title: Drosophila odorant receptors are novel seven transmembrane domain proteins that can signal independently of heterotrimeric G proteins.
abstract: Olfaction in Drosophila is mediated by a large family of membrane-bound odorant receptor proteins (Ors). In heterologous cells, we investigated whether the structural features and signalling mechanisms of ligand-binding Drosophila Ors are consistent with them being G protein-coupled receptors (GPCRs). The detailed membrane topology of Or22a was determined by inserting epitope tags into the termini and predicted loop regions. Immunocytochemistry experiments in Drosophila S2 cells imply that Or22a has seven transmembrane domains but that its membrane topology is opposite to that of GPCRs, with a cytoplasmic N-terminus and extracellular C-terminus. To investigate Or signalling mechanisms, we expressed Or43b in Sf9 and HEK293 cells, and show that inhibitors of heterotrimeric G proteins (GDP-beta-S), adenylate cyclase (SQ22536), guanylyl cyclase (ODQ), cyclic nucleotide phosphodiesterases (IBMX) and phospholipase C (U73122) have negligible impact on Or43b responses. Whole cell patching of Or43b/Or83b-transfected HEK293 cells revealed the opening of plasma membrane cation channels on addition of ligand. The response was blocked by lanthanum and by 2-APB, but not by Ruthenium red or SKF96365. Based on these data, we conclude that Drosophila Ors comprise a novel family of seven transmembrane receptors that in HEK293 cells signal by opening cation channels, through a mechanism that is largely independent of G proteins.
authors.affiliation: School of Biological Sciences, Monash University, Melbourne, Vic. 3800, Australia.
authors.forename: Renee
authors.initials: R
authors.lastname: Smart
authors.forename: Aidan
authors.initials: A
authors.lastname: Kiely
authors.forename: Morgan
authors.initials: M
authors.lastname: Beale
authors.forename: Ernesto
authors.initials: E
authors.lastname: Vargas
authors.forename: Colm
authors.initials: C
authors.lastname: Carraher
authors.forename: Andrew V
authors.initials: AV
authors.lastname: Kralicek
authors.forename: David L
authors.initials: DL
authors.lastname: Christie
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.forename: Richard D
authors.initials: RD
authors.lastname: Newcomb
authors.forename: Coral G
authors.initials: CG
authors.lastname: Warr
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D004330:Drosophila / Q000737:chemistry / Q000378:metabolism*; D029721:Drosophila Proteins / Q000737:chemistry / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D007473:Ion Channels / Q000378:metabolism; D017434:Protein Structure, Tertiary; D018035:Receptors, Odorant / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D029721:Drosophila Proteins; D007473:Ion Channels; C496847:OR22a protein, Drosophila; C432947:OR43b protein, Drosophila; D018035:Receptors, Odorant; D019204:GTP-Binding Proteins