␤1 and ␤2 subunits were described previously (19,22,24). A polyclonal anti-␤3 antibody was a gift from Dr. William A. Catterall (University of Washington, Seattle, WA) and has been described previously (20).…”

“…␤1 and ␤2 colocalize with sodium channel ␣ subunits at nodes of Ranvier, and ␤1 interacts with contactin and with Nf186 in vitro (9,17,19,20). Nf186, Nr-CAM, ␤1, and ␤2 each interact in vitro with a key cytoskeletal anchoring protein, ankyrin G , that is also localized to nodes of Ranvier (21)(22)(23). ␤1-or ␤2-mediated homophilic cell-adhesive interactions result in ankyrin recruitment in Drosophila S2 cells, and the interaction of sodium channel ␣ subunits with ankyrin G is greatly enhanced in the presence of ␤1 subunits in vitro (22,24).…”

“…Nf186, Nr-CAM, ␤1, and ␤2 each interact in vitro with a key cytoskeletal anchoring protein, ankyrin G , that is also localized to nodes of Ranvier (21)(22)(23). ␤1-or ␤2-mediated homophilic cell-adhesive interactions result in ankyrin recruitment in Drosophila S2 cells, and the interaction of sodium channel ␣ subunits with ankyrin G is greatly enhanced in the presence of ␤1 subunits in vitro (22,24). We propose that the sodium channel signaling complex at nodes of Ranvier includes the CAMs contactin, NrCAM, and Nf186 as well as ankyrin G and that sodium channels interact with these proteins via ␤1 subunits.…”