Synergism of accessory factors in functional expression of mammalian odorant receptors.
The discovery of odorant receptors led to endeavors in matching them with their cognate ligands. Although it has been challenging to functionally express odorant receptors in heterologous cells, previous studies have linked efficient odorant receptor expression with N-terminal modifications and accessory proteins, including the receptor-transporting proteins (RTPs) and Ric8b. Here we have shown that a shorter form of RTP1, RTP1S, supports robust cell-surface and functional expression of representative odorant receptors. Using a combination of accessory proteins, including RTP1S, Ric8b, and G(alphaolf), a diverse set of untagged odorant receptors were successfully expressed heterologously due to the synergistic effects among the various accessory proteins. Furthermore, the addition of an N-terminal rhodopsin tag to the odorant receptors, along with the same set of accessory proteins, exhibits an additional level of synergism, inducing enhanced odorant receptor responses to odorants and thus defining a more efficient heterologous expression system. We then showed that the presence or absence of different N-terminal tags has little effect on the ligand specificity of odorant receptors, although the amount of receptor expressed can play a role in the ligand response profile. The accuracy of the odorant receptor heterologous expression system involving tagged odorant receptors and various accessory proteins promises success in high throughput de-orphaning of mammalian odorant receptors.
Zhuang
Hanyi
H
Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Matsunami
Hiroaki
H
D002460:Cell Line; D044385:GTP-Binding Protein alpha Subunits / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D026901:Membrane Transport Proteins / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D018035:Receptors, Odorant / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
D044385:GTP-Binding Protein alpha Subunits; D026901:Membrane Transport Proteins; D009687:Nuclear Proteins; C495316:RTP1 protein, human; D018035:Receptors, Odorant; D011993:Recombinant Fusion Proteins; C084618:olfactory G protein subunit alpha olf