Functional and Nonfunctional Forms of CquiOR91, an Odorant Selectivity Subunit of Culex quinquefasciatus.
In Culex quinquefasciatus, CquiOR91 is the ortholog of 2 larvae-specific odorant receptors (ORs) from Anopheles gambiae (Agam\Or40, previously shown to respond to several odorant ligands including the broad-spectrum repellent N,N-diethyl-3-methylbenzamide, DEET) and Aedes aegypti (Aaeg\Or40). When we cloned full-length CquiOR91 from a Culex quinquefasciatus larval head RNA sample, we found 2 alleles of this OR, differing at 9 residues. Functional analysis using the Xenopus oocyte expression system and 2-electrode voltage clamp electrophysiology revealed one allele (CquiOR91.1) to be nonfunctional, whereas the other allele (CquiOR91.2) was functional. Receptors formed by CquiOR91.2 and Cqui\Orco responded to (-)-fenchone, (+)-fenchone, and DEET, similar to what has been reported for Agam\Or40. We also identified 5 novel odorant ligands for the CquiOR91.2 + Cqui\Orco receptor: 2-isobutylthiazole, veratrole, eucalyptol, d-camphor, and safranal, with safranal being the most potent. To explore possible reasons for the lack of function for CquiOR91.1, we generated a series of mutant CquiOR91.2 subunits, in which the residue at each of the 9 polymorphic residue positions was changed from what occurs in CquiOR91.2 to what occurs in CquiOR91.1. Eight of the 9 mutant versions of CquiOR91.2 formed functional receptors, responding to (-)-fenchone. Only the CquiOR91.2 Y183C mutant was nonfunctional. The reverse mutation (C183Y) conferred function on CquiOR91.1 , which became responsive to (-)-fenchone and safranal. These results indicate that the "defect" in CquiOR91.1 that prevents function is the cysteine at position 183.
Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, R-189, PO Box 016189, Miami, FL 33101, USA and.|Present address: Department of Internal Medicine, The Ohio State University Wexner Medical Center, Columbus, OH 43210, USA.
David T
DT
Hughes
Department of Molecular and Cellular Biology, University of California, 1 Shields Avenue, Davis, CA 95616, USA.|Present address: School of Life Sciences, Keele University, Keele, Staffordshire ST5 5BG, UK.
Julien
J
Pelletier
Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, R-189, PO Box 016189, Miami, FL 33101, USA and.
Suhaila
S
Rahman
Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, R-189, PO Box 016189, Miami, FL 33101, USA and.|Present address: Leidos Biomedical Research, Inc., Rockville, MD 20892, USA.
Sisi
S
Chen
Department of Molecular and Cellular Biology, University of California, 1 Shields Avenue, Davis, CA 95616, USA.
Walter S
WS
Leal
Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, R-189, PO Box 016189, Miami, FL 33101, USA and.
Charles W
CW
Luetje
D000483:Alleles; D000818:Animals; D001892:Camphanes; D009033:Culicidae / Q000737:chemistry*; D003671:DEET / Q000378:metabolism; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D008024:Ligands; D009154:Mutation; D009636:Norbornanes / Q000378:metabolism; D021122:Protein Subunits; D018035:Receptors, Odorant / Q000235:genetics* / Q000378:metabolism
D001892:Camphanes; D019476:Insect Proteins; D008024:Ligands; D009636:Norbornanes; D021122:Protein Subunits; D018035:Receptors, Odorant; C027327:fenchone; D003671:DEET
Xenopus oocytes; electrophysiology; insect; olfaction; polymorphism; receptor