TY - JOUR
T1 - Odorant response assays for a heterologously expressed olfactory receptor
AU - Katada, Sayako
AU - Nakagawa, Takao
AU - Kataoka, Hiroshi
AU - Touhara, Kazushige
N1 - Funding Information:
We thank Dr. D.W. Saffen for zif268 promoter, Dr. E. Okuda-Ashitaka for CFTR cDNA, Dr. H. Sekine for PC12h cells, Dr. T. Okuda for the modified pSPUTK, Dr. M. Tanaka for assisting vector constructions, and Dr. M. Omura for critical reading of the manuscript. This work was supported in part by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan. K.T. is a recipient of grants from Uehara Memorial Foundation, Kato Memorial Bioscience Foundation, The Naito Foundation, and The Mochida Memorial Foundation.
Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 2003/6/13
Y1 - 2003/6/13
N2 - Odorant responsiveness of a mouse olfactory receptor, mOR-EG, was investigated in various heterologous cells using a variety of detection methods. Odorant-induced Ca2+ response was observed in HEK293 cells that coexpressed mOR-EG and the promiscuous G protein, Gα15. Without Gα15, a robust increase in cAMP level was observed upon odorant-stimulation in various mammalian cells. A luciferase reporter gene assay using zif268 promoter was adopted to amplify the cAMP signals. In Xenopus laevis oocytes, odorant-stimulated currents were recorded when mOR-EG cRNA was co-injected with either Gα15 or cAMP-dependent channel. These results suggest that odorant responsiveness can be monitored via a signaling pathway mediated by endogenous Gαs or transfected Gα15 in heterologous cell systems. Various functional assays for a heterologously expressed olfactory receptor reported in this study, are potentially useful for high-throughput ligand screening and functional analyses of hundreds of olfactory receptors.
AB - Odorant responsiveness of a mouse olfactory receptor, mOR-EG, was investigated in various heterologous cells using a variety of detection methods. Odorant-induced Ca2+ response was observed in HEK293 cells that coexpressed mOR-EG and the promiscuous G protein, Gα15. Without Gα15, a robust increase in cAMP level was observed upon odorant-stimulation in various mammalian cells. A luciferase reporter gene assay using zif268 promoter was adopted to amplify the cAMP signals. In Xenopus laevis oocytes, odorant-stimulated currents were recorded when mOR-EG cRNA was co-injected with either Gα15 or cAMP-dependent channel. These results suggest that odorant responsiveness can be monitored via a signaling pathway mediated by endogenous Gαs or transfected Gα15 in heterologous cell systems. Various functional assays for a heterologously expressed olfactory receptor reported in this study, are potentially useful for high-throughput ligand screening and functional analyses of hundreds of olfactory receptors.
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U2 - 10.1016/S0006-291X(03)00863-5
DO - 10.1016/S0006-291X(03)00863-5
M3 - Article
C2 - 12767924
AN - SCOPUS:0038242292
VL - 305
SP - 964
EP - 969
JO - Biochemical and Biophysical Research Communications
JF - Biochemical and Biophysical Research Communications
SN - 0006-291X
IS - 4
ER -