Functional analysis of a Drosophila melanogaster olfactory receptor expressed in Sf9 cells.
Olfactory receptors (ORs) are seven transmembrane proteins that are responsible for the transduction of volatiles into neuronal signals. Their low sequence homology means that the prediction of ligands for ORs based on extrapolation from empirical data of other ORs is difficult, so an experimental approach must be used. Here, we report a functional assay for insect ORs using calcium-imaging in Sf9 cells. We find that the interaction of the odorant, ethyl butyrate, with the Drosophila melanogaster olfactory receptor Or22a is both dose-dependent and highly sensitive, with Or22a responding to ethyl butyrate with an EC(50) of (1.58+/-0.82)x10(-11)M. This degree of sensitivity does not require the addition of odorant binding proteins or downstream signal transduction elements. Furthermore, we demonstrate that Or22a expressed in Sf9 cells has a similar response profile to a range of odorants previously tested in vivo. This functional assay system will provide a useful tool for the de-orphaning of ORs from a wide range of insect species that are yet to have ligands assigned, and will help provide insight into OR specificity and mechanism of activation.
Kiely
Aidan
A
The School of Biological Sciences, The University of Auckland, Auckland, New Zealand.
Authier
Astrid
A
Kralicek
Andrew V
AV
Warr
Coral G
CG
Newcomb
Richard D
RD
D000818:Animals; D002087:Butyrates; D002460:Cell Line; D029721:Drosophila Proteins / Q000235:genetics* / Q000502:physiology*; D004331:Drosophila melanogaster / Q000502:physiology*; D004594:Electrophysiology / Q000379:methods*; D009812:Odorants; D018035:Receptors, Odorant / Q000235:genetics* / Q000502:physiology*; D012903:Smell / Q000502:physiology*; D018411:Spodoptera; D014162:Transfection
D002087:Butyrates; D029721:Drosophila Proteins; C496847:OR22a protein, Drosophila; D018035:Receptors, Odorant; C045572:ethyl butyrate