The Sodium Channel Accessory Subunit Navb1 Associates With Brain Kv4.2 And Modulates The Functional Expression Of Kv4.2-encoded Channels

Abstract

Somatodendritic A-type (IA) voltage-gated K+ (Kv) channels are key regulators of neuronal excitability, functioning to modulate the back-propagation of action potentials and the frequency of repetitive firing. Although progress has been made in identifying components of Kv4-encoded IA channels, increasing evidence suggests that additional regulators are involved in modulating the functional expression of these channels. To identify novel components of native Kv4.2-encoded IA channel complexes, an anti-Kv4.2 antibody was used to immunoprecipitate Kv4.2 from mouse brain and co-purifying proteins were identified using a mass spectrometry (MS)-based proteomic approach. In addition to the Kv4 pore-forming alpha subunits (Kv4.2, Kv4.3, Kv4.1) and the previously described accessory subunits KChIP1-4 and DPP6/DPP10, eight tandem MS spectra identifying the Navb1 peptide YENEVLQLEEDERFEGR were acquired from Kv4.2 immunoprecipitates from wild-type brain lysates, but were absent from brains of animals (Kv4.2-/-) lacking Kv4.2. Although Navb1 has previously been reported to regulate the functional expression of voltage-gated Na+ channels in heterologous expression systems, these MS-based results suggest the interesting possibility that Navb1 might also function in the regulation of Kv4.2-encoded IA channels. To explore this possibility, the ability of Navb1 to modulate the functional expression of Kv4.2 channels was investigated in HEK-293 cells. Biochemical experiments confirmed that Navb1 co-immunoprecipitates with Kv4.2, but not with KChIP2. Whole-cell voltage-clamp recordings revealed that co-expression of Navb1 significantly increases Kv4.2-encoded current densities without measurably affecting the time- and/or voltage-dependent properties of the currents. Western blot and cell surface biotinylation experiments revealed that Navb1 increases the total and the cell surface expression of Kv4.2 proteins. Taken together, these results suggest that Navb1 associates with Kv4.2, increases the stability of the Kv4.2 protein and the cell surface expression of functional Kv4.2-encoded channels.