The Transient Receptor Potential Vanilloid 1 (TRPV1, vanilloid receptor 1) ion channel plays a key role in the perception of thermal and inflammatory pain, however, its molecular environment in dorsal root ganglia (DRG) is largely unexplored. Utilizing a panel of sequence-directed antibodies against TRPV1 protein and mouse DRG membranes, the channel complex from mouse DRG was detergent-solubilized, isolated by immunoprecipitation and subsequently analyzed by mass spectrometry. A number of potential TRPV1 interaction partners were identified, among them cytoskeletal proteins, signal transduction molecules, and established ion channel subunits. Based on stringent specificity criteria, the voltage-gated K(+) channel beta 2 subunit (Kvβ2), an accessory subunit of voltage-gated K(+) channels, was identified of being associated with native TRPV1 channels. Reverse co-immunoprecipitation and antibody co-staining experiments confirmed TRPV1/Kvβ2 association. Biotinylation assays in the presence of Kvβ2 demonstrated increased cell surface expression levels of TRPV1, while patch-clamp experiments resulted in a significant increase of TRPV1 sensitivity to capsaicin. Our work shows, for the first time, the association of a Kvβ subunit with TRPV1 channels, and suggests that such interaction may play a role in TRPV1 channel trafficking to the plasma membrane.
Keywords: 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate; 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; 4′,6-diamidino-2-phenylindole; A kinase anchor protein; AKAP; ATP; Accessory subunit; BSA; CGRP; CHAPS; CHO; CMC; Chinese hamster ovary; DAPI; DNA; DRG; DTT; Dorsal root ganglia; EDTA; EGTA; GAPDH; HEPES; Icaps; IgG; Kvβ2; PBS; PMSF; SDS; Signaling complex; TRPV1; Transient Receptor Potential Vanilloid 1; WT mouse; adenosine tris-phosphate; bovine serum albumin; cDNA; calcitonin gene-related peptide; capsaicin-activated current; complementary deoxyribonucleic acid; critical micelle concentration; deoxyribonucleic acid; di-tiothreitol; dorsal root ganglia; ethylene glycol tetraacetic acid; ethylenediaminetetraacetic acid; glyceraldehyde-3-phosphate dehydrogenase; immunoglobulin G; phenylmethylsulfonyl fluoride; phosphate buffer saline; sodium dodecyl sulfate; voltage-gated K(+) channel subunit beta 2; wild type mouse.
© 2013.