Analysis of the native quaternary structure of vanilloid receptor 1.
Vanilloid receptor subtype 1 (VR1) is a ligand-gated channel that can be activated by capsaicin and other vanilloids as well as by protons and heat. In the present study, we have analyzed the oligomeric state of VR1. Co-immunoprecipitation of differently tagged VR1 molecules indicated that VR1 can form oligomers. Using two different heterologous VR1 expression systems as well as endogenous VR1 expressed in dorsal root ganglion cells, we analyzed oligomer formation using perfluoro-octanoic acid polyacrylamide gel electrophoresis. Results were confirmed both with chemical cross-linking agents as well as through endogenous cross-linking mediated by transglutaminase. Our results clearly show that VR1 forms multimers in each of the expression systems with a homotetramer as a predominant form. The oligomeric structure of VR1 may contribute to the complexity of VR1 pharmacology. Finally, differences in glycosylation between the systems were observed, indicating the need for caution in the use of the heterologous expression systems for analysis of VR1 properties.
Kedei
N
N
Laboratory of Cellular Carcinogenesis and Tumor Promotion, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Szabo
T
T
Lile
J D
JD
Treanor
J J
JJ
Olah
Z
Z
Iadarola
M J
MJ
Blumberg
P M
PM
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D019556:COS Cells; D002210:Caprylates / Q000737:chemistry; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005466:Fluorocarbons / Q000737:chemistry; D049452:Green Fluorescent Proteins; D008024:Ligands; D008164:Luminescent Proteins / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D020836:Protein Structure, Quaternary; D011522:Protons; D011955:Receptors, Drug / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D050916:TRPV Cation Channels; D014162:Transfection; D011503:Transglutaminases / Q000378:metabolism
D002210:Caprylates; D003432:Cross-Linking Reagents; D005466:Fluorocarbons; D008024:Ligands; D008164:Luminescent Proteins; D011522:Protons; D011955:Receptors, Drug; D011993:Recombinant Fusion Proteins; D050916:TRPV Cation Channels; C494484:TRPV1 receptor; D049452:Green Fluorescent Proteins; C023036:perfluorooctanoic acid; D011503:Transglutaminases