Functional characterization of two human olfactory receptors expressed in the baculovirus Sf9 insect cell system.
Olfactory receptors (ORs) are the largest member of the G-protein-coupled receptors which mediate early olfactory perception in discriminating among thousands of odorant molecules. Assigning odorous ligands to ORs is a prerequisite to gaining an understanding of the mechanisms of odorant recognition. The functional expression of ORs represents a critical step in addressing this issue. Due to limitations in heterologous expression, very few mammal ORs have been characterized, and so far only one is from human origin. Consequently, OR function still remains poorly understood, especially in humans, whose genome encodes a restricted chemosensory repertoire compared with most mammal species. In this study, we have designed cassette baculovirus vectors to coexpress human OR 17-209 or OR 17-210 with either G(alpha olf) or G(alpha16) proteins in Sf9 cells. Each OR was found to be expressed at the cell surface and colocalized with both G(alpha) proteins. Using Ca2+ imaging, we showed that OR 17-209 and OR 17-210 proteins are activated by esters and ketones respectively. Odorant-induced calcium response was increased when ORs were coexpressed with G(alpha16) protein, whereas coexpression with G(alpha olf) abolished calcium signaling. This strategy has been found to overcome most of the limitations encountered when expressing an OR protein and has permitted odorant screening of functional ORs. Our approach could thus be of interest for further expression and ligand assignment of other orphan receptor proteins.
Matarazzo
Valéry
V
UMR 6149 et GDR 2590 CNRS et Université de Provence, IFR du Cerveau, 31 Chemin J. Aiguier, F-13402 Marseille Cedex 20, France.
Clot-Faybesse
Olivier
O
Marcet
Brice
B
Guiraudie-Capraz
Gaëlle
G
Atanasova
Boriana
B
Devauchelle
Gérard
G
Cerutti
Martine
M
Etiévant
Patrick
P
Ronin
Catherine
C
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D044385:GTP-Binding Protein alpha Subunits / Q000235:genetics / Q000378:metabolism*; D023281:Genomics; D006801:Humans; D007313:Insecta / Q000166:cytology; D018613:Microscopy, Confocal; D009812:Odorants; D018034:Olfactory Receptor Neurons / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism
D044385:GTP-Binding Protein alpha Subunits; D043562:Receptors, G-Protein-Coupled; C084618:olfactory G protein subunit alpha olf; D002118:Calcium